Chemical Processes With Participation of Biological and Related Compounds

Chemical Processes
with Participation of Biological

and Related Compounds
Chemical Processes
with Participation of Biological
and Related Compounds
Biophysical and -Chemical Aspects of Porphyrins,

Pigments, Drugs, Biodegradable Polymers
and Nanofibers
Edited by
T.N. Lomova
G.E. Zaikov
LEIDEN • BOSTON
2008
This book is printed on acid-free paper.
ISBN 978 90 04 16210 5
Copyright 2008 by Koninklijke Brill NV, Leiden, The Netherlands.

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printed in the netherlands

Contents
Preface xi
Foreword 1
Chapter 1
Synthetic and Natural Bacteriochlorins: Synthesis, Properties and
Applications 5
M.A. Grin and A.F. Mironov
Introduction 5
1 Synthetic bacteriochlorins 6
2 Natural bacteriochlorins and their chemical modifications 14
3 Amphiphilic and water-soluble derivatives of bacteriochlorins 30
4 Conclusion 41
References 41
Chapter 2
meso-Phenylporphyrins as Synthetic Models of Natural Porphyrins:
Synthesis and Modification 45
A.S. Semeykin, S.A. Syrbu and O.I. Koifman
Introduction 45
1 Synthesis of meso(5,10,15,20)-tetrasubstituted porphyrins 46
1.1 Synthesis of meso-tetra-β-octasubstituted porphyrins 55
1.2 Synthesis of nonsymmetric meso-tetrasubstituted porphyrins 58
2 meso(5,15)-Disubstituted porphyrins 66
3 Synthesis of meso(5)-Phenyl-b-octaalkylporphyrins 74
4 Reactions of introduction and modification of substituents in phenyl
rings of meso-phenyl-substituted porphyrins 77
4.1 Introduction of substituents into phenyl rings of meso-phenylporphyrins 77
4.2 Modification of functional groups in phenyl rings of
meso-phenylporphyrins 79
4.2.1 meso-Oxyphenylporphyrins and their modification 80
4.2.2 meso-Aminoporphyrins and their modification 82
References 84
vi CONTENTS
Chapter 3
The Mechanism of Catalytic Action of the Coordination Centres
of Catalase Synthetic Models 93
T.N. Lomova, M.E. Klyueva and M.V. Klyuev
Introduction 93
1 Substituted copper(II) porphyrins as catalysts of the hydrogen peroxide
disproportionation reaction 95
2 Kinetic regularities and mechanisms of peroxide decomposition reactions
in the presence of acido complexes of highly substituted manganese
porphyrins 96
2.1 Kinetics and reaction mechanism of oxidation of manganese(III)
porphyrins by hydrogen peroxide 98
2.2 Kinetics of peroxide disproportionation in the presence of
manganese(III) porphyrins 108
3 Conclusion 114
References 115
Chapter 4
Complexation of Porphyrins with Ions and Organic Molecules 117
N.Zh. Mamardashvili, V.V. Borovkov, G.M. Mamardashvili,
Y. Inoue and O.I. Koifman
Introduction 117
1 Complexation of porphyrins with ions 118
2 Complexation of porphyrins with organic molecules:
the thermodynamic aspect 125
3 Complexation of porphyrins with organic molecules:
the chirality aspect 144
3.1 Host–guest systems based on monomeric porphyrins 144
3.2 Host–guest systems based on dimeric and oligomeric porphyrins 151
4 Conclusion 162
References 163
Chapter 5
Chemical Activation of Porphyrins in Coordination Core Reactions 169
D.B. Berezin and B.D. Berezin
Introduction 170
1 Porphyrin ligands with localized and delocalized NH bonds 171
1.1 Factors causing the delocalization of NH bonds in porphyrin
molecules 172
1.1.1 Spatial structure and polarization of the porphyrin molecule 172
1.1.2 NH activation in the course of porphyrin–solution
component interaction and porphyrin–solid phase interaction 181
1.2 Interaction of organic solvents and porphyrins with delocalized-type
bonds 185
1.2.1 Acid-base interactions 185
1.2.2 Tautomeric processes 190
1.3 Quantitative assessment of the state of NH bonds in porphyrin
molecules 199
CONTENTS vii
1.3.1 Spectral criterion 199

1.3.2 Kinetic criterion 200
1.3.3 Quantum chemical criterion 203
1.3.4 Insufficiency of absorption spectrum analysis of porphyrins 203
1.4 Nonplanar structure of the macrocycle and chemical NH activity
in its coordination core 204
2 Reactivity of the coordination core in molecules of porphyrin ligands 208
2.1 Complexation reactions 208
2.2 Proton ionization of NH bonds in H2P molecules 210
2.3 Nucleophilic substitution reactions in the coordination core 211
3 Biosignificance of the phenomenon of NH activation 211
References 212
Chapter 6
Synthesis, Structure Peculiarities and Biological Properties of
Macroheterocyclic Compounds 219
M.K. Islyaikin, E.A. Danilova, Yu.V. Romanenko, O.G. Khelevina
and T.N. Lomova
Introduction 219
1 Synthesis, structure and properties of the initial compounds 221
2 ABBB-type macroheterocyclic compounds 228
3 Azolophthalocyanines 230
4 State of triazoleporphyrazines in proton-donor media 236
5 ABAB-type macroheterocyclic compounds 244
6 ABABAB-type macroheterocyclic compounds 249
7 Coordination properties 253
8 Studies of the practically valuable properties of macroheterocyclic
compounds and their metal complexes 262
8.1 Biological properties 262
References 266
Chapter 7
The Photochemical Aspect of Reactions of Flavonols with Molecular
Oxygen 271
E.A. Venedictov
Introduction 271
1 Structure of flavonols 272
2 Spectral luminescent properties of flavonols 276
3 Photoproduction of 1O2 (1∆g) 278
4 1O2 Reactions 279
5 Photochemical properties of coordination compounds of quercetine 284
6 Conclusion 288
References 288
Chapter 8
Solvation of Drugs as a Key for Understanding Partitioning and
Passive Transport Exemplified by NSAIDs 291
German L. Perlovich and Annette Bauer-Brandl
Introduction 292
viii CONTENTS
1 Partitioning and diffusion 292

1.1 The partitioning (distribution) process 293
1.2 Influence of the solution pH on the partition/distribution coefficients 294
1.3 Diffusion of drugs 294
2 Solvation of drugs: relevance and theoretical approaches 296
2.1 The main definitions 296
2.2 Models describing the solvation of molecules 297
3 Experimental methods to measure solvation characteristics and choice
of subjects 299
3.1 Sublimation experiment 299
3.2 Method of isothermal saturated solubility 301
3.3 Isothermal calorimetry 301
3.4 Choice of drugs 301
4 Crystal structures of NSAIDs 302
4.1 Description of hydrogen bond networks topology by graph set
assignment 302
4.2 Analysis of packing architectures of NSAIDs crystal lattices 303
5 Thermochemical and thermodynamic properties of NSAIDs 307
5.1 Thermodynamic characteristics of sublimation of NSAIDs 307
5.1.1 Differences of racemate and enantiomer ibuprofen
crystal lattices 309
5.2 Thermochemical characteristics of NSAIDs 312
6 The difference between partitioning and distribution of NSAIDs
from the thermodynamic point of view 314
6.1 Solvation characteristics of dissociated and non-dissociated
(+)- and (±)-IBP 316
6.2 Solvation characteristics of dissociated and non-dissociated
forms of the other NSAIDs 317
6.3 Solvation characteristics of transfer process of dissociated and
non-dissociated molecules from buffer to n-octanol 318
7 Correlation between biopharmaceutically relevant parameters and
solvation characteristics 322
References 324
Chapter 9
Biodamage of Materials: Adhesion of Microorganisms on
the Surface of Materials 327
K.Z. Gumargalieva, I.G. Kalinina, S.A. Semenov and
G.E. Zaikov
Introduction 328
1 Results and discussion 328
References 339
Chapter 10
Controlled Release of Aseptic Drug from Poly(3-hydroxybutyrate)
Films: A Combination of Diffusion and Zero-order Kinetics 341
R.Y. Kosenko, Y.N. Pankova, A.L. Iordanskii, A.P. Bonartsev, and
G.E. Zaikov
Introduction 342
CONTENTS ix
1 Experimental 342
3 Conclusion 347
4 Acknowledgement 347
References 347
Chapter 11
Transport of Water as a Structurally Sensitive Process
Characterizing the Morphology of Biodegradable Polymer Systems 349
A.L. Iordanskii, Yu.N. Pankova, R.Yu. Kosenko, A.A. Ol’khov and
G.E. Zaikov
Introduction 350
1 Experimental 350
2 Hydrophobization of poly-(3-hydroxybutyrate) 351
3 Hydrophilization of poly-(3-hydroxybutyrate) 354
References 359
Chapter 12
A Novel Technique for Measurement of Electrospun Nanofiber 361
M. Ziabari, V. Mottaghitalab and A.K. Haghi
Introduction 361
1 Methodology 363
1.1 Simulation of electrospun web 363
1.2 Fiber diameter measurement 363
1.2.1 Manual method 364
1.2.2 Distance transform 364
1.2.3 Direct tracking 365
1.3 Real webs treatment 366
2 Experimental 367
4 Conclusion 373
References 374
Chapter 13
Image Analysis of Pore Size Distribution in Electrospun Nanofiber
Webs: New Trends and Developments 375
M. Ziabari, V. Mottaghitalab and A.K. Haghi
Introduction 375
1 Methodology 376
1.1 Sieving methods 378
1.2 Mercury porosimetry 378
1.3 Flow porosimetry (bubble point method) 379
1.4 Image analysis 379
1.4.1 Real webs 380
1.4.2 Simulated webs 381
2 Experimental 382
4 Conclusion 388
References 389
x CONTENTS
Chapter 14
Electrospun Biodegradable and Biocompatible Natural Nanofibers:
A Detailed Review 391
A.K. Haghi and R.K. Haghi
1 Introduction 391
1.1 Electrospinning setup 392
2 Effect of systematic parameters on electrospun nanofibers 393
2.1 Solution properties 393
2.1.1 Viscosity 393
2.1.2 Solution concentration 393
2.1.3 Molecular weight 395
2.1.4 Surface tension 396
2.1.5 Number of entanglements 396
2.1.6 Solution conductivity 396
2.1.7 Effect of salt addition 397
2.1.8 Solvent 399
2.2 Processing condition 399
2.2.1 Applied voltage 399
2.2.2 Feed rate 402
2.2.3 Distance of needle tip to collector 403
3 Theory and modeling 405
4 Natural fibers 407
5 Electrospinning of silk fibers 408
5.1 Introduction 408
5.2 Crystal structure of silk (fibroin) at various stages of electrospinning 408
5.3 Spinning dope preparation for electrospinning 411
5.3.1 Degumming 411
5.3.2 Dissolving of fibroin 411
5.4 Electrospinning of silk fibroin 411
5.4.1 Effect of silk polymer concentration on fiber diameter 412
5.4.2 Effect of voltage and spinning distance on morphology and
diameter 414
5.5 Characterization 416
6 Electrospinning of cellulose and cellulose acetate 417
6.1 Electrospinning of CTA solution 418
7 Concluding remarks 419
References 420
Nomenclature 422
To learn anything without thinking is absolutely useless,
Thinking about something without analysing and
Studying the subject of thinking is dangerous
Confucius, 551–479 BC
Ancient China
Chemistry is a miracle, interest, delight,

The future and basis of well-being of people
Yury M. Luzhkov
Mayor of Moscow
November 12, 2003
Preface
Everything around us has been created by chemistry (glass, gas, medicines, food, metals,
polymers, etc.). There is no pathos of “chemical” enthusiasm in this definition. Chemistry
is the basis of everything, which life produces (microbes, plants, animals, human beings).
Any living body is a giant chemical reactor with millions of coordinated chemical reactions
proceeding in it. Molecular biology, molecular genetics and gene engineering, biotechnol-
ogy and intellect – all this is chemistry.
Nobelist Prof. N.N. Semenov and his pupils (Profs. M.N. Emanuel, V.N. Kondratyev,
V.I. Goldansky, A.L. Buchachenko, K.I. Zamoraev, Yu.B. Monakov) insisted on that. And
we fully agree with them.
There is a certain shift in the world of chemistry from studies of more or less simple
reactions to get, for example, sulfuric acid, ammonia, phenol and acetone to research into
complicated biochemical and biological processes. This is required for solving medical
problems, because medicine is our health and our good life. It is common knowledge that
it is better to be healthy and rich than to be poor and sick.
Chemistry is more than a science. It is a festivity. This volume presents the reviews of
chemists working in the field of biochemistry, biology and, in the final analysis, for medi-
cine and health.
We look forward to readers’ comments, will be grateful for them and will definitely
use them in our future research.
Prof. Gennady E. Zaikov
Moscow, Russia
Prof. Tatyana N. Lomova
Ivanovo, Russia
x INTRODUCTION
To propagate education is to extend prosperity.
I mean the general prosperity but not one’s private wealth.
With extension of prosperity, part of evil disappears.
Alfred Nobel, Sweden
Foreword
Research into the kinetics and thermodynamics of processes involving biologically active
substances and multistage mechanisms of their interactions in living systems is of primary
interest for modern science. Many problems arising in the course of the studies may be
solved by investigating the reactivity of BAS in model systems and synthetic analogs of nat-
urally occurring biomolecules with a complex and often unknown structure. The authors of
this book have been for many years engaged in studies of the reactivity of BAS in various
aggregate states and in relation to the molecular structures and supramolecular forms.
The book covers many aspects of the effects of relatively simple biomolecules as mod-
el enzymes, molecular receptors, photosensitizers, pharmacophores, and biopharmaceutical
agents. The quantitative characteristics of the transitions of cations, anions and small or-
ganic molecules, enzymic catalysis, and diffusion of molecules through biological mem-
branes are presented. The mechanisms of the processes are discussed. The biological
activity of the compounds studied is assessed.
Hydrated porphyrin forms and their unique properties are of great interest. Chapter 1
deals with naturally occurring bacteriochlorins, their properties, isolation and chemical
modifications. A separate part is concerned with synthetic bacteriochlorins; here, general
and particular methods of production of these compounds, their spectral and physicochem-
ical characteristics, are considered. Special consideration is given to the use of naturally
occurring and synthetic bacteriochlorins in the production of new-generation photosensi-
tizers for photomedicine.
Chapter 2 describes methods of synthesis of meso-mono-, di-, tri-, and tetraphenyl-
porphyrins along with schemes of addition and modification of substituents in them.
Approaches are shown to fine setting of the physicochemical properties of porphyrins
obtained.
Chapter 3 represents the results of research into the catalytic activity of manga-
nese(III)- and copper(II)-porphyrins alkyl- and phenyl-substituted in β- and meso-positions
in the reaction of decomposition of hydrogen peroxide in the DMFA–KOH–H 2O system.
The ion-molecular mechanism of the decomposition with kinetically significant stages of
2 Foreword
two-electron oxidation and subsequent partial reduction of metalporphyrin was determined

as well as acid–basic equilibria of peroxide. It is shown that the efficient catalysis of
decomposition of hydrogen peroxide is determined by the degree of binding of porphyrin
in a complex with metal, by the structure of the mixed coordination sphere, and by the
mutual influence of the ligands; the compounds under study behave as catalases in living
systems.
Investigation of the interactions between porphyrin molecules and protein surround-
ings in biological systems is an important research area in modern biochemistry. These
interactions may proceed in various ways, e.g., through the formation of covalent bonds,
by ionic association, as donor–acceptor interactions, or through hydrogen bonds between
separate covalent-bound fragments.
Chapter 4 gives a systemic description of complexation properties of porphyrins and
their ability to discern charged particles and small organic molecules.
Depending on the molecular structure and medium, NH bonds of the coordination
nucleus of porphyrins are considered as localized or partially localized ones. Their delocal-
ization may be caused by internal molecular effects, such as polarization of the molecule
by substituents or by specifically nonplanar conformations of the macrocycle during solva-
tion in solvents containing electron–donor components, by polymerization, sorption or
transition to the solid state. The activation of NH bonds in porphyrin molecules is associated
with drastic changes in the coordination nucleus reactivity.
Chapter 5 makes an analysis of the causes and pathways of activation of NH bonds
with regard to the classification of porphyrins by their capability for NH activating. Some
reliable quantitative criteria for determining the degree of NH activity were suggested for
porphyrins and their analogues. Activation of NH bonds may manifest itself not only in the
reactivity of porphyrin coordination nuclei but may lead to the reorganization of the π-chro-
mophore and reacting sites and to tautomeric processes. Products of this reorganization are
good models of supramolecular biological systems. Chemical activation of porphyrins is
considered separately as a potent tool of controlling the activity of molecules along with
biological aspects.
Chapter 6 reviews the latest achievements in the chemistry of macroheterocyclic com-
pounds – structural analogs of porphyrin and hexapyrins. The aromas of various macro- cy-
clic molecules and their fragments were investigated in accordance with the geometric (EN,
GEO, and HOMA) and magnetic (NICS) criteria based on experimental data and results of
DFT quantum-chemical calculations. The coordination and biological properties of macro-
heterocyclic compounds and their complexes with metal cations are considered.
Chapter 7 presents the results of studies on the kinetics of photochemical reactions of
biologically active pigments and related compounds with molecular oxygen.
Chapter 8 is devoted to the search of correlations of thermodynamic characteristics
(the Gibbs energy and enthalpy and entropy constituents of the Gibbs energy) of solvation
of molecules of drugs prepared by traditional experimental methods and their diffusive
properties and biopharmaceutically important properties.
Chapters 9–11 deal with biodamage of materials (adhesion of microorganisms on the
surface of materials), aspects of controlled release from polymer films and transport of
water as a structurally sensitive process that characterizes the morphology of biodegradable
polymer systems.
Chapters 12–14 discuss the development of an image analysis based method (direct
tracking) for measuring the diameter of electrospun fiber, consider new trends and devel-
opments in image analysis of pore size distribution and present a detailed review of different
Foreword 3
aspects of electrospinning biodegradable and biocompatible natural nanofibers.

This collection of papers dealing with the results of studies on various classes of bio-
logically active compounds will add to understanding the problems of their reactivity and
the nature of processes occurring in living objects with their participation.
Editors:
Prof. Gennady E. Zaikov
Institute of Biochemical Physics, Russian Academy of Sciences
Moscow, Russia; GEZaikov@yahoo.com
Prof. Tatyana N. Lomova
Institute of Solution Chemistry, Russian Academy of Sciences
Ivanovo, Russia; tnl@isc-ras.ru
4 Foreword
- нет ссылки 57 в списке лит.
-
1 Synthetic and Natural

Bacteriochlorins: Synthesis,
Properties and Applications
M.A. Grin and A.F. Mironov

Lomonosov Moscow State Academy of Fine Chemical
Technology, 86 Vernadsky Prospekt, Moscow, 119571,
Russia; email: httos.mitht@g23.relcom.ru
This chapter considers natural bacteriochlorins, their properties, isolation and

chemical modifications. Part of the discussion is given to synthetic bacteriochlor-
ins, with consideration of common and specific methods of producing these sub-
stances, their spectral and other physicochemical characteristics. Special attention
is paid to the use of natural and synthetic bacteriochlorins in developing new-
generation photosensitizers for photomedicine.
Introduction
In the recent decade, attention of scientists, working in the field of developing new photo-
sensitizers (PS) for photodynamic therapy (PDT) of cancer, has been focused on com-
pounds with intensive absorption in the range of 770 up to 850 nm. The use of PS with this
therapeutic window of absorption opens new possibilities for the diagnostics and treatment
of malignant neoplasms. The light with these wavelengths scatters weakly and, therefore,
can penetrate deeper into the tissue (Scheme 1) [1]. This is of special significance in pig-
mented tumors, for instance, melanoma. Besides, it is important that accessible and cheap
semiconductor lasers can be used for this range.
Such compounds include derivatives of synthetic and natural bacteriochlorins. It is
known that in porphyrin systems two peripheral double bonds in opposite pyrrole rings
(B and D) are cross-conjugated, and their presence is not required for aromaticity to be
preserved.
6 M.A. Grin and A.F. Mironov
Penetration into tissues, mm
nm
Porphyrins Chlorins Bacteriochlorins
Scheme 1 Dependence of the penetration of light into tissues on the wavelength.
In the reduction of one bond (dihydroporphyrins–chlorins) or both bonds (tetrahydro-

porphyrins–bacteriochlorins), aromaticity is preserved, and the change of symmetry leads
to a batochromic shift of the Q band. Figure 1 shows the real spectra of three compounds:
porphyrin, chlorin and bacteriochlorin, which have the same m-hydroxyphenyl substituents
in meso-positions of the macrocycle. Bacteriochlorins intensively absorb in the near IR re-
gion of the spectrum (λmax = 760–780 nm, ε = 4×104 –1×105 M –1 cm –1) [2] and, there-
fore, possess optimal properties for their use as photodynamic agents. Besides, they
generate active oxygen species (AOS) with a high quantum yield, which depends on the
nature of the central metal and peripheral substituents [3].
However, the macrocycle with two reduced double bonds is chemically unstable,
therefore, derivatives of bacteriochlorophyll a are apt to be oxidized to respective chlorins
and porphyrins. This fact largely restricts introduction of PS of bacteriochlorin series into
clinical practice. A high hydrophobicity and related low solubility in polar solvents also
complicate their use in medicine.
The main works on the chemical conversions of bacteriochlorophyll a (Bchl a) aim to
increase the chemical stability and to develop water-soluble forms of bacteriochlorin pho-
tosensitizers.
Works on the chemical modification of bacteriochlorophyll a are comparatively a few.
Major research in this field has been done at the Weizmann Institute of Science, Rehovot,
Israel and Photodynamic Therapy Center, Roswell Park Cancer Institute, Buffalo, USA.
Our group also does intensive research in this field, and major results will be presented in
this review.
There are two approaches to production of bacteriochlorins. The synthetic way in-
cludes the reduction of double bonds in pyrrole rings B and D in porphyrins, and the semi-
synthetic route is when bacteriochlorophylls isolated from natural sources are modified to
increase their stability, improve the spectral characteristics and solubility in polar solvents.
1 Synthetic Bacteriochlorins
There are two approaches to the reduction of porphyrins to chlorins and bacteriochlorins.
The first includes catalytic hydrogenation or treatment with metals in an alcohol medium
Synthetic and Natural Bacteriochlorins: Synthesis, Properties and Applications 7
nm
nm
nm
Figure 1 Electronic spectra of porphyrin (1), chlorin (2) and bacteriochlorin (3).
R R
NH N R R NH N R
R A
N HN N HN
R R
4
B
R R
R N N R N N R
A R
Zn Zn
N N N N
R R
5
A: p-MeC6H4SO2NHNH2, K2CO3/Py, t [N2H2]
B: Zn(OAc)2
Scheme 2 Reduction of porphyrins and their metal complexes with diimide.
(boiling with sodium in amyl alcohol). The other approach is related to the use of diimide
NH=NH, which is formed from p-tosylhydrazine, as a reducing agent. When porphyrins are
reduced as free bases, a mixture of respective chlorin and bacteriochlorin 4 is formed,
whereas the reduction products of the zinc complex of porphyrin are chlorin and isobacte-
riochlorin 5 (Scheme 2) [4].
This approach was realized by R. Bonnett et al. in the reduction of 5,10,15,20-
tetrakis(m-hydroxyphenyl)porphyrin (m-THPP) 1 to form respective chlorin (m-THPC) 2
and bacteriochlorin (m-THPBC) 3 [5].
Though the molar extinction coefficients and λmax of the long-wavelength absorption
band increase in the sequence porphyrin > chlorin > bacteriochlorin, the photophysical
properties of the reduced species, including the quantum yields of the triplet and singlet ox-
ygen, differ insignificantly [6]. The photodynamic efficiency in the given sequence increas-
es at each stage of reduction,. Thus, at a tumour photonecrosis depth of 5 mm the doses of
PS introduced decrease from 6.25 mmol/kg for porphyrin to 0.75 mmol/kg for chlorin and
0.39 mmol/kg for bacteriochlorin [5].
However, in in vivo experiments bacteriochlorin, possessing a high photodynamic ac-
tivity, proved to be much less stable as compared with the chlorin analogue (Foscan® ); its
considerable amount was oxidized in cells within 24 h.
An interesting approach to the synthesis of di- and tetrahydroporphyrin derivatives
was proposed by Callot et al. [7], who showed that, during the action of diazoacetic-acid
methyl ester on tetraphenylporphyrin (TPP) 6, carbene formed in the reaction attacks the
double bonds in the pyrrole rings B and D to form cycloaddition products chlorin 7 (λmax =
650 nm) and bacteriochlorin 8 (λmax = 720 nm) (Scheme 4).
OH
HO NH N
N HN OH
OH
OH
HO NH N 2
+ B C
N HN OH
OH
OH
1 HO NH N
N HN OH
OH
A: p-MeC6H4SO2NHNH2, K2CO3/Py, t [N2H2]

B: [N2H2]
C: ɨ-chloranil
Scheme 3 Reduction of 5,10,15,20-tetrakis(m-hydroxyphenyl)porphyrin with diimide.
Ph CO2Me CO 2Me
H Ph
Ph H
H H
H H
NH N NH N
NH N
A
Ph Ph
Ph Ph
+ Ph Ph
N HN N HN
N HN
H
MeO2C
Ph H Ph
Ph H
6 7 8
A: N2CHCO2Me, CuI
Scheme 4 Interaction of TPP with diazoacetic-acid methyl ester.
Another method of bacteriochlorin synthesis is based on the treatment of porphyrins

with osmium tetroxide. The reaction proceeds similarly to the above-described reduction
of porphyrins with diimide: interaction of a free base with OsO4 yields tetrahydroxybacte-
riochlorin 9, whereas osmylation of the Zn complex produces mainly isobacteriochlorin 11.
NH N NH N N N
A B
Zn
N HN N HN N N
HO
HO 10
C A A
HO
OH HO
OH
N N
NH N NH N
Zn
N N
N HN N HN
HO
HO HO
HO 9 11
O
A C
OH O
OH
O
NH N NH N NH N
C
+
N HN N HN N HN
Et
O O 13
O 12
Scheme 5 Interaction of octaethylporphyrin and its Zn complex with OsO4.
Pinacoline regrouping of vicinal tetrahydroxybacteriochlorins formed leads to an isomeric

mixture of ketobacteriochlorins 12 and 13 (Scheme 5) [8].
Later on, R. Pandey et al. used this approach to produce vicinal dihydroxy- and keto-
bacteriochlorins from natural chlorins: methyl ester of mesopyropheophorbide a 14 and tri-
methyl ester of mesochlorin e6 17 (Scheme 6) [9]. The first to be formed in the course of
the reaction are osmate complexes, which include the pyridine molecule; their reductive
splitting by hydrogen sulfide leads to respective vicinal diols 15 and 18.
Despite the improved spectral characteristics of the latter due to the batochromic shift
of the Q band to the red region, their photodynamic activity in vivo is lower than that of
respective keto derivatives 16 and 19. This is, apparently, due to a more hydrophobic char-
acter of the keto group, which enables ketobacteriochlorins to stay longer in tumor cells.
Another example of producing bacteriochlorins from chlorophyll a derivatives is os-
mylation of 3-formyl-3-devinylpurpurin 18 methyl ester 20 to form dihydroxybacteriochlo-
rin 21, that as the result of the pinacoline regrouping yields a mixture of 7-oxo- and
8-oxobacteriochlorins 22 and 23, which is the result of two variants of migration of alkyl
groups in pyrrole B (Scheme 7) [10].
O
OH
OH
NH N NH N
NH N A B
N HN N HN
N HN H H
H
H H
H O
O
O
14 MeO2C 15 MeO2C 16
MeO2C
O
OH
OH
NH N NH N NH N
A B
H N HN H N HN H N HN
H H H
CO2Me CO2Me CO 2Me
CO2Me CO2Me CO2Me
MeO2C MeO2C MeO2C
17 18 19
A: OsO4/Py, H2S; B: H2SO4
Scheme 6 Oxidation of methyl ester of mesopyropheophorbide a 14 and trimethyl ester of meso-

chlorin e6 and regrouping of vicinal diols.
CHO CHO OH
OH
NH N NH N
A
N HN N HN
PMe O O PMe O O
O O
20 B 21
O
CHO CHO
O
NH N NH N
N HN N HN
PMe O O PMe O O
O O
786 nm 777 nm
22 23
Scheme 7 Oxidation of 3-formyl-3-devinylpurpurin 18.

H H
B B
NH N O NH N O N
A D
B Me
P
N HN N HN O
D D
HO
PMe HO PMe PMe
PMe
26 27
O O O
A B B
NH N O NH N O NH N O
A
N HN N HN N HN
D C D C
HO OH
Me HO P Me
P Me Me OH
PMe P PMe P
28 29 30
PMe = CH2CH2CO2CH3
Scheme 8 Effect of electron-acceptor substituents in porphyrins on the regioselectivity of OsO4

oxidation.
R. Pandey et al. studied the effect of electron-acceptor substituents in porphyrins and

chlorins on the regiospecificity of OsO4 oxidation [11]. The presence of an electron-
acceptor substituent in one of the pyrrole rings of the porphyrin macrocycle was shown to
direct hydroxylation to the opposite pyrrole. Thus, the reaction with 3-acetyldeutero-
porphyrin IX 24 leads to the hydroxylation of ring C 25, whereas a similar reaction with
8-acetyldeuteroporphyrin IX 26 yields a vicinal diol in ring D 27. The reaction of 3,8-di-
acetylporphyrin 28 with OsO4 showed no stereoselectivity, and a mixture of diols 29 and
30 was formed (Scheme 8). A similar study was carried out on chlorins, the treatment of
which by OsO4 leads exceptionally to the hydroxylation of the pyrrole ring B.
The character of the substituents in the macrocycle also affects the progress of the pi-
nacoline regrouping [12]. An interesting regularity was found to exist between the total
number of electron-acceptor groups in the molecule of vicinal diol and the selectivity of for-
mation of 7- or 8-ketobacteriochlorins. Thus, irrespective of the arrangement of one carbo-
nyl group (the pentanone exocycle or the formyl group), in the macrocycle 31 under acidic
conditions 8-ketobacteriochlorin 32 is always formed, whereas the presence of two elec-
tron-acceptor groups (131-keto and 3-formyl) in chlorin 33 leads to a mixture of 7- and 8-
ketobacteriochlorins 34 and 35. The presence of the anhydride or imide exocycle conjugat-
ed with the macrocycle, as well as of three carbomethoxy groups in the lower part of the
chlorin macrocycle 36 and 37 also leads to a mixture of 7- and 8-ketobacteriochlorins with
various ratios of the isomers (Scheme 9).
Bacteriochlorins can also be obtained from divinylporphyrins by the Diels–Alder
reaction involving two vinyl groups (Scheme 10) [13, 14]. In turn, the latter are obtained
from vicinal dihydroxy derivatives by boiling in benzene in the presence of p-toluenesulfo
acid. A “double” Diels–Alder reaction by both vinyl groups in 8,18- 38 and 3,13- 40 in
OH
OH
O
NH N NH N
NH N B
A
N HN N HN
N HN H H
H
H H
H
O O
O
CO2Me 31 CO2Me CO2Me 32
CHO OH
CHO
OH
Me O
N
NH N
NH N
A B
35
N HN
H
N HN H +
O
H
H
O
O N
CO2Me
CO2Me 33
34
OH O
OH N
NH N NH N
A B
+
H N HN N HN O
H
H H N
CO2Me CO2Me
CO2Me CO2Me
MeO2C MeO2C
36
CHO CHO
OH
O
OH N
NH N NH N
A B
H N HN
+
H N HN O
H H
CO2Me CO2Me N
CO2Me CO2Me
MeO2C MeO2C
37
Scheme 9 Effect of electron-acceptor substituents in chlorins on the progress of the pinacoline re-
grouping.
CO2Me CO2Me
CO2Me
MeO 2C
NH N NH N
A
N HN N HN
CO2Me
MeO2C
38 CO2Me 39 CO2Me
MeO2C
CO2Me
NH N A Me NH N
N HN N HN
CO2Me
CO2Me
CO2Me CO2Me
40 41
A: dimethyl ester of acetylenedicarboxylic acid (DMAD), diazabicycloundecene (DBU)
Scheme 10 A Diels–Alder reaction with divinylporphyrins.
divinylporphyrins with various dienophiles, e.g., with dimethyl ester of acetylenedicarbox-

ylic acid (DMAD) made it possible to produce bacteriochlorins 39 and 41, in whose spectra
the Q band is shifted to the region of 800 nm.
A similar approach was used by Pandey et al. to produce bacteriochlorin from methyl
ester of 3-ethyl-7,8-dihydroxypurpurine 18 42 [15]. Boiling of the latter in o-dichloroben-
zene led to methyl ester of 8-vinyl-3-ethylpurpurine 18 43 with a 60% yield. The above re-
action with various dienophiles (TCE and DMAD) yielded adducts 44 and 45 (Scheme 11).
Similar adducts were obtained for chlorin 48; the DMAD adduct was present as cis-
50 and trans- 51 isomers (Scheme 12).
A close reaction is intramolecular cyclization of Ni complexes of 5,10- or 5,15-bis-
(vinylformyl)porphyrins 52 to form bacteriochlorins with two six-membered exocycles
conjugated with the main macrocycle 53 (Scheme 13) [16]. The latter proved unstable and
were oxidized in air.
2 Natural Bacteriochlorins and Their Chemical Modifications

Bacteriochlorophylls represent an independent group of natural chlorophylls and are wide-
spread in nature, mainly in numerous photosynthesizing bacteria [17–19]. The pigments
differ by the extent of hydrogenation of the macrocycle and by the character of the substit-
CN
NC CN CN
OH
OH
NH N A NH N NH N
B
H H H
O O O O O O O O O
MeO2C 42 MeO2C MeO2C
43 44
C
H+
MeO2C H
CO2Me CO2Me CO2Me
H MeO2C MeO2C
H
H
NH N B: NH N
NH N
E D
H N HN N HN H N HN
H
H H H
O O O O O O O O O
MeO2C MeO2C MeO2C
47 45 46
A: ɨ-dichlorobenzene, boiling
B: tetracyanoethylene (TCE)
ɋ: DMAD
D: Et3N
E: DBU
Scheme 11 Interaction of 8-vinylpurpurine 18 with dienophiles.
uents. Several modifications of bacteriochlorophylls are known. Thus, bacteriochlorophylls

a and b have been isolated from purple bacteria; green bacteria were the source of bacteri-
ochlorophylls a, c, d and e; sulfur bacteria, of bacteriochlorophylls c, d and e; bacteriochlo-
rophylls g were isolated from some types of photosynthesizing bacteria.
The mentioned bacteriochlorophylls are usually divided into two sufficiently large
groups (Scheme 14). The first group, which includes bacteriochlorophylls a, b and g, is
characterized by the presence of the tetrahydroporphyrin macrocycle 54 and, as the alkoxy
radical R4 for the first two, the residues of phytol (a), geraniol (b) and 2,10-phytadienol;
and for the third, farnesol (d) and geranyl geraniol. The second group with the dihydropor-
phyrin macrocycle 55, for which the name of chlorobium chlorophylls is also used, includes
bacteriochlorophylls c, d and e.
These bacteriochlorophylls are characterized by the presence of the pentanone ring,
α-hydroxyethyl group in position 3, methyl substituent at the δ-meso-carbon atom and
etherifying alcohol R4 – 2,6-phytadienol (CH3)2CH(CH2)3CH(CH3)(CH2)3C(CH3)=CH
(CH2)2C(CH3)=CHCH2OH and 2,16,20-phytatrienol (CH3)2C=CH(CH2)2C(CH3)=CH
(CH2)2CH(CH3)(CH2)3-C(CH3)=CHCH2OH.
CN
NC
OH
OH
NH N NH N NH N H
A B
H N HN 49
48
H
CO2Me CO2Me C
MeO2C
CO2Me CO2Me
H CO2Me H
MeO2C MeO2C
MeO2C
E NH N H D NH N
NH N H
51 50
A: ɨ-dichlorobenzene, boiling
B: TCE
ɋ: DMAD
D: Et3N
E: DBU
Scheme 12 Interaction of 8-vinyl-8-deethylmesochlorin p6 with dienophiles.
N N
CHO A N N
Ni Ni
N N N
N
CHO
53
52
A: HCl
Scheme 13 Intramolecular cyclization of bis(vinylformyl)porphyrins.
Pigments of the first group have intensive absorption bands in the near IR region. Of
special interest are bacteriochlorophylls a and b, whose maxima are not only shifted to the
red region, but also have the highest extinction coefficients. Of the two, Bchl a is usually
taken as the initial material to develop novel photosensitizers for photodynamic therapy
(PDT) of cancer and other possible photomedical applications.
Bacteriochlorophyll a is a porphin derivative, which contains the pentanone ring (exo-
cycle) condensed with the tetrahydroporphyrin macrocycle, various peripheral substituents
(CH3)2C=CH(CH2)2C(CH3)=CH(CH2)2C(CH3)=CH(CH2)2C(CH3)=CHCH2OH (a);
(ɋH3)2ɋH(ɋH2)3ɋ(ɋH3)=CH(CH2)2CH(CH3)(CH2)3-C(CH3)=CHCH2OH (b);
(CH3)2C=CH(CH2)2-C(CH3)=CH(CH2)2C(CH3)=CHCH2OH (c)
R1 H HO H R1
R2 R2
N N R3 N N
Mg R5 Mg
N N N N
R3
H H
H
H
H
O O
CO2Me
R4O2C R4O2C
Bacteriochlorophyll ɚ: R1=COCH3 Bacteriochlorophyll c: R1=R3=R5=CH3

R2=H, R3=C2H5; R2=C2H5;
Bacteriochlorophyll b: R1=COCH3 Bacteriochlorophyll d: R1=CH3, R2=C2H5 - C5H11,
R2 + R3= (=CHCH3); R3= C2H5;R5=H
Bacteriochlorophyll g: R1= - CH=CH2 Bacteriochlorophyll e: R1=CHO;R2=C2H5 - C5H11
R2 + R3= (=CHCH3); R3= C2H5;R5=CH3
Scheme 14 Major types of natural bacteriochlorophylls.
and the central atom of magnesium. To enumerate hydrogen and nitrogen atoms forming
the molecule of Bchl a and its derivatives, in this review we use the IUPAC nomenclature.
Derivatives of natural Bchl a can be divided into two groups [20]. The former includes
compounds, which contain the exocyclic fragment:
O
R1 H
bacteriochlorophyll ɚ: Ɇ=Mg, R1=Me,
H R2=COOMe, R3= phytyl;
N N bacteriopheophetin a: Ɇ=2H, R1=Me,
R2=COOMe, R3= phytyl;
Mg
bacteriopheophorbide a: Ɇ=2H, R1=Me,
N N R2=COOMe, R3=H;
H
bacteriopyropheophorbide a: Ɇ=2H, R1=Me,
R2=R3=H;
phytyl – a residue of the alcohol phytol
H
H
O HO
R2
R3O2C
The second group includes derivatives containing no exocyclic fragment:

O
R1
H
H bacteriochlorin ɟ6: R1=Me, R2=R4=COOH,
R3= CH2COOH
NH N
bacteriochlorin p: R1=Me, R2=R3=R4=COOH
H HN
trimethyl ester of bacteriochlorin p:
R1=Me, R2=R3=R4=COOMe
H
R4 R3 R2
Sources for production of Bchl a are biomass of the purple bacteria Rh. sphaeroides,
Rh. roseapersiana and Rh. capsulata. At our laboratory, Bchl a is isolated from the biomass
of Rh. capsulata [21], which contains no other bacteriochlorophylls; this greatly facilitates
the isolation and purification of the main pigment [22, 23]. Based on Bchl a, various groups
made research, the aim of which was to obtain stable individual bacteriochlorins with spec-
tral characteristics not inferior to initial bacteriochlorophyll and in some cases even exceed-
ing them. New photosensitizers should be less hydrophobic as compared with initial
bacteriochlorophyll and be sufficiently well soluble in polar solvents. It is also desirable for
these compounds to have functional groups, which enable adding other bioactive molecules
to them to increase the tropicity to cancer cells and targeted intracellular transport.
The first step on the way to solving these problems was to include an additional anhy-
dride exocycle into the main macrocycle, which led to the increase of stability of the pig-
ment [24]. For this, bacteriochlorophyll a 56 was extracted from biomass of the purple
bacteria Rh. capsulata and then, without isolation and additional purification, was oxidized
in an alkaline medium by oxygen of the air. The subsequent treatment with hydrochloric
acid led to the formation of the anhydride ring and production of bacteriopurpurine 57
(Scheme 15).
O O
C C
H H
H H
N NH N
Biomass of A
N
Mg B
Rhodobacter
capsulatus
H N N H N HN
H H
HO2C O O
H39C20O2C H O O
H3CO2C
56 57
Ⱥ: isopropanol; B: 1. O2/KOH, 2. HCl
Scheme 15 Production of bacteriopurpurine.
Transformation of the pentanone ring into the anhydride cycle, which includes oxida-
tion of carbon atom C-132 in the exocycle by oxygen of the air in alcoholic solutions of
chlorophyll a, was observed well back at the beginning of the 20th century by the founder
O
O O
A B
NH NH N
N N A N
Mg
N HN
N N N HN H
H D C H
H HR
H B
CO2Me O
CO2Me O CO2Me O
MeO2C
PhytylO2C PhytylO2C
58 61.R= OH
56
62.R= OH
O O
O
NH N NH N NH N
D C
N HN N HN N HN
H H
Me
H H
HO O O CO2Me O CO2Me O
HOOC
HOOC MeO2C MeO2C
63 E 59 60
O
O O
NH N
NH N NH N
N HN
N HN N HN
CO2Me
O CO2Me CO2Me
O O O CO2Me
ROOC
ROOC MeO2C 64 65
57a. R=H F
57b. R=Me
57c. R=Pr
A: 0.1% HCl; B: 5% H2SO4/MeOH; C: O2 of the air; D: KOH/CH3OH; E: CH2N2; F: KOH/CH3OH, CH2N2
Scheme 16 Chemical transformations of Bchl a in acidic and alkaline media.
of chlorophyll chemistry, Nobelist R. Willstätter [25]. He introduced the term “allomeriza-

tion” for the autooxidation process, which is catalyzed by bases [26]. It is established at
present that the mechanism of allomerization includes the formation of enol in the pen-
tanone ring, which is then oxidized to form lactone (unstable chlorin), which, in turn, is
transformed into the anhydride cycle conjugated with the chlorin macrocycle (purpurine
18). The occurrence of the exocycle, leading to an increase of the conjugation chain in the
molecule, causes a batochromic shift of the absorption bands Qx (545 nm) and Qy (818 nm)
and the emergence of a purple-red stain, with which the name of the compound (purpurine)
is associated [26].
The first to observe allomerization of bacteriochlorophyll a was H. Fischer in 1938
[27]. Recently, A. Scherz et al. reported the formation of 132-hydroxy allomers in a metanol
solution of Bchl a [28].
The oxidation process of Bchl a in BP 57 was studied in detail at the laboratory of
R. Pandey (Scheme 16) [29]. The treatment of bacteriopheophetine a 58 with a solution of
5% H2SO4 in methanol yielded, along with the required bacteriopheophorbide methyl ester
O
modification of C H
A B
the acetyl group
NH N H
Bacteriochlorophyll ɚ D C
H N HN
E
H
HO2C O O
etherification O
modification of
the anhydride cycle
Scheme 17 Possible chemical modifications of bacteriopurpurine.
59, a mixture of oxidation products. Analysis of the latter showed it to consist of chlorin 60
and a mixture of diastereomers of 132-(R/S)-hydroxyderivatives 61 and 62 (epimers). The
presence of the hydroxyl group in the pentanone ring was reliably proven by the spectra of
1
H NMR; in this case, the ratio between R and S isomers was 1:4. These detailed studies
have shown that the S epimer, in which the carbomethoxy group in position 132 and the
propionic acid residue in position 17 of the macrocycle are directed to different sides rela-
tive to the plane of the macrocycle, is thermodynamically more stable under acidic condi-
tions [30, 31].
However, work with derivatives of Bchl a in acidic media is strongly complicated due
to the rapid oxidation of the pigment. The authors also observed allomerization in the pres-
ence of alkali. Thus, bacteriopheophorbide 59 in an air-bubbled solution of KOH–propanol
is allomerized to form “unstable bacteriochlorin” 63. The mechanism of allomerization un-
der alkaline conditions is unknown, but an indirect proof of the formation of intermediate
63 can be production of bacteriochlorin with the glyoxalic acid residue in position 15 of the
macrocycle 64 at the action of diazomethane on bacteriopheophorbide 59. “Unstable bac-
teriochlorin” 63 is converted into BP in the evaporation of the solvent; both free acid 57a
and ester 57c can be formed. As BP 57a possesses a low solubility in organic solvents, it is
expedient to convert it into methyl ester 57b, which facilitates chromatographic purifica-
tion. To increase the yield of BP, our laboratory developed a method of its production with-
out isolating bacteriopheophorbide as an intermediate. In this case, allomerization of Bchl a
in an air-bubbled KOH–isopropanol solution takes longer (1.5–2 h), apparently, due to the
presence of carotenoids, which are radical traps [32].
For bacteriopurpurine, a number of chemical transformations are possible (Scheme
17). They include the modification of the acetyl group with its reduction to an α-hydroxy-
ethyl group and its conversion into a vinyl group; conversion of the anhydride cycle into
the imide cycle; etherification of the propionic-acid residue by alcohols and serine.
Bacteriopurpurine is stable only in neutral and acidic media; in the presence of bases,
there occurs a rapid opening of the anhydride cycle, which leads to the recovery of the main
spectral band in the electronic spectrum.
Stability was increased and the spectral characteristics were improved by producing
cyclic imides of chlorins and bacteriochlorins by substitution of oxygen in the exocycle for
the nitrogen atom. The reaction was adjusted on methyl ester of purpurine 18 66 (Scheme
18) [33, 34]. The latter reacted to hexylamine at room temperature, giving a mixture of
NH N NH N
A
H N HN H N HN
H H
R1 R2
O O O
CO2Me CO2Me 67
66 a. R1 = CO2H, R2 = CONH(CH2)5CH3
b. R1 = CONH(CH2)5CH3, R2 = CO2H
NH N NH N
C
H N HN H N HN
R
H H
O N O O
X Y
CO2Me CO2Me
C6H13 68
69 a. X = O, Y = N(CH2)5CH3
b. X = N(CH2)5CH3, Y = O
A: C6H13NH2; method 1 – B: DCA; ɋ: DBU; method 2 – from 67 to 69: 1. CH2N2, 2. KOH/CH3OH
Scheme 18 Synthesis of cyclic imides in the chlorophyll a series.
isomeric amides in positions 131 67a and 151 67b at a ratio of 6:1 with a yield of 95%. In
order to obtain cyclic isoimides, the free carboxyl group of amides was activated using two
techniques.
In the first technique, a mixture of amides was treated with DCC to form two isoimides
(λmax = 690 and 696 nm) at a ratio of 6:1 with the total yield of 96%. In one of them, the
nitrogen of hexylamine is in position 131 68a; in the other, in position 151 68b. The mixture
of isoimides at the action of diazabicycloundecene (DBU) in toluene in an alkaline medium
at 60° is converted into the required cycloimide 69 with a low yield. The second approach
includes etherification of intermediate amides by diazomethane, followed by the treatment
of formed esters by a KOH methanol solution. In this case, cycloimide 69 is formed with
the yield of more than 80%. The authors note that the substitution of DBU by a stronger
KOH or NaOH base leads to a further increase of the yield.
Optimized conditions were used for the synthesis of bacteriochlorin derivatives
(Scheme 19) [34]. The mixture of amides 70a and 70b, obtained as the result of opening
the anhydride cycle of bacteriopurpurine by hexylamine, yields unstable carbodiimide
O O
H H
H H
NH N NH N
A
H N HN H N HN
H H
R1 R2
O O O
CO2C3H7 CO2C3H7 70
1 2
a. R = CO2CH3, R = CONH(CH2)5CH3
b. R2 = CONH(CH2)5CH3, R2 = CO2CH3
O O
H H
H H
NH N NH N
C
H N HN H N HN
H H
O N O X Y
O
R CO2C3H7
CO2C3H7 71
72. R = C6H13 a. X = O, Y = N(CH2)5CH3
b. X = N(CH2)5CH3, Y = O
A: C6H13NH2; method 1 – B: DCA; ɋ: DBU; method 2 – from 70 to 72: 1. CH2N2, 2. KOH/CH3OH
Scheme 19 Synthesis of cyclic imides in the bacteriochlorophyll a series.
derivatives, which are rapidly converted to the more stable cyclic isoimides 71a and 71b.
The latter were separated chromatographically into individual isomers (λmax = 804 and 796
nm) at a ratio of 6:1. The base-catalyzed intramolecular cyclization of isoimides led, with
a yield of 45%, to cycloimide 72 (λmax = 822 nm). Similar results were obtained with
amides, whose carboxyl groups were methylated prior to the treatment by the base.
In an alkaline medium, along with the synthesis of the required cycloimide 72, there
occurred the oxidation of 12-CH3 group to form minor products: 12-formyl (4–6%) 75 and
12-hydroxymethyl (2–4%) 76 derivatives (Scheme 20). The authors believe that this un-
usual oxidation is the consequence of enolization leading to tautomers 73 and 74, which are
apt to oxidation in air. In turn, keto-enolic tautomerism in cycloimide is possible owing to
the strong electron-acceptor effect of the imide exocycle conjugated with the main tetra-
pyrrole system [35].
An unusual bacteriochlorin 78, having a Qy band at 849 nm, was obtained from 12-
hydroxymethylcycloimide 76 under acidic conditions (Scheme 21). The 2D ROESY spec-
tra showed the interaction of 10-H meso-proton with the adjacent 12-CH2 group and the ab-
sence of 8-H proton. The authors assume cycloimide 78 to have the structure depleted of
8-H proton to shift the double bonds inside the macrocycle. As the mechanism, they propose
N HN N HN H N HN H
A
H H
B
N O N O N
O O O O
Hexyl PrO2 C Hexyl Hexyl
PrO2C PrO2C
72 73 74
N HN N HN
CHO CH2OH
and
O N O O N O
Hexyl PrO2C Hexyl
PrO2C
75 76
A: KOH; B: HCl
Scheme 20 A possible mechanism of formation of 12-formyl- and 12-hydroxymethyl cycloimide

of bacteriochlorin p in an alkaline medium.
O O O
NH N NH N NH N
A
N HN N HN N HN H
CH2OH CH2
H
O N O O N O N
O O
PrO2C Hexyl Hexyl Hexyl
PrO 2C PrO2C
76 77 78
+
A: H
Scheme 21 Transformations of 12-hydroxymethylcycloimide of bacteriochlorin 76 in an acidic

medium.
elimination of the hydroxyl group under acidic conditions to form a carbocation of the ben-
zyl type 77, which is stabilized due to the abstraction of the proton at C-8 [29].
In contrast with natural bacteriochlorophyll a, the isoimide and imide analogues are
more stable, and their spectral characteristics compare favourably to respective precursors.
The photodynamic activity of the derivatives of chlorin p6 and bacteriochlorin p can
be further increased by introducing trifluoromethyl groups into cycloimide molecules
(Scheme 22) [36].
Fluorine-containing substituents enhance the solubility of compounds in lipids, which
increases the transport rate of such molecules through lipid membranes. The authors note
that the activity of PS in vivo is affected not only by the nature of the substituents but also
by how they are arranged in the macrocycle. Thus, cycloimide of chlorin p6 79b, containing
a bis-(trifluoromethyl)benzyl grouping at the nitrogen atom of the macrocycle, evokes a
more efficient inhibition of tumour growth as compared with the isomer containing this
O Butyl
NH N NH N NH N
A B
H H H
O O O O N O R O N O
R
CO2Me CO2Me CO2Me
R=CH3 R 79 R
C R=CF3
a. R=CH3
b. R=CF3
OR
NH N NH N
D a. R = H2C
H N HN H N HN
H H
O N O O N O CF3
CO2Me Butyl CO2Me Butyl
b. R = H2 C
80
CF3
A: 3,5-dimethylbenzylamine or 3,5-bis(trifluoromethyl)benzylamine;
B: HBr/CH3COOH, C4H7OH; C: C4H9NH2; D: HBr/CH3COOH, 3,5-dimethylbenzyl alcohol or
3,5-bis(trifluoromethyl)benzyl alcohol
Scheme 22 Synthesis of fluorine-containing cyclic imides of chlorin p6.
substituent in the upper part of the chlorin macrocycle 80b (respectively, 100 and 66% of
tumour regression in 90 days).
Studies of the relation between the structure and activity were continued on cycloimi-
des of bacteriochlorin p. For this, bacteriopurpurine methyl ester was treated with 3,5-bis-
(trifluoromethyl)benzylamine. Along with required cycloimide, a Schiff base was formed;
the base proved to be labile in in vivo experiments. However, during the reduction of the
C=N bond, stable cycloimide 81 with the Qy band in the region of 796 nm was formed [37].
CF3
NH CF3
H
NH N
H N HN
H
O N O CF3
CO2Me
81 CF3
81
O NOH NOH
H H H
H H H
NH N NH N NH N
A A
H H H
HO2C O O HO2C O O O HO2C O N O
O
57 82 OH 83
B C or D
2
O NOH NOR
H H H
H H H
NH N NH N NH N
A
H H H
NaO2C COONa NaO2C COONa
COONa R1O 2C O N O
COONa
OR3
84 85 86
a: R = H; R2 = R3 = COCH3
1
b: R1 = R3 = CH3; R2 = H
A: NH2OH.HCl, Py; B: NaOH/CH3OH; C: Ac2O; D: CH2N2, (C2H5)2O
Scheme 23 Interaction of bacteriopurpurine with hydroxylamine.
Our laboratory was the first to perform the synthesis of cycloimides of chlorins and
bacteriochlorins by acting with highly nucleophilic agents hydroxylamine and hydrazine
hydrate on bacteriopurpurine.
Initially, for modification of bacteriopurpurine we used our earlier proposed method
of converting purpurine 18 to N-hydroxycycloimide [38, 39]. Herewith, the presence of the
acetyl group in bacteriopurpurine instead of the vinyl substituent in purpurine 18 leads to
the emergence of an additional reaction centre. Conducting the reaction of bacteriopurpu-
rine with hydroxylamine, it was shown (Scheme 23) that oxime 82 was initially formed
[40]. To prove this, we performed chemical conversions, which included the opening of the
anhydride cycle in oxime 82 and the treatment of bacteriochlorin p 84 by hydroxylamine
in pyridine. The identity of the products obtained indicates that in the treatment of BP by
hydroxylamine the first to enter into the reaction is the acetyl group.
Subsequent studies of the interrelation between bacteriopurpurine and hydroxylamine
have shown that, when excess reagent is used and time is increased up to 10 h, the second
molecule of hydroxylamine enters into the reaction with the anhydride cycle to yield oxime
of N-hydroxycycloimide of bacteriochlorin p 83. The latter compound can be obtained both
from bacteriopurpurine 57 and from intermediate oxime 82. The course of the reaction was
followed chromatographically and spectrally by the change of position of the long-wave Qy
band. In the first three hours, the maximum at 818 nm shifted to 792 nm, which corresponds
to the formation of oxime 82, after which it gradually returned to the long-wave region (812
nm). We believe that at this stage oxime interacts with the second molecule of hydroxy-
lamine to form compound 83. Thus, a derivative of hydroxamic acid was first obtained in
OH
N N
HO H H
NH N H NH N H
N HN N HN
H H
H H
O O O N O
N
CO2Me CO2Me
OMe OMe
Scheme 24 Syn- and anti-isomers of oxime 86b.
O O OH
H H H
H H H
NH N NH N NH N
A B
H H H
CO2H CO2H CO2H CO2H
O O O
88
57 CO2H 87 CO2H
CO2H E C or D
H H OH
H H
H
H
NH N NH N
F NH N
H N HN H N HN
H N HN
H H
H
O N O O O O
O O O
CO2R
CO2R OR1
CO2H 89
91 90
a. R=H, R1=H
b. R=CH3, R1=H a. R=H
G b. R=CH3
H c. R=C2H5
c. R1=R2=CH3
d. R=CH3, R=Ts
A: NaOH, CH3OH; B: NaBH4; C: HCl, dioxan; D: TsCl, C5H5N; E: TsOH, CHCl3; F: NH2OH HCl, Py;
G: CH2N2, Et2O; H: TsCl, Py
Scheme 25 Synthesis and modifications of 3-vinyl-3-deacetylbacteriopurpurine 90.
the bacteriochlorophyll a series. The derivative, having a mobile hydrogen atom in its com-
position, readily enters into the acylation and alkylation reactions [41]. Thus, the treatment
with acetic anhydride leads to the formation of diacetate 86a, which proved rather labile
and in storage was decomposed into monoacetate and the initial compound 83.
The treatment of oxime of hydroxamic acid 83 by diazomethane yields N-methoxy-
cycloimide 86b with a good yield. This compound proved much more stable than acetate
and showed a good photodynamic activity in in vitro and in vivo experiments. Two inter-
esting facts were discovered in the course of the studies. The acidity of the hydrogen atom
in the oxime function is decreased so much that the function is not involved in the reaction
with diazomethane. Besides, comprehensive analysis (TLC, mass spectra) of N-methoxy
derivative 86b showed the substance to be a mixture of two isomers with very close values
of Rf. The isolated isomers were characterized by the 1H NMR spectra, including 1D NOE
spectroscopy; based on the data obtained, it was concluded that oxime 86b existed in the
form of two stereoisomers (sin- and anti-) (Scheme 24). Such isomers are formed in the in-
teraction of ketones with hydroxylamine [42].
As the acetyl group in BP strongly complicates the progress of the reaction with hy-
droxylamine, it was decided to convert it into the vinyl group.
The vinyl group in compound 90 was obtained from the α-hydroxyethyl group in al-
cohol 88 under the action of p-toluenesulfo acid (Scheme 25) [43]. The acetyl group was
reduced not on bacteriopurpurine 57, but on bacteriochlorin 87, as the treatment of bacte-
riopurpurine with sodium boron hydride had been earlier shown to contribute to the con-
version of the anhydride exocycle into the δ-lactonic one [44].
Triacid 87 obtained from BP was reduced by sodium boron hydride to respective al-
cohol 88, which has an absorption maximum at 740 nm. The treatment of the latter by hy-
drochloric acid in dioxane led only to the closure of the anhydride cycle, but not to the
dehydration of the α-hydroxyethyl group. Compound 89 was also observed to be formed
at the action of tosylchloride on alcohol 88; if the reaction was continued for more than 2 h,
the predominant product formed was O-tosylate.
A more efficient method of producing the vinyl group was developed based on the
treatment of alcohol 88 by p-toluenesulfo acid. Herewith, it was found that, along with the
formation of the vinyl group, there occurs the closing of the anhydride exocycle in com-
pound 90, which leads to the recovery of the main spectral band Qy to the region of 783 nm.
Interestingly, if the reaction was performed in chloroform, the product was obtained as free
acid 90a, whereas addition of methyl or ethyl alcohols directly to the reaction medium led
to the rapid etherification of propionic acid residue to form respective methyl 90b or ethyl
90c esters. This activity of the carboxyl group is, evidently, due to the formation of the
mixed anhydride of the residue of propionic acid and p-toluenesulfo acid.
3-Vinyl-3-deacetylbacteriopurpurine 90a is a structural analogue of purpurine 18 and,
for this reason, it was of interest to perform the above-described reaction with hydroxy-
lamine to assess the effect due to the conversion of the acetyl group into the vinyl group.
The treatment of compounds 90a and 90b by hydroxylamine produced respective
N-hydroxycycloimides 91a and 91b. Although the absolute values of the wavelengths of
the Q band in the spectra of N-hydroxycycloimides 86b and 91c are very close to 812 and
809 nm, the relative change of the long-wave absorption in the reaction products strongly
differs as compared with the initial substances. In the case of bacteriopurpurine, the forma-
tion of oxime by the acetyl group leads to a virtually complete levelling down of the effect
from the introduction of the imide exocycle. As the result, the terminal cycloimide is only
6 nm worse in the long-wave absorption than the initial pigment. In the case of the vinyl
derivative of bacteriopurpurine, transformation of the anhydride cycle to the imide cycle
gives a significant increment of the wavelength of the Q band up to 27 nm.
Recently, Sasaki and Tamiaki [45] synthesized a series of derivatives of bacteriopy-
ropheophorbide 92, which contain various substituents in pyrrole A.
The vinyl group was obtained by a different method as compared with that described
above. The high yield of compound 94 was achieved at the expense of the outgoing mesyl
group. Further on, the vinyl group was oxidized to the formyl group 95 using OsO4 and the
subsequent splitting of diol NaIO4 (Scheme 26). As the reactivity of the formyl group in
NH N
Rhodobacter A B C
sphaeroides BChl - a
(purple bacterium) N HN
O
CO2Me
92
OH H 0
Ph
NH N D NH N E NH N
N HN N HN N HN
O O O
CO2Me CO2Me CO2Me
93 94 95
Scheme 26 Synthesis of bacteriopyropheophorbide derivatives with various substituents in

pyrrole A.
the chlorin series has been well studied, the authors carried out a number of similar trans-
formations in the bacteriochlorin series, which ncluded the oxidation and reduction of the
formyl group to the carboxyl and hydroxymethyl groups to form compounds 96 and 97, as
well as the Knoevenagel reaction with dicyanomalonic ester, which led to compound 98
(Scheme 27).
Studies of N-hydroxycycloimides of bacteriochlorin have shown that the hydroxyl
group can be successfully used to produce alkyl-substituted derivatives. At the same time,
introduction of acyl radicals by this way is less promising due to the lability of such deriv-
atives.
In this connection, we developed a method of producing N-aminocycloimides of bac-
teriochlorin p [46]. As is known, hydrazine and its derivatives possess a high nucleophilic-
ity with respect to sp2-carbon atoms [47]. For this reason, anhydrides of acids are
convenient reagents for acylation of hydrazine.
In the case of BP 57, we have shown (Scheme 28) that its treatment by hydrazine hy-
drate in pyridine initially leads to monohydrazide, evidently, in the form of two isomers 99a
and 99b. The acetyl group in this case is converted to hydrazone [45]. In the subsequent
treatment of the reaction mass by HCl, there occurs the intramolecular cyclization to form
an additional six- or seven-membered cycle 100a or 100b. Hydrazone at this stage was con-
verted to initial ketone.
It is known that during the acylation of hydrazine after the addition of the first acyl
group the subsequent acylation, due to a partial deactivation of the first nitrogen atom, usu-
ally occurs by the second amino group [48]. Thus, in the interaction of hydrazine with an-
hydrides of aromatic dicarboxylic acids symmetric hydrazides are formed with the increase
of the initial five-membered cycle to the six-membered cycle (Scheme 29) [49].
In the case of bacteriopurpurine, this could lead to the formation of hydrazide with six-
100a or seven-membered 100b cycles. The use of 2D heteronuclear resonance and the study
HO 0 0
NH N NH N
N HN N HN
O O
CO2Me CO2Me
96
A C
0 OH
H
NH N NH N
B
N HN N HN
O O
CO2Me CO2Me
95 97
D E
CN
OH
NC
NH N N N
Zn
N HN N N
O O
CO2Me CO2Me
98
A: NH2SO3H/NaClO2, THP, 2-methyl-2-butene; B: t-BuNH2*BH3, CH2Cl2;

C: AcOH/EDC*HCl/DMAP, CH2Cl2; D: malonic acid dinitrile, Et3N, THP;
E: Zn(OAc)2, CH2Cl2/CH3OH
Scheme 27 Chemical transformations of 3-formyl-3-deacetylbacteriopyropheophorbide.
of 1H NMR spectra of the obtained compound at various temperatures, as well as chemical

modifications including the formation of Schiff bases, alkylation and acylation of the amino
group confirmed the six-membered structure of the exocycle 100a. The developed method
of producing N-aminocycloimide derivatives of bacteriochlorophyll a is distinguished by
simplicity and high yields [50].
N NH2
O
H H
H H
NH N NH N
A
H N HN H N HN
H H 2
O O COR1 COR
O
CO2H CO2H
57 99
a: R1=NHNH2; R2=OH
b: R1=OH; R2=NHNH2
B,ɋ
O O
H H
H H
NH N NH N
H N HN H N HN
H H
O O O N N O
N
CO2Me NH2 CO2Me H H
100a 100b
.
A: N2H4 H2O, pyridine; B: 1N HCl; C: CH2N2, Et2O
Scheme 28 Reaction of bacteriopurpurine with hydrazine hydrate.
O O
NH
O + NH2 NH2
NH
R R
O O
Scheme 29 Formation of cyclic hydrazides in the interaction of hydrazine with anhydrides of aro-
matic dicarboxylic acids.
3 Amphiphilic and Water-soluble Derivatives of Bacteriochlorins

Natural bacteriochlorins are known to be distinguished with an increased hydrophobicity
[2]. To be successfully used in PDT, they should have a more balanced ratio of hydrophobic
and hydrophilic substituents in the macrocycle. Usually, this is achieved by introduction of
one, two or three carboxyl groups, amino acid residues or hydroxyl-containing functions
into the molecule [51–54]. Thus, introduction of serine by the residue of propionic acid by
enzyme re-etherification of Bchl a makes it possible to significantly increase its solubility
in water. Such conjugates of bacteriochlorophyllide a (Bchlfd a) with serine preserve the
photophysical properties of Bchl a, generate AOS with a high yield, but, unfortunately, are
subject to photooxidation, demetallation in a weakly acidic medium and biodegradation,
which restricts their use in clinic.
Substitution of the central atom of Mg by Pd, etherification of the propionic acid
O O
H H
H H
N N N N
A
Mg Mg
H N H N N
H H
HO2C
HO2C O O O
MeO2C MeO2C
101 H2N O 102
O
O
H
H
H
H
NH N
C N N
Pd
H N HN H N N
HO2C H
HO2C H
O O O O
H2N MeO2C
O 103 H2N MeO2C 104
O
A: serine; B: HCl; C: Pd(CH3COO)2
Scheme 30 Production of complexes of bacteriochlorophyll a derivatives with serine.
residue and re-etherification of the carbomethoxy group in the pentanone ring lead to stable
derivatives of [Pd]-Bchlfd a with a high photodynamic activity [55]. The quantum yield of
AOS for these compounds is sufficiently high – from 1 in nonpolar solvents down to 0.5 in
aqueous solvents. Bacteriochlorophyllide a 101 and its complex with serine 102 showed a
high anti-tumour activity on the M2R mouse melanoma cell line (LD50 0.2–0.5 µM)
(Scheme 30) [56]. Substitution of Mg by Pd 104 increased the photodynamic activity,
which led to a decrease of the value of LD50 to 0.01–0.03 µM. Besides, owing to the high
fluorescence of 102 in cancer tissue as compared with healthy tissue (the selectivity reaches
8–10), the latter, along with PDT, can be also used for fluorescent diagnostics (FD).
A series of new negatively charged water-soluble derivatives of Bchl a was obtained
at the Weizmann Institute of Science, Israel. These compounds proved promising for vas-
cular-targeted photodynamic therapy (VPT). The damage of vessels providing for the blood
supply of the tumour was found to be the essential factor of its necrosis. VPT is efficient in
the treatment of hard tumours, as well as non-tumour processes associated with increased
vascularization, e.g., age-related macular degeneration. Regression and necrosis of the tu-
mour occur as the result of both the death of cancer cells and due to the occlusion or perfo-
ration of tumour vessels. However, hemorrhagic necrosis of the tumour, caused by the
above-named water-soluble derivatives of bacteriochlorophyll a, can have undesirable con-
sequences, especially when tumours are localized in such vital organs as bronchi and lungs.
The method of VPT leads to necrosis of the central part of the tumour, which can spread to
95% of tumour’s volume. However, cancer cells on the periphery of the tumour can fail to
O
O
N N
N N
M
Pd
N N
N N
O O NH
CO2CH3 CO2CH3
R O CO2-K+
109 SO3- K+
56:M=Mg, R=phytyl
105:M=2H, R=OH
106:M=Pd, R=OH (WST09, Tookad)
107:M=Pd, R= O-succinimide-SO3-Na+a+
108:M=Pd , R=NH-(CH2)3-SO3-Na+
SO3- K+
O N O
N N N N
N N
Pd Pd
Pd
N N N N
N N
O NH O NH
NH NH CO2CH3 O NH
O O
CO2CH3 CO2-K+ CO2CH3
SO3- K+ SO3- K+
SO3- K+
SO3- K+ SO3- K+
110 111 112: M = Pd

113: M = 2H
114: M = CuII
115: M = Zn
116: M = MnIII
Scheme 31. Negatively charged PS based on bacteriopheophorbide and its metal complexes.
die and grow again. In this connection, the VPT method is not universal and can be consid-
ered only in combination with chemotherapy, radiotherapy and photodynamic therapy.
Water-soluble negatively charged PS were obtained by aminolysis of the pentanone
exocycle in bacteriopheophorbide and its metal complexes (Scheme 31). Their hydrophilic
properties and capability of aggregation in aqueous solutions was studied [57]. Amphiphi-
licity was assessed by the octanol/water distribution coefficient (P). The effect of periph-
eral substituents on the ability of obtained compounds to be solved in water and polar
organic solvents was shown. It was found that the opening of the exocycle in bacte-
riopheophorbide significantly increased the hydrophilicity of bacteriochlorins and, as a
consequence, enhanced their ability to be solved in polar organic solvents, including meth-
anol, ethanol, DMFA and DMSO. Besides, compounds with the open cycle 109–116 are
well soluble in aqueous solutions (PBS) – up to 40 mg/ml, which is much greater than the
solubility of compounds with the pentanone ring, but having polar substituents on the pe-
riphery of the macrocycle (compound 108, 4 mg/ml). The coefficient P strongly increases
(1:19) at the substitution of the central metal ion Pd(II) by Mn(III) 116. At the same time,
a decrement of the hydrogen chain in the alkyl substituent 109 as compared with 112 does
not in practice affect the distribution coefficient. All compounds with the open exocycle in
aqueous solutions form aggregates (2–8 molecules), which dissociate upon dilution, inclu-
sion into micelles, as well in solutions with physiological concentrations of serum albumin.
Among new PS of the bacteriochlorin series developed at the Weizmann Institute of
Science in cooperation with Steba-Biotech (France) and Negma-Lagards (France), we
should note the second-generation Tookad, which is a Pd complex of bacteriopheophorbide
106. Tookad exhibits a high activity to such tumours as rat glioma [58], prostate cancer [59],
HT29 human colon carcinoma in experiments on laboratory animals.
The main mechanism of action includes tumour vessel damage, which leads to hypoxy
and necrosis of the tumour. At present, the preparation is at the second stage of clinical tests,
which are carried out at medical centres of Canada, Europe and Israel. As Tookad is poorly
soluble in aqueous solutions (octanol:water = 24:1), it is introduced as a suspension with
Cremophor.
At the action on the Pd complex of bacteriopheophorbide by taurine (2-sulfo-
ethylamine) there occurs the opening of the pentanone cycle and the formation of dianion
(code name WST11) 112 [60]. The preparation possesses a good solubility in phosphate
buffer (up to 50 mg/ml), where it is present in the shape of small aggregates. In blood serum-
containing solutions, it is subjected to deaggregation and occurs as monomers in a complex
with serum albumin (BSA) and high-density lipids.
O
Absorption
N N
Pd
N N
O SO3Na
N
CO2CH3 H
CO2H
nm
112
To increase the hydrophilicity of cycloimides of bacteriochlorin p, we synthesized de-

rivatives, which in pyrrole A contain hydroxyl-containing substituents attached to the mac-
rocycle by an ether bond (Scheme 32). For this, initial bacteriopurpurine 57 was
transformed into bacteriochlorin p 87, the acetyl group in which was reduced by NaBH4
88. The subsequent closing of the anhydride cycle using TFA 89 and the treatment by hy-
droxylamine and diazomethane yielded 3-(α-hydroxymethyl)-3-deacetyl-N-hydroxymeth-
ylcycloimide of bacteriochlorin p 117. The hydroxyl group in pyrrole A was activated by
means of trifluoroacetic acid anhydride; the obtained trifluoroacetate 118 was condensed
with the methyl ester of ethylene glycol, di- and triethylene glycol and glycerol to yield
compounds 119–122.
Experiments in vitro on HeLa and A549 cell lines and in vivo on lymphoma mice have
shown that the phototoxicity of the above-named compounds is 20 times as high as that of
the initial bacteriopurpurine [61].
The cause of the so high activity of preparations obtained is due to the increased se-
lectivity to cancer cells (tropicity 8–13) and the significant quantum yield of the generation
of singlet oxygen (0.54–0.57). Analysis of the distribution of PS in tissues showed the larg-
est accumulation in tumour vessels, leading to hypoxy and necrosis of the tumour, which
O
C CF3
OH O
H H R1 H
H H H
NH N NH N NH N
A B
H N HN H N HN
H N HN
H H
H O O
O O N
O O N
N OMe
OMe CO2Me
CO2Me
CO2Me OMe
117 118 119 - 122
119 O 121 O O
O O OH
HO
120 O OH 122 O OH
O
A: (CF3CO)2O; B: 1. CH2(OH)CH2OMe, 2. HO(CH2CH2O)2H, 3. HO(CH2CH2O)3H,

4. HOCH2CH(OH)CH2OH
Scheme 32 Synthesis of cycloimides of bacteriochlorin p with polar substituents in pyrrole A.
O O O
H H H
H H H
NH N NH N NH N
H H H
MeO2C O MeO2 C O O MeO2C O O
N O N N
NH2 HN O HN O
C C
N N I
A: isonicotinic acid chloroanhydride, Py; B: CH3I, boiling
Scheme 33 Synthesis of cationic cycloimide of bacteriochlorin p.
is absolutely consistent with the data by Israeli investigators for the action mechanism of
bacteriochlorophyll a derivatives.
Much less is known of the introduction of positively charged substituents into bacte-
riochlorins. However, just these photosensitizers may prove to be the most efficient in pho-
todynamic antimicrobial therapy. The method is based on the inactivation of viruses,
bacteria, yeasts and protozoa by active oxygen species, which are generated by photosen-
sitizers in illumination [62].
Available literature data show that this method is considerably behind photodynamic
O O
H O
H H
H H H
NH N NH N NH N
C
H N HN HN
H N H N HN
H H H
O N O O N O O N O
CO2Me CO2Me
HN O N O CO2Me
H 3C NHMe
123 125 127
N N
A
B
O O
H O
H
H H
H
H
NH N NH N
NH N
H N HN H N HN
H N HN
H H
H
O N O O N O
O N O
CO2Me
CO2Me N O
HN OH CO2Me
CH3 NH2
100
N
N
123 a 126
Scheme 34 Chemical transformations of cycloimide 123.
therapy of cancer by the level of fundamental elaboration and practical application. There
are only separate data on the photosensitization of nonpathogenic yeasts in the presence of
porphyrins and phthalocyanines [63–65]. As is known, the outer surface of bacteria carries
a negative charge, in connection with which the most efficient coupling to bacterial cells
and the photodynamic action is expected from cationic photosensitizers.
Such photosensitizers based on N-cycloimides of bacteriochlorin have been synthe-
sized at our laboratory. Initially, it was planned to use for these purposes the N,N-dimeth-
ylamino derivative. However, the treatment of it by excess methyl iodide or dimethyl
sulfate failed to lead to the quaternization of the nitrogen atom [46].
A more successful effort was introduction into the N-amino derivative 100 of the isoni-
cotinic acid residue 123 followed by the quaternization of the nitrogen atom in the pyridine
ring 124 (Scheme 33) [66]. For this, cycloimide 100 was treated with isonicotinic acid chlo-
roanhydride in pyridine. We showed hydrazide obtained to exist in two isomeric forms 123
and 123a, which are formed owing to keto-enolic tautomerism or due to the absence of free
rotation around the bond C(O)–N.
O O
H H
H H
NH N NH N
A B
100
H N HN H N HN
H H
O N O O N O
CO2Me CO2Me
HN O HN O
128 129
N
N
C C
O O
H H
H H
NH N NH N
H N HN N HN
H
H H
O N O O N O
CO2Me CO2Me
H2N O H2N O
N
I
130 N
131
A: 6-methylnicotinic acid chloroanhydride, pyridine; B: 6-methylpicolinic acid chloroanhydride, pyridine;

C: CH3I, boiling
Scheme 35 Synthesis and quaternization of cycloimides with residues of pyridinecarboxylic acids.
O N O
N O
H
N
Me
Figure 2 Formation of a five-membered ring in hydrazide 129.
To establish the structure of the isomers, chemical modification of cycloimide 123 was
carried out; it included the treatment of the latter by diazomethane (Scheme 34). As the
result, two substances with the same molecular mass but different chromatographic mobil-
ity were obtained, which enabled their isolation in the individual form. A more mobile iso-
mer was assigned the structure of N-methylhydrazide 125; and the other isomer, of
O-methylimidate 126.
The structure of N-methylhydrazide 125 was confirmed by countersynthesis. For this

purpose, N-aminocycloimide 100 was treated with a stoichiometric amount of methyl io-
dide, and N-monomethylaminocycloimide 127 was condensed with isonicotinic acid chlo-
roanhydride. The obtained compound by its molecular mass and chromatographic mobility
proved to be identical to the fast-moving isomer 125.
The other isomer 126 proved rather labile and rapidly degraded to form N-aminocy-
cloimide 100.
The 1H NMR spectra of the fast-moving isomer 125 preserved the signal doubling sim-
ilar to that observed for initial hydrazide 123. This phenomenon is, apparently, due to the
existence in amides of one more type of isomerism, which emerges owing to the absence
of free rotation around the bond C(O)–N. As the rotation barrier for such isomers is com-
paratively low, it can be overcome by increasing the temperature. Indeed, during the record-
ing of the 1H NMR spectra for compounds 123 and 125 at 50° the signals merged. At the
same time, in isomer 126, whose 1H NMR spectrum was recorded in the first hour after iso-
lating the substance, there was no such signal doubling, which corresponds to the presence
of only one Z or E isomer.
The cationic photosensitizer 124 is a much more hydrophilic compound as compared
with initial bacteriochlorins 100 and 123. In this connection, it is characterized by a better
solubility in water–alcoholic solutions, which makes it more promising for medical appli-
cations.
Along with isonicotinic acid, other pyridinecarboxylic acids, as well as more complex
compounds of the quinoline series were used for the synthesis of cationic photosensitizers
[66]. Interaction of N-aminocycloimide 100 with chloroanhydrides of 6-methylnicotinic
and 6-methylpicolinic acids occurred similar to the reaction with isonicotinic acid, and
compounds 128 and 129 were obtained with high yields (Scheme 35). However, an attempt
of the quaternization of the obtained hydrazides showed significant differences in their re-
activities. In the case of the hydrazide of 6-methylnicotinic acid 128, quaternization was
successful, and cationic cycloimide 130 was obtained with a high yield. In contrast, com-
pound 131 was not formed even at multi-hour boiling of 6-methylpicolinic acid hydrazide
129. Apparently, this is due to the formation of the five-membered cycle (Fig. 2) at the ex-
pense of the intramolecular hydrogen bond between nitrogen atoms of the heterocycle and
amide hydrogen.
The data of the 1H NMR spectra, which lack the proton signal doubling, are also in
favour of the rigid fixation of the pyridine ring in the plane of the amide bond.
We observed similar differences in chemical activity also for hydrazides with residues
of quinolinecarboxylic acids (Scheme 36). Condensation of N-aminocycloimide 100 with
chloroanhydrides of 2- and 6-quinolinecarboxylic acids yielded hydrazides 132 and 133, of
which only compound 132 was converted into quaternary base 134.
A logical follow-up of the above-described research into the synthesis of hydrophilic
photosensitizers of the bacteriochlorin series was the development of the routes of synthe-
sizing zwitterionic cycloimides, which have the N-methylpyridine and carboxyl groups in
the lower part of the macrocycle 137. The methyl ester of propionic acid residue in cycloim-
ide 124 was hydrolyzed to introduce a negative charge spatially close to the cationic group-
ing (Scheme 37).
However, the traditional method of hydrolyzing esters in an alkaline medium was
unacceptable due to the lability of the imide exocycle under such conditions. Therefore,
hydrolysis of cycloimide methyl ester was performed in an HCl/dioxane medium in an ar-
gon atmosphere. Another route of producing zwitterionic cycloimides assumed the use of
O O
H H
H H
NH N NH N
A B
100
H N HN H N HN
H H
O N O O N O
CO2Me CO2Me
HN O HN O
132 133
N
C C
O O
H H
H H
NH N NH N
H N HN H N HN
H H
O N O O N O
CO2Me CO2Me
HN O HN O
N
134 135
I
I
N
A: 6-quinolinecarboxylic acid chloroanhydride, pyridine; B: 2-quinolinecarboxylic acid chloroanhydride,

pyridine; C: CH3I, boiling
Scheme 36 Synthesis and quaternization of cycloimides with residues of quinolinecarboxylic acids.
free acid in the synthesis (Scheme 38). For this purpose, N-aminocycloimide of bacterio-
chlorin with the residue of propionic acid 138 was taken as the initial compound. All stages
of the synthesis, including acylation by the chloroanhydride of isonicotinic acid 139 and
the subsequent quaternization of the nitrogen atom 136, were performed at the presence of
an unprotected carboxyl group in the molecule. This route proved to be less successful, and
the total yield did not exceed 18%.
The obtained betaine derivative of cycloimide of bacteriochlorin 137 possessed a
sufficiently high solubility in water, which made it possible to abandon Cremophor usually
O O O
H H H
H H H
NH N NH N NH N
A B
H N HN H N HN ɋ H N HN
H H H
O N O O N O O N O
CO 2Me H 2N CO 2H H 2N CO2 H 2N
O O O
124 136 137

N I N I N
A: (CH3)2CO/HCl, Ar, t; B: KOH/H2O; C: HCl
Scheme 37 Synthesis of zwitterionic cycloimide of the bacteriochlorin series.
O O
H H
H H
NH N NH N
A B
H N HN H N HN
H H
HO2C O HO2C O O
N O N
NH2 HN O
138 139
N
O O
H H
H H
NH N NH N
C
D
H N HN H N HN
H H
HO2C O O O O
N N
HN O O HN O
O
136 137
N x N
Scheme 38 Synthesis of zwitterionic PS.
used for dissolution of hydrophobic compounds and possessing an intrinsic toxicity [67,
68].
Introduction of monosaccharides into bacteriochlorin derivatives also makes it possi-

ble to regulate the amphiphilicity of photosensitizers. Besides, carbohydrate-containing
porphyrins are capable of selective accumulation in neoplastic tissues, by specifically in-
teracting with receptors on the surface of tumour cells [69–71]. We have performed the
synthesis of a new carbohydrate-containing photosensitizer based on cycloimide of bacte-
riochlorin p. As the initial compound, we took the above-described cycloimide containing
the residue of isonicotinic acid 123. The carbohydrate component – 6-dehydroxy-6-
iodo-D-galactopyranoside – was obtained by the scheme including the reaction of D-galac-
tose peracetate with bromoethanol under conditions of acidic catalysis, substitution of the
bromine atom by iodine by boiling with sodium iodide in acetone and the subsequent elim-
ination of protective groupings under the action of 0.1 M solution of sodium methylate. In-
troduction of the carbon fragment into the pigment molecule was performed by way of
quaternization of the nitrogen atom of the pyridine ring by the above-described derivative
of galactose. Glycoconjugate 140, as well as its acetylated analogue were obtained with a
high yield (70%) using 1-O-(2-iodoethyl)2,3,4,6-tetra-O-acetyl-β-D-galactopyranoside.
Due to the positive charge and the hydrophilic carbohydrate residue, the compound ob-
tained is well soluble in water and, as shown by preliminary biological tests, can be consid-
ered as a promising photosensitizer for photodynamic therapy of cancer.
O
H
H
NH N
H N HN
H
O N O
CO 2Me HN
O
OH
N
O
HO O
X
OH
OH
140
Besides the development of PS themselves for PDT of cancer, of great interest in the
recent years is the development of means of delivering PS inside cancer cells to damage-
sensitive targets. Targeted intracellular transport can help to achieve a significant enhance-
ment of photodynamic action, which makes it possible to reduce the concentration of the
drug. Some of such endocyted conjugates are capable of accumulating in lysosomes or pen-
etrating into the nuclei. The group of Prof. A.S. Sobolev (Moscow) develops targeted de-
livery systems, which include the model ligand (insulin, alpha-melanocyte-stimulating
hormone), photosensitizer (chlorin e6, bacteriochlorin p), nuclear localization signal of
SV-40 large T-antigen, and, as a carrier, E. coli hemoglobin-like protein (Fig. 3) [72–75].
Earlier studies by this laboratory have shown that the conjugate of bacteriochlorin p
His - tag D -Tox HMP NLS D - MSH
His-tag – (His)6;
D-Tox – translocation domain of diphtheria toxin;
HMP – E.coli hemoglobin-like protein;
NLS – nuclear localization signal of SV-40 large T-antigen;
α-MSH – alpha-melanocyte-stimulating hormone.
The pocket for porphyrins is in HMP.
Figure 3 Design of a targeted PS delivery system.
with the modular recombinant transporter (MRT) carrying the alpha-melanocyte-stimulat-

ing hormone for its delivery to melanoma cell nuclei has a phytocytotoxicity exceeding this
parameter in free photosensitizers 230 times [76]. Recent works by this group showed [77]
that the binding of bacteriochlorin p with MRT carrying the epidermal growth factor (EGF)
increased the activity of PS of the order of 1000 times (IC50 = 3000 nM for bacteriochlorin
p and IC50 = 4.2 nM for the conjugate) for human carcinoma A431 cells. The authors ex-
plain this increased activity by an enhanced expression of EGF receptors on the surface of
cancer cells.
4 Conclusion
This review has a chemical trend, as it was written by organic chemists. It does not include
numerous works on the biological tests of bacteriochlorophyll a derivatives. It is beyond
doubt that these compounds are in extreme demand by oncologists and specialists of other
medical fields (dermatology, ophthalmology) in view of their prospective use as photosen-
sitizers for photodynamic therapy.
The first works on the synthesis of bacterio-derivatives, in which our group was also
involved, appeared 10–12 years ago; now some of these compounds are at various stages
of clinical tests. All this is indicative of rapid developments in this field of the chemistry of
tetrapyrrole compounds.
It is perfectly evident that for the successful use of PDT in clinical practice health pro-
cessionals should have a series of PS with various therapeutic absorption windows. Deriv-
atives of bacteriochlorophyll a, which absorb in the near IR region of the spectrum, have
their own field of use, where other PS prove little efficient.
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- после 25-ой схемы идет 27; две схемы 33
2 meso-Phenylporphyrins as
Synthetic Models of Natural
Porphyrins: Synthesis and
Modification
A.S. Semeykin, S.A. Syrbu and O.I. Koifman

Ivanovo State University of Chemistry and Technology,
1 F. Engels Prospect, Ivanovo, 153000, Russia
email: koifman@isuct.ru
Methods of synthesis of meso-mono-, di-, three- and tetraphenylporphyrins as well

as methods of introducing the supplement substituents and their further modifica-
tion are considered. Possible ways for fine regulation of the physicochemical pro-
perties of porphyrins are shown.
Introduction
Porphyrins and metalloporphyrins are widespread in nature and are of great biological sig-
nificance. Their most important representatives are chlorophyll, which, as part of the
protein– lipid complex, performs the initial stage of photosynthesis in green plants, and
blood heme, which, in combination with the protein globin, performs the reverse binding
and transportation of oxygen to living cells. Metalloporphyrins are also present in some
enzymes – catalases, peroxidases, etc. At present, interest in porphyrins and their metal
complexes is extremely great. Results obtained in studies of the chemistry and properties
of these compounds are discussed in detail in a number of monographs [1 – 10].
Real ways have been outlined for practical applications of porphyrins and their metal
complexes as efficient catalysts of oxygen electroreduction, oxidation of sulfur dioxide in
electrochemical synthesis of sulfuric acid and fuel hydrogen, polymerization regulators of
acrylates, as photooxidation sensitizers, medical preparations and analytical reagents, semi-
conductors and model compounds in studies of such biological processes as photosynthesis,
46 A.S. Semeykin, S.A. Syrbu and O.I. Koifman
reversible binding of oxygen, enzyme catalysis [6]. The successful development of these
trends depends on reliable methods of synthesis and modification of porphyrins possessing
the required physicochemical properties and resistant to aggressive media and reagents.
The broad use of porphyrins in engineering, technology and medicine is restrained by
the low accessibility of most porphyrins, many of which are obtained with very low yields.
In this connection, of special interest and topicality are issues of the chemistry of synthetic
porphyrins. In this context, one’s attention is attracted by porphyrins containing in meso-
positions aryl substituents, which can be subjected to various chemical transformations. At
present, the properties of natural porphyrins are modelled mainly using their synthetic ana-
logues meso-tetraphenylporphyrins 1, readily obtained by condensation of commercially
available pyrrole with benzaldehydes. However, in some cases they are not very con-
venient, as, unlike natural porphyrins, they have no alkyl or pseudoalkyl substituents in
β-positions of the porphyrin cycle, whereas meso-positions are substituted. On the other
hand, sufficiently accessible octaalkylporphyrins 2 are not always convenient, either, as they
have no active groups, which can be changed to impart them with various physicochemical
properties. Therefore, of great interest are porphyrins, which combine the advantages of
these two classes, such, for instance, as 5,15-diaryloctaalkylporphyrins 3 and 5-aryl-
octaalkylporphyrins 4. These porphyrins are produced mainly by condensation of α-unsub-
stituted linear derivatives of pyrroles with benzaldehydes.
Ar R R R Ar R R Ar R
R R R R R R
NH N NH N NH N NH N
Ar Ar
N HN N HN N HN N HN
R R R R R R
Ar R R R R R Ar R
1 2 3 4
1 Synthesis of meso(5,10,15,20)-tetrasubstituted porphyrins

meso-Tetrasubstituted porphyrins were first obtained by Rotmund, who found [11] that
meso-substituted porphyrins 5 were formed in the interaction of pyrrole with aldehydes
(Scheme 1). Condensation of acetaldehyde with pyrrole leads, with a low yield, to meso-
tetramethylporphyrin (5, R=Me), and of pyrrole with formaldehyde to porphin (V, R=H)
with a negligible yield of 0.03% [12]. In the subsequent works by Rotmund [12, 13], the
yield of porphyrins was significantly improved due to the performance of the reaction in
pyridine in a sealed ampoule at a high temperature (140–240°C). Condensation of pyrrole
with benzaldehyde yielded meso-tetraphenylporphin (5, R=Ph, H2TPP) [13], which is
formed with a sufficiently high yield and is readily isolated from the reaction mixture [13].
Later, the works [14, 15] showed the yield of H2TPP to increase upon addition of zinc
acetate to the reaction mixture. The obtained zinc complex of tetraphenylporphin (ZnTPP)
is then transferred by the action of a mineral acid to free porphyrin. However, the yield of
H2TPP even under these conditions, optimal for the condensation reaction in pyridine, does
not exceed 18%.
At present, 2,4,6-trimethylpyridine (collidine) [16, 17] and quinoline are also used as
a reaction medium besides pyridine; they boil at a higher temperature than pyridine (171
meso-Phenylporphyrins as Synthetic Models of Natural Porphyrins 47
o NH N
140-240 C
+ RCHO R R
N N HN
H
R
5
Scheme 1
and 237 against 115°C, respectively), which makes it possible to perform the condensation
reaction at an atmospheric pressure and in the presence of oxygen of the air as an oxidant.
By the Rotmund’s method, the reaction is performed at high concentrations of the re-
acting substances (more than 1 M), but a comparatively low yield of meso-substituted por-
phyrins restricts its application. Methods of high-temperature synthesis have been modified
to date, which sometimes makes it possible to completely avoid the use of the solvent. Thus,
the reaction of a mixture of pyrrole and benzaldehyde in the presence of a metal salt (in the
absence of a solvent) in a sealed ampoule at 150– 250°C produces H2TFP with a yield ex-
ceeding 50% [18]. Treatment of a mixture of pyrrole, benzaldehyde and silica gel or ceolite
in a microwave oven leads after a chromatographic purification to H2TFP with a yield
reaching 9% [19–21]. Introduction of pyrrole into the ampoule containing aldehyde in the
gas phase (200–250°C) in a air atmosphere leads to the production of some meso-substi-
tuted porphyrins with yields of up to 23% [22].
Recently [23] tetraphenylporphins were reported to be obtained with yields of more
than 20% in a condensation reaction in 2,4,6-trichlorophenol during the boiling in the pres-
ence of air at reagent concentrations of about 0.6 mol/kg. It is difficult to assign this reaction
to the Adler’s method (see further) due to the small acidity of this phenol (pKa = 6.1). Be-
sides, it is of interest that manganese complexes of many even di-ortho-substituted tetraphe-
nylporphyrins are obtained with yields exceeding 50%, which implies the presence of a
template effect on the Mg(II) ion.
It should be noted that synthesis of meso-substituted porphyrins in basic media, unde-
servedly forgotten in the recent years due to the discovery of acid-catalysis methods, has
its specific areas of application, such, for instance, as synthesis of meso-phenylporphyrins
having in phenyl rings groups labile in an acidic medium or containing in meso-positions
some heterocyclic residues (e.g., furanic or pyrrolic).
Based on the works [13, 24] (their authors obtained with a small yield H2TPP at a pro-
longed boiling of a mixture of pyrrole and benzaldehyde in a methanol– pyridine medium),
Adler [25], when studying the effect of various solvents, found that the condensation reac-
tion is catalyzed by acids more efficiently than by bases. This conclusion was confirmed in
the work by Treibs [26]. Adler [25] determined that when performing the reaction in
aerated, boiling, acid-containing organic solvents the yield of H2TPP by the spectrophoto-
metric estimate reached 40%. At present, the condensation reaction of pyrrole with alde-
hydes (Scheme 2) in acid-containing medium in the presence of the air is one of the major
methods of producing meso-substituted porphyrins (the Adler’s method). The reagents
mainly used as acidic solvents are acetic acid [25, 27], propionic acid [28, 29]; mixed sol-
vents: pyridine-acetic acid [26], benzene-chloroacetic acid [25], toluene-p-toluene sulfo
NH N
air
+ RCHO R R
H+
N N HN
H
R
5
Scheme 2
acid [30], xylene-chloroacetic acid [31– 33] and some others. The reaction is usually per-
formed at the boiling temperature of the solvent, sometimes at the passage of air through
the reaction mixture [26]. It has been found [34] that the yield of porphyrins depends on the
temperature of the reaction medium, and the largest yield is obtained at temperatures close
to 140°C. At lower temperatures, the porphyrin formation rate is small, and at higher tem-
peratures the rate of oxidation of porphyrins produced is high. An optimal concentration of
the reagents in the reaction medium is of the order of 0.2– 0.4 mol/l.
Finally, there are new modifications of the Adler’s method for the synthesis of meso-
substituted porphyrins: condensation of pyrrole with aldehydes in a medium of dimethyl-
formamide (dimethylsulfoxide)-aluminium trichloride (yield of H2TFP, 30%) [35], or in
propionic acid with microwave irradiation (yield of meso-tetraarylporphyrins, 20–43%)
[36]. The reaction of anisic aldehyde with pyrrole at 120°C in propionic acid containing
30% nitrobenzene, which in this case is an oxidant, made it possible to obtain a respective
porphyrin with the yield of 45% [37]. The yields of 5–20% were observed for a number of
other arylaldehydes [37, 38].
R HH
H
H
NH N
R R
N HN
R
6
It should be noted that under conditions of the reaction of acidic condensation, respec-
tive chlorins 6 are also formed besides porphyrins, and in some cases chlorins become the
main reaction products [39]. However, they can easily be transformed into respective por-
phyrins by the treatment with benzoquinone derivatives: para-chloranil (p-CA) or 2,3-
dichloro-5,6-dicyanobenzoquinone-1,4 (DDQ) [40–43].
The use of a mixture of valeric acid and nitrobenzene at 160°C for the condensation
of pyrrole with benzaldehydes makes it possible to significantly increase the yield of por-
phyrins, which contain no chlorins [44].
Isolation of porphyrins causes no problems, when they are crystallized during the
H R R
CF3COOH
or NH HN NH N
BF3etherate R H DDQ R
+ R-CHO R
N CH Cl , 25oC H
2 2 NH HN
R
N HN
H
R H R
7 5
Scheme 3
cooling of the reaction mass in a sufficiently pure form [26, 29]; in other cases, the solvent
is driven off under vacuum [45] or with steam [31] and the residue is chromatographed. In
the case of using acids as solvents the reaction mixture can be neutralized by a solution of
ammonia, the residue be filtered and porphyrin also be isolated by chromatography on a
suitable sorbent [46].
The Adler’s method proved itself to be excellent for preparative-scale synthesis of por-
phyrins from aldehydes, which are relatively stable. The possibility to obtain quickly and
easily an approximately 20% yield of such porphyrins at reagent concentrations of up to
0.4 M makes the Adler’s method sufficiently convenient. The use of the Adler’s method is
limited in syntheses of meso-substituted porphyrins from aldehydes having substituents,
which do not withstand the action of acids at high temperatures, many 2,6-disubstituted ben-
zaldehydes and many aliphatic aldehydes.
A comparatively recently developed new method of synthesizing meso-substituted
porphyrins under mild conditions [47, 48] consists in the condensation of pyrrole with al-
dehydes in chloroform or methylene chloride in the presence of trifluoroacetic acid or boron
trifluoride etherate in an inert atmosphere at room temperature to respective porphyrinogen
7 with subsequent oxidation of the reaction mixture by a stoichiometric amount of DDQ or
p-CA (the Lindsey method) (Scheme 3). There is an example, where oxidation of porphy-
rinogen 7 is performed by hydrogen peroxide in acetic acid [49]. Besides the chemical
methods, meso-alkylsubstituted porphyrinogens, which are more resistant to oxidation, can
be converted to porphyrins photochemically [37].
The reaction, as found in [47], is sensitive to reagent concentrations. The highest yields
of H2TPP (35–40%) are obtained in the reaction of 10 mM benzaldehyde and pyrrole each,
and the yield decreases approximately two times at reagent concentrations of 100 mM each
and 1 mM each. The decrease of the yield at a higher concentration of the reagents can be
partially reduced by increasing the amounts of acid catalyst. For instance, at 100 mM ben-
zaldehyde and pyrrole each, the yield of H2TPP was 23% with 1 mM BF3 etherate; 30%,
with 3.2 mM and 29% with 10 mM BF3 etherate. These magnitudes are still lower than the
35– 40% yield obtained at reagent concentrations of 10 mM each in CH2Cl2 with BF3 ether-
ate or trifluoroacetic acid as a catalyst [50]. The reaction is also sensitive to the concentra-
tion of acid catalyst: for the reagent concentrations of 10 mM each, BF3 etherate is efficient
at a concentration of 1 mM, whereas trifluoroacetic acid is required at a much higher con-
centration of 20– 50 mM [47]. Addition of sodium or ammonium chlorides was found to
significantly increase the yield of porphyrins [51, 52].
By the example of tetra(mesityl)porphin (5 R = 2,4,6-trimethylphenyl; H2TMP), it has
been shown [53– 55] that, for synthesis of tetraphenylporphyrins containing electron-donor
substituents in both ortho-positions of the phenyl rings, at the stage of condensation it is re-
quired to use BF3 etherate as a catalyst in the presence of a cocatalyst, 0.75% ethanol. This
method was improved to obtain gram amounts of H2TMP [56, 57]. Cocatalysts in the syn-
thesis of H2TMP and similar porphyrins can be, besides ethanol, also ethylene glycol,
2-methoxyethanol [55] or methanol [58]. Syntheses of tetraphenylporphyrins containing
electron-acceptor substituents in both ortho-positions of the phenyl rings do not require the
use of cocatalyst. Application of 2,2-dimethoxypropane as a dehydrating agent also leads
to an increase of the yield of H2TMP [58].
A further improvement of the method was to increase the reagent concentrations and
to use oxygen of the air as an oxidant of porphyrinogen in the catalysis by iron phthalocy-
anine and p-CA [51]. This slightly decreases the yield of the end product, but makes it pos-
sible to use the method in large-scale syntheses [59].
Mild conditions of the reaction at the stages of condensation and oxidation enable the
use of the Lindsey method for a broad range of aldehydes. This method is of special signif-
icance for synthesis of spatially hindered porphyrins [54, 57, 60] and meso-alkylporphyrins
[61, 62], which are obtained by other methods with low yields. The yield of porphyrins can
reach 50% depending on the aldehyde. At present, this is the most widely used method of
synthesizing meso-substituted porphyrins.
There are several modifications of this method: use of pyrrole with aldehydes of clays,
such as monomorillonite and the like as condensation catalysts [20, 21, 63, 64]. The activity
of clays is ascribed to their porosity but not acidity, and their efficiency as condensation
catalysts is associated with stabilization of porphyrinogens in clay nanotubes [21, 65]. For
synthesis of porphyrins with polar groups, which are obtained by the Lindsey method with
low yield [47, 66], the method of micellar synthesis was proposed [67], which consists in
conducting the condensation reaction in hydrochloric-acid solution of dodecylsulfate used
as a surfactant, followed by oxidation of the reaction mixture by a solution of p-CA in tet-
rahydrofurane. This method makes it possible to obtain tetraphenylporphins having car-
boxy, hydroxy, acetamido, ether and ester groups in the phenyl rings.
R R
R
NH N NH N
N HN N HN
R R R
R R
D,E,E,E- 8 D,E,D,E- 8
R
R
R R R
NH N NH N
N HN N HN
R
R R
D,D,E,E- 8 D,D,D,D- 8
The Lindsey method has the mildest reaction conditions. The yields are in most cases
higher than those for the Adler’s method, though the reaction at reagent concentrations from
0.01 M up to 0.1 M requires large amounts of solvent to be removed. The method is appli-
cable in the case of aldehydes having substituents unstable to the action of acids, 2,6-dis-
ubstituted benzaldehydes, aliphatic aldehydes and little-accessible expensive aldehydes
(for production of porphyrins from which high yields are of great importance).
meso-Tetraphenylporphyrins, having in one of ortho-positions of the phenyl rings
bulky substituents, are formed as a statistical mixture of atropisomers 8 owing to the impos-
sibility of the turn of teh phenyl rings; in some cases, when substituents are polar, the mix-
ture can be separated chromatographically [68, 69].
The synthesis mechanism of meso-substituted porphyrins in an acidic medium
(Scheme 4) includes the interaction of pyrrole by the carbonyl group of aldehyde to form
pyrrylcarbinol 9. Further, it is protonated, yielding stabilized carbocation 10, which then
H+ H+ +
+ RCHO CHOH CH + CH
N N N N
R R R
H H H H
9 10
pyrrole RCHO
CHOH H CH OH
N H R N H+ N H R N N
R R n
H H H H H
11 12
H R H R H R H H
R R
N
H
NH HN NH HN NH HN
R H RCHOR H R H pyrrole NH HN
H R H R H R H H
NH HN NH HN NH NH HN
R R
H H NH
R OH R OH
1. cyclization
2. oxidation
R R R
R R
N
NH N H
NH HN NH N
N N
R R R R R R
N HN N HN N R R
NH NH HN
R R
13 5 14 15
Scheme 4
reacts to the additional molecule of pyrrole to form meso-substituted dipyrrolylmethane 11.

Further the reaction is repeated with the buildup of the polypyrrole up to 12, which is cy-
clized at n = 4 to porphyrinogen 7, oxidized further by oxygen of the air (Adler’s method)
or by benzoquinone derivatives (the Lindsey method) to meso-tetrasubstituted porphyrin 5.
Using the Alder’s method, the linear polypyrrole chain can be also partially oxidized to the
closure into the cycle. Depending on the reaction conditions, relative concentrations of the
reagents and catalyst used the end result of the cyclization and subsequent oxidation can
differ. Thus, conditions were determined, at which, besides porphyrins 5, also corroles 13
can be formed (excess of pyrrole, yield up to 10%) [70], N-confused porphyrins 14 (yield
up to 10%) [71, 72] and tetraphenylsapphirins 15 (yield up to 1%) [73, 74] (Scheme 4).
The condensation reaction of pyrrole with aldehydes is also used for the synthesis of
compound structures containing porphyrin fragments. Thus, in condensation of dialde-
hydes, in which phenyl rings via ortho-positions are connected by a sufficiently long di-
oxymethylene chain [R = –O(CH2)nO–, n ≥ 4], with pyrrole, a mixture of three isomeric
belted porphyrins 16–18 is formed; herewith, the most strained porphyrin 16 is not formed
if the chain is too small (n = 4) [75–77]. At an even smaller chain length (n = 2, 3 and 4 for
meta-positions), under the condensation conditions, dimeric sandwiched porphyrins 19 are
formed [78, 79].
R R O R R O
O
O O O O O
NH N NH N NH N
N HN N HN N HN
O
O
R R
O O
16 17 18
O
R O R
O
O
N
HN N
HN
NH
N NH
N
O
O
R O R
O
19
Condensation of various-structure tetraaldehydes with pyrrole yielded the so called

capped porphyrins 20– 23 [33, 80– 85]. Often, syntheses of this kind are made using
high-dilution methods [75, 76].
All listed porphyrins at small lengths of connecting bridges have a distortion of the
plane structure of the porphyrin macrocycle, caused by the tension of the bonds between
phenyl rings in meso-positions and meso-substituted macrocycle. Moreover, sandwiched
dimers 19 having an even number of methylene links (n = 2, 4) are distorted more than their
analogues with an odd number (n = 3) [78].
It should be noted that such porphyrins also have hindrances in conducting the reac-
tions by the central atoms of nitrogen on one or both sides of the macrocycle, i.e., they pos-
sess the properties of both distorted and hindered porphyrins.
An increase of the length of connecting bridges to greater than an extreme value leads
to the disappearance of the distortion of the porphyrin macrocycle due to the elimination of
R R R R
R R
R R
R R
R R
NH N NH N
NH N
N HN N HN
N HN
20 21 22
NH NH
O O
HO
O
O O
NH N
N HN
O
23
the strain from the phenyl–macrocycle link [86].

By condensation of mono-4-formylphenyltriphenylporphyrins with pyrrole, pentam-
eric porphyrins 24 were synthesized [87, 88].
R
R
X
R
NH N NH N
X X X=
N HN N HN
R
X
R
R
24
The adduced exotic structures are usually obtained with yields not exceeding 10%;
however, synthesis of respective polyaldehydes presents no large problems in most cases,
which makes such porphyrins interesting in many biological and catalytic studies of por-
phyrin systems.
The first intermediate compounds in the condensation of pyrrole with aldehydes are
pyrrylcarbinols 9. Therefore, of great interest are studies of the possibility of using these
compounds for synthesis of meso-substituted porphyrins.
Three methods of synthesis of pyrrylcarbinols have been developed to date (Scheme
I RCOCl
N
MgX R LiAlH4 RLi H
(NaBH4) CHR III
N N N
RCON(CH3)2 O OH O
II H H H
N POCl3 25 9 26
H
Scheme 5
5). The first two methods are based on the initial production of 2-ketopyrroles: the reaction
of the pyrrole Grignard reagent with chloroanhydride of the acid yields 2-ketopyrrole 25
and 3-ketopyrrole with yields of ~60 and 15%, respectively (I) [89]; an alternative method
of producing 2-ketopyrroles consists in the interaction of N,N-dimethylamides with pyrrole
according to Wilsmeyer in the presence of phosphorus oxychrolide (II) [90– 92]. 2-Keto-
pyrroles 25 are then reduced by LiAlH4 or NaBH4 to pyrrylcarbinols 9. The direct one-stage
method of producing pyrrylcarbinols is performed by the reaction of pyrrole-2-carboxyal-
dehydes 26 with lithium or magnesium organic compounds (III) [90– 92]. The last two
methods have no drawback of the first method – nonselectivity of substitution in acylation
of pyrrole. Besides, it has been found [93, 94] that pyrrylcarbinols with perfluoroalkyl res-
idues are easily formed in condensation of hydrated perfluoroaldehydes with pyrrole in the
presence of alkali.
Pyrrylcarbinols 9 are self-condensed, yielding meso-substituted porphyrins under con-
ditions of Rotmund’s reaction (Scheme 6) [95]. The yields of porphyrins in this reaction are
comparable with those for condensation of pyrrole with aldehydes. Besides, coloured inter-
mediate compounds observed in condensation of pyrrole with aldehyde, are the same as in
self-condensation of respective pyrrylcarbinol. These results prove that pyrrylcarbinols 9
are indeed intermediate products in the condensation reaction in an acidic medium [95].
NH N
CHR H+
N R R
OH [O]
H N HN
R
9 5
Scheme 6
meso-Substituted porphyrins are usually synthesized under conditions similar to those

of the Adler’s reaction [90-92, 97, 96]; in some cases, the yield of porphyrins is observed
to be significantly increased upon addition of zinc acetate to the reaction mixture [98].
Sometimes, the two-stage Lindsey method is also employed [93, 94, 99]. The method using
pyrrylcarbinols is mainly applied to produce meso-alkylporphyrins and hindered porphy-
rins. Of interest is the fact that bulky pyrrylcarbinols obtained from 2-isobutyryl or
2-pyvaloylpyrrole can not be converted to respective porphyrins [98].
1.1 Synthesis of meso-tetra-β-octasubstituted porphyrins

Besides pyrrole, its 3,4-disubstituted derivatives can enter into the condensation reaction
with aldehydes to form, after oxidation, meso-tetra-β-octasubstituted porphyrins possessing
a distorted porphyrin cycle owing to the steric interaction of substituents in adjacent meso-
and β-positions.
The properties of porphyrins are mainly determined by two factors. One of them, elec-
tronic, is due to the occurrence of a broad aromatic π-conjugation contour embracing the
macrocycle. The other, geometric, is represented by a plane tetrapyrrole system possessing
a rigidity and relative stability to strain.
At present, however, there are numerous data, which show that the porphyrin macro-
cycle is sufficiently flexible and is capable of the existence in non-plane conformations
[100– 102]. These properties of porphyrin molecules are extensively studied, as the distor-
tion of the porphyrin cycle can play a significant role in biological photosynthetic and redox
systems [103–105]; besides, the conformation distortions of the porphyrin chromophore
can serve as a tool of fine tuning of its physicochemical properties.
We should note the distinction of hindered porphyrins from distorted ones. The pecu-
liarity of the former is in shielding the central cavity of the porphyrin molecule by bulky
substituents from attacks of various reagents. This spatial hindrance strongly affects the re-
activity of porphyrins, but has a minor effect on their physical properties. In contrast to these
compounds, distorted porphyrins have a non-plane deformed macroring, owing to which
fact both the chemical and physical characteristics of these porphyrins significantly change
as compared with analogues possessing the plane structure of the porphyrin core.
Distortion of the porphyrin cycle can in an ideal form produce two extreme – saddled
and ruffled – conformations [102].
The data of X-ray diffraction studies show that most symmetric distorted porphyrins
and their complexes have a saddled (or close to it) conformation, and only in sufficiently
rare cases, for some metal complexes, the pure ruffled conformation is realized [102, 106].
_ + + + +
0
_ _ _
+
NH N NH N
_ _ 0 0 _
+
0 _ 0 0 0
+ N HN
N HN _ _
+ _
0 _ + + +
+
saddled ruffled
The most known and basic method of distortion of the porphyrin cycle is introduction
of substituents to its periphery to adjacent meso- and β-positions; the distortion rises with
their number and size increasing.
Thus, in condensation of benzaldehyde with pyrrole and its 3,4-disubstituted analogue
under the action of boron trifluoride etherate, a mixture of porphyrinogens was obtained;
condensation of the mixture by DDQ led to six possible porphyrins 27–32 (Scheme 7)
[107– 109]. The X-ray diffraction analysis of specimens of these porphyrins showed [107]
the deformation of the macrocycle to grow from almost plane tetraphenylporphin 27 (A =
Ph) to strongly distorted dodeca-substituted porphyrin 32 (A = Ph, B = Et, Ph).
Condensation of 3,4-diphenylpyrrole with formaldehyde and benzaldehyde under the
32 (A = Ph, B = Et, Ph).

B B
1. BF3 etherate
ACHO + + 27-32
N N 2. DDQ
H H
A B A B A
B B
NH N NH N NH N
A A A A A A
N HN N HN N HN
B
A A A B
27 28 29
B A B B A B B A B
B B B B B B
NH N NH N NH N
A A A A A A
N HN N HN N HN
B B B
A B A B A B
30 31 32
Scheme 7
action of HBr in ethanol, followed by oxidation of the mixture of porphyrinogens by DDQ

in boiling toluene, produced six possible porphyrins 32– 37 (A = B = Ph) (Scheme 8) [110]
with the deformation increasing from plane β-octaphenylporphin 33 to dodecaphenylpor-
phyrin 32.
Similar results were obtained in condensation of 3,4-disubstituted pyrroles and their
2-hydroxymethyl derivatives 38 with benzaldehydes in methanol containing HBr followed
by the oxidation of the mixture of porphyrinogens by p-CA in THF (Scheme 8) [111].
Of interest is the fact that, proportionally to the growth of distortion of the porphyrin
cycle, the batochromic shift of all bands in absorption spectra of these porphyrins also rises
[107, 109– 111].
Scheme 8
B B
1. HBr - EtOH
ACHO + CH2O + 32–37
N 2. DDQ
H
or
B B B B
1. HBr - EtOH
ACHO + + CH2OH 32–37
2. DDQ
N N
H H
38
B B B A B B A B
B B B B B B
NH N NH N NH N
N HN N HN N HN
B B B B B B
B B B B B A B
33 34 35
B A B B A B B A B
B B B B B B
NH N NH N NH N
A A A A A
N HN N HN N HN
B B B B B B
B B B B B A B
36 37 32
The most strongly distorted dodeca-substituted porphyrins 32 are obtained by conden-

sation of aldehydes with 3,4-disubstituted pyrroles; methods of their synthesis differ little
from those for tetraphenylporphins (Scheme 9). In the main, these are two known modifi-
cations of the method: condensation of pyrrole with aldehyde in organic acid-containing
boiling solvent in the presence of oxygen of the air (Adler’s method) [27, 112, 113], or con-
densation under the action of a catalyst (acid, boron trifluoride etherate) under mild condi-
tions to porphyrinogen followed by its oxidation without isolation (or with isolation) to
porphyrin by benzoquinone derivatives (the Lindsey method) [114– 120]. The high resis-
tance of porphyrinogens to oxidation makes it possible to isolate these compounds in pure
form and then oxidize them to porphyrins.
Scheme 9
B A B
B B
B B
[O] NH N
+ ACHO A A
N H+
N HN
H
B B
B A B
32
It has been found [121–123] that the strong distortion of the porphyrin cycle also
results from the occurrence of bulky substituents, such, e.g., as tert-butyl or adamantyl in
meso-positions in the absence of substituents in β-positions of the cycle.
In production of distorted β-alkylsubstituted porphyrins 32 (B = Alk), it is better to use
the condensation in acid-containing alcoholic solvents followed by the isolation of porphy-
rinogen and its subsequent oxidation by DDQ in THF. For synthesis of β-phenylporphyrins
32 (B = Ar), the condensation is better to be conducted in boiling acetic acid followed by
oxidation of porphyrinogen or without its oxidation in boiling with DDQ, or else with
isolation and subsequent oxidation in boiling toluene also using DDQ. The use of the Lind-
sey method [120, 124] is justified in the synthesis of porphyrins 32 having o-disubstituted
phenyl rings in meso-positions.
It should be noted that condensation between benzaldehyde and pyrrole nonsymmet-
rically substituted in β-positions produces a statistical mixture of four randomeric porphy-
rins (39– 42).
B C A B C B B C A A C A
A B A A A B B B
NH N NH N NH N NH N
C C C C C C C C
N HN N HN N HN N HN
B A A A A A A A
A C B B C B B C B B C B
39 (type I) 40 (type II) 41 (type III) 42 (type IV)
Interesting is the fact that condensation of 3,4-dialkylpyrrole with alkylaldehydes fol-

lowed by oxidation of the reaction mixture by benzoquinone does not lead to distorted dode-
ca-alkylporphyrins, but is stopped at the stage of respective porphodimethenes 43 [119]. In
this case, distortion of the macrocycle skeleton is so great that the oxidation potential of ox-
idants used is not sufficient for its aromatization. On the contrary, practically nondistorted
porphyrins 44, 45 are obtained with high yields [119, 125].
,
B H A B A
R
B B
NH N NH N R
A A NH N
A A
N HN N HN
B B N HN
R
B A H B A
A,B = Alk A = Alk R
43 44 45
1.2 Synthesis of nonsymmetric meso-tetrasubstituted porphyrins

Condensation of aldehyde with pyrrole makes it possible to obtain porphyrin having four
identical meso-substituents. However, there are many applications, which require various
substituents in meso-positions of the porphyrin cycle. One of the simplest approaches to this
goal is the so called mixed-aldehyde condensation (Scheme 10). The reaction of pyrrole
with a mixture of aldehydes makes it possible to obtain a mixture of six porphyrins 46– 51,
which can be separated by means of thin-layer chromatography [126], and in the case of a
polar group in one of the aldehydes, using column chromatography [127].
The expected ratio of porphyrins in mixed-aldehyde condensation is set by the binom-
inal distribution. At an aldehyde ratio of 1:1, the distributions are as follows: A4, 6.25%;
A3B, 25%; cis-A2B2, 25%; trans-A2B2, 12.5%; AB3, 25%; B4, 6.25%. This result assumes
the equal reactivity of both aldehydes at all stages of the porphyrin formation reaction. A3B
porphyrins are obtained the most often in mixed-aldehyde condensation. The yield of A3B
porphyrin is maximal at a ratio of the reacting aldehydes 3:1: A4, 31.64%; A3B, 42.19%;
cis-A2B2, 14.06%; trans-A2B2, 7.03%; AB3, 4.69%; B4, 0.39%. A larger content of alde-
hyde A increases the yield of A4 porphyrin at a decrease of the absolute amount of A3B por-
Scheme 10
+ ACHO + BCHO 46–51

N
H
A B B
NH N NH N NH N
A A A A A B
N HN N HN N HN
A A A
A4 A3B cis-Ⱥ2ȼ2 (ȺȺȼȼ)
46 47 48
B B B
NH N NH N NH N
A A A B B B
N HN N HN N HN
B B B
trans-Ⱥ2ȼ2 (ȺȼȺȼ) Ⱥȼ3 ȼ4
49 50 51
phyrin. However, the ratio of the reacting aldehydes, which gives the highest isolated yield
of A3B porphyrin, depends on the real reactivities of these aldehydes, as well as on the ease
of separation of the porphyrin mixture [128]. When a mixture of strongly-polar hard-of-
access aldehyde and low-polar aldehyde is used to produce AB3 porphyrins, it is sometimes
more advantageous to use a large excess of low-polar aldehyde, so that the strongly-polar
one reacts more completely. Herewith, symmetrically substituted B4 porphyrin is mainly
produced, which is, however, easily separable chromatographically from the required AB3
porphyrin [129–131]. Using this method, a number of dimeric porphyrins linked via meso-
positions by various spacers was synthesized [131–136].
In practice, isolated yields of AB3 porphyrins in Adler’s method are mainly at the level
of 5%, with an approximate total yield of the porphyrin mixture 20% and a statistical dis-
tribution of the mixture.
Reactivities of aldehydes in mixed-aldehyde condensation sharply differ in most cas-
es; for this reason, namely the Lindsey method is the most suitable for such syntheses, or a
modified Adler method, in which the first stage of the reaction is performed in the absence
of oxygen of the air to avoid oxidation of porphyrinogen formed by more reactive aldehyde
and to achieve an equilibrium between porphyrinogens, which is set by the ratio of the ini-
tial aldehydes.
Sometimes, the reaction is conducted with two aldehydes of close reactivities or with
aldehydes stable under the synthesis conditions. Further, by modification of substituents,

they are imparted with required polarity or structure [20, 97, 137].
Of interest is the fact that aldehydes, which do not yield meso-tetrasubstituted porphy-
rins, often enter into mixed-aldehyde condensation as one of the components [95, 138, 139].
Mixed-aldehyde condensations are usually restricted by the use of two reagents.
Three-component condensations are very rare. Condensation with three different aldehydes
and pyrrole made it possible to obtained a porphyrin mixture, from which 5-(2-hydroxyphe-
nyl)-15-(2-nitrophenyl)-10,20-di-p-tolylporphin was isolated with the yield of less than
0.1% [95]. Joint condensation of benzaldehyde, terephthalaldehyde and 4-hydroxybenzal-
dehyde with pyrrole made accessible monohydroxyporphyrin dimer isolated as a mixture
of isomers with the 0.7% yield [140].
By condensation of a mixture of trialdehyde 52 and aldehydes 53 with pyrrole by the
Adler’s method, capped porphyrins with additional covalently bound extra ligand 54 was
synthesized, though with low yields, but in a minimal number of stages (Scheme 11) [141].
Scheme 11
O O
O O
O O
O N O
O + O
O H O
O O
O OHC R air O R
+ EtCOOH NH N
CHO OHC
N HN
CHO N O O
O O N
52 53 54
Mixed-pyrrole condensations can also be performed. Thus, at a ratio of 3:1, a mixture

of pyrrole and pyrrole containing a carboxyl group, which is attached to it via polymethyl-
ene bridge 55, reacts to benzaldehyde (or p-toluyl aldehyde) in boiling propionic acid. The
further treatment of the reaction mixture makes it possible to obtain porphyrin having one
β-substituted pyrrole residue, which can provide for the attachment of this porphyrin to
polymer 56 (Scheme 12) [142]. Mixed-pyrrole condensations are used much more rarely
Scheme 12
H3C CH3
(CH2)nCO 2H CHO
N
(CH2)nCO 2H
3 + + 4 NH HN
N N
H H N
CH3
H3C CH3
55 56
than mixed-aldehyde ones, undoubtedly, due to the lower accessibility of substituted

pyrroles.
A more promising synthesis of nonsymmetric meso-tetrasubstituted porphyrins is the
method, which makes use of preliminarily obtained meso-substituted dipyrrolylmethanes
11. Condensation of unsubstituted pyrrole with aldehydes leads to a mixture of oligomeric
products: dipyrrolylmethanes 11, tripyrranes 57, bilanes 12 (n = 4), cyclic porphyrinogens
7 and higher linear and cyclic oligomers. However, conducting this reaction at a significant
excess of pyrrole makes it possible to stop it at the stage of meso-substituted dipyrrolyl-
methanes 11 (Scheme 13). A large number of methods was reported for direct condensation
leading to dipyrrolylmethanes [93, 143– 154]. As condensation catalysts, use is usually
Scheme 13
H R
NH R NH HN
TPAA or R
RCHO + NH +
BF3 etherate H H NH
excess NH
11 57
Ⱥ ɛ ɛ
made of trifluoroacetic acid or boron trifluoride etherate; the reaction is run at room tem-
perature in a solvent or without it. Removal of excess pyrrole under vacuum yields raw
dipyrrolylmethane, which is recrystallized [147], chromatographed [147] or distilled [146,
155] to obtain a pure product. The isolation of respective tripyrrane 57 from nonpurified
dipyrrolylmethane has been also reported [155].
In a similar way, using excess pyrrole, meso-substituted dipyrrolylmethane 11 was
synthesized from pyrrylcarbinol 9 with the yield of 95% (Scheme 14) at the catalysis by
trifluoroacetic acid or BF3 etherate [147].
Scheme 14
R NH
CHOH + NH PhCHO
N TPAA or
H R H NH
excess BF3 etherate
9 11
Condensation of meso-substituted dipyrrolylmethane 11 with aldehyde leads to

ABAB(trans)-type meso-tetrasubstituted porphyrins 58 [156–158]; the use of two alde-
hydes in condensation makes it possible to obtain ABAC-type porphyrin 59; however, in a
mixture with two porphyrins ABAB and ACAC [149, 159– 161], if one of aldehydes has a
substituent similar to dipyrrolylmethane one, the formed product is mainly BA3-type por-
phyrin 60 [162– 164] (Scheme 15).
In condensation of a mixture of two meso-substituted dipyrrolylmethanes with alde-
hyde similar to one of the meso-substituents of dipyrrolylmethane, the product obtained
is mainly BA3-type porphyrin 60, and using aldehyde with a substituent not similar to meso-
Scheme 15
B B
NH N NH NH N
BCHO A BCHO
A A A A
N HN CCHO H
NH N HN
C B
ABAC + ABAB+ACAC ABAB
59 11 58
ACHO
BCHO
B
NH N
A A
N HN
A
BA3 + ABAB+A4
60
substituents of dipyrrolylmethanes, ACBC porphyrin 61. However, in both cases other por-
phyrins are also formed (Scheme 16); their chromatographic separation is possible only in
the occurrence of polar groups in substituents.
Scheme 16
C A
NH N NH HN NH N
CCHO A B ACHO
A B + A B
H NH HN H N HN
N HN
C A
ACBC + ACAC+BCBC BA3 + ABAB + A4
61 60
It should be noted that for such kind of syntheses, the suitable method is only the
low-temperature Lindsey method, as acidolysis of meso-substituted dipyrrolylmethanes in
acid media at high temperatures and regroupings leads in the subsequent oxidation to the
formation of a compound mixture of porphyrins (Scheme 17).
However, in [165] it was shown that during the condensation in propionic acid in the
presence of zinc acetate only trans-porphyrin 58 without regrouping was formed, which is,
possibly, due to oxidation processes prior to the closure of the cycle. This reaction conduct-
ed by the two-stage Lindsey method forms a mixture of isomeric porphyrins, though with
a large yield [162, 165]
Scheme 17
NH pyrrole B NH
A H+ + BCHO +
CH + CH + H
H NH N N H+ N H NH
A B
H H H
A number of studies [157, 158] have shown dipyrrolylmethanes having spatially hin-
dered or perfluoroalkyl substituents in meso-positions to be the most resistant to acidolysis. .
The condensation reactions using meso-substituted dipyrrolylmethylcarbinols 62 in-
stead of a mixture of aldehyde and dipyrrolylmethane make it possible to avoid acidolysis
due to the impossibility of forming nonstabilized dicarbocation (Scheme 18).
Scheme 18
B B
+
CHOH CH
NH NH
A H+ A H+ + +
HC CH +
H H N N
NH NH A B
H H
62
Scheme 19 presents some possible routes of using meso-substituted dipyrrolylcarbi-

nols 62 and dipyrrolyldimethylcarbinols 63 for synthesis of nonsymmetrically meso-tetra-
substituted porphyrins of various types. Of special interest is the latter route, which makes
it possible to obtain totally nonsymmetric meso-tetrasubstituted porphyrins 64 [166– 168].
Scheme 19
B B
CHOH
A NH NH N
A A
H NH N HN
B
ABAB
62 58
B B
CHOH
NH HN NH N
A C
+ A C
H NH HN H N HN
HOHC
D D
ABCD + ABAB+CDCD
64
B B
CHOH
A NH HN NH N
C
+ A C
H NH H
HN N HN
CHOH
B B
ABCB
63 11 65
B B
CHOH
A NH HN NH N
C
+ A C
H NH H
HN N HN
CHOH
D D
ABCD
66 11 64
ɛ
To obtain carbinols 62 and 63, use can be made of the methods applied for synthesis
of pyrrylcarbinols (Scheme 5): acylation of the magnesium derivative of dipyrrolylmethane
by acylchlorides or according to Wilsmeyer to acyldipyrrolylmethane followed by its re-
duction to carbinol or interaction of respective carbaldehyde with aryllithium derivatives.
A new convenient method of synthesis of carbinols has been developed [169, 170]. The re-
action of o-mercaptophenol with carboxylic acid or its derivatives in the presence of BF3
etherate yields benzoxathiol tetrafluorate 67, a masked equivalent of aroyl. Benzoxathiol
Scheme 20
B S BO
B -
+ BF4 S
O S NH NHO NH
pyridine 66
+ A A A
CH3CN ɩpyridine
NH NH CH3CN NH
S
B O
67 11 68 69
BF3 Et2O HBF4 OEt2 HgO HBF4 OEt2 HgO

B B B B
BCOOH
+ OH O OH O
OH NH NH NH NH
LiAlH4 LiAlH4
A A A A
SH NH NH NH NH
OH O
B B
62 63
tetrafluorate readily reacts in acetonitrile containing one equiv. pyridine at room tempera-
ture with dipyrrolylmethane 11 to yield 5- 68 or 5,5′-disubstituted dipyrrolylmethane 69
(the use of another benzoxathiazol tetrafluorate at the 2nd stage ultimately leads to nonsym-
metrically substituted dimethylcarbinol 66), whose oxidative hydrolysis to acyldipyrrolyl-
methanes and subsequent reduction yields required carbinols 62 or 63 [166] (Scheme 20).
There are methods of synthesis of nonsymmetric meso-tetrasubstituted porphyrins us-
ing tripyrranes ([3+1] condensation) [171] (Scheme 21) and bilanes [166] (Scheme 22). The
reaction of pyrrole with benzoxathiazole tetrafluoroborate 67 successively yields 2-substi-
Scheme 21
O
A A A
A O OH
- S
+ BF4 pyridine LiAlH4
O S NH HgO
+ NH NH
N CH3CN BF3 OEt2
H S
A O OH
A A
O
71
67
70
B
HO TFA, 25oC
excess
pyridine
HN
A B A
N N
HO
B H
NH HN NH
1. TFA H
N
N 2. DDQ
A B A
73 72
Scheme 22
B B B
O OH
NH NH NH NH HN
1. EtMgBr LiAlH4 BF3-Et2O
A A A A
2. BCOCl excess
ɩɢɪɪɨɥ
NH NH NH pyrrole
ɢɡɛɵɬɨɤ NH HN
O OH
B B B
11 63 74
B
1. CCHO, TFA
2. DDQ
NH N
A C
N HN
B
65
tuted pyrrole and then 2,5-disubstituted pyrrole 70 unlike the repeated substitution in posi-
tion 4 in the acylation reaction. The use of dicarbinol 71, obtained after the reduction, for
synthesis of tripyrrane 72 in excess pyrrole makes it possible to perform further the 3+1 con-
densation to obtain cis-A2B2 porphyrin 73.
The use of dipyrrolylmethane dicarbinol 63 makes it possible to obtain bilane 74 for
synthesis of ABCB-type meso-tetrasubstituted porphyrin 65 [166].
Thus, the use of linear meso-substituted oligopyrroles makes it possible to obtain in-
dividual nonsymmetric meso-tetrasubstituted porphyrins without impurities of isomeric
products. However, these methods require sufficiently complex syntheses of initial oli-
gopyrroles.
2 meso(5,15)-Disubstituted Porphyrins
5,15-Diphenylporphyrin and its β-alkylsubstituted analogues are of interest because these
compounds have some common features peculiar of 5,10,15,20-tetraphenylporphyrin,
namely the presence of two meso-phenyl groups, with some features of β-alkylporphyrins
owing to the occurrence of unsubstituted meso-positions, and porphin, in the absence of
β-substituents; herewith, such a porphyrin is not so inaccessible as the totally unsubstituted
initial porphin.
5,15-Diphenylporphyrins 4 of symmetric structure (trans-) have been known since
rather long in the form of octaalkylsubstituted derivatives, but in the form of β-unsubstitut-
ed analogues they appeared only recently, when unsubstituted dipyrrolylmethane 75 [26,
93, 172, 173] and meso-phenyldipyrrolylmethanes 11 [147, 172, 174] became relatively ac-
cessible.
Dipyrrolylmethane 75, which has neither meso- nor β-substituents, is obtained in three
stages from pyrrole and thiophosgene via thioketone 76 [175], which by the action of hy-
drogen peroxide in an alkaline medium is converted to ketone 77 [175], reduced to 75 under
the action of diborane with the total yield of about 40% [173, 176] (Scheme 23). Recently,
it was found that 75 could be obtained by direct desulfonation of thioketone 76 by Reney
nickel or sodium boron hydride [172]. Besides, it was possible to perform a small-scale con-
densation of polyformaldehyde (paraform) with excess pyrrole in the presence of boron tri-
fluoride etherate or trifluoroacetic acid, which yields dipyrrolylmethane 75 with the yield
of 40% in one stage [177, 178].
Proceeding from the features of structure of meso(5,15)-disubstituted porphyrins, sev-
eral possible routes for their synthesis can be presented (Scheme 24). All these methods
Scheme 23
S O
CSCl2 H2O2
OH- NH HN
N NH HN
H
76 77
(CH2O)n
BF3 Ni or NaBH4 B2H6
NH HN
75
were tested for synthesis of trans-substituted porphyrins, but the most applicable are routes
A and B, as routes C and D require an additional stage of producing formyldipyrrolyl-
methane. However, the use of route D makes it possible to obtain porphyrins nonsymmet-
rically substituted via phenyl rings without impurities of other porphyrins.
Scheme 24
R R
CHO
R R
NH H NH
+ ArCHO
A R Ar R C Ar
NH NH
R R
R R
R NH N R
78 80
R N HN R R
X R R CHO X
R R R
B D
H NH R Ar R H NH HN H
+ HC(OMe)3 4 +
Ar NH Ar NH HN Ar
R X = H, CO2H R R
X CHO X
R R R
79 81 79
It should be noted that β-substituted dipyrrolylmethanes used for synthesis of

5,15-substituted porphyrins via routes A, B and D should have a symmetric system of sub-
stitution relative to the meso-carbon atom, otherwise two isomeric porphyrins are formed.
Synthesis of symmetric (relative to the meso-carbon atom) meso-substituted β-substi-
tuted dipyrrolylmethanes 86 used to produce meso-substituted porphyrins consists in acid-
catalyzed condensation of aldehydes with α-unsubstituted pyrroles 82, in which the second
α-position is substituted by a readily removed group (usually, carbethoxyl or carb-
oxybenzyl) [165, 179, 180] (Scheme 25). Then follows the removal of protective groups
Scheme 25
B B B
COOR COOH
A A A
COOR
B NH NH NH
CCHO C OH- R=Et C t C
NH
A H+ H H2(Pd/C) R=Bz H
NH H NH
NH
A A A
COOR COOH
B B B
83 84 85 86
by, respectively, alkaline hydrolysis 84 (R = Et) or hydrogenolysis 84 (R = Bz) [173] with

subsequent decarboxylation 85 by heating in high-boiling solvents, such as ethylene glycol
[165], ethanolamine, DMFA [173], diethylformamide [181, 182]. For carbethoxy deriva-
tives 84 (R = Et), the protective grouping could be removed in one stage by heating in an
aqueous alkaline solution in a sealed ampoule at 180°C [80] or by boiling in alkaline eth-
ylene glycol in an inert atmosphere [183].
Symmetric meso-unsubstituted dipyrrolylmethanes 88 are obtained by self-condensa-

tion of pyrrylcarbinyl cations 87 in binary solvents containing water, with removal of form-
aldehyde, often with an almost qualitative yield [126, 184, 185] (Scheme 26). This method
is the most convenient for synthesis of such dipyrrolylmethanes.
Scheme 27
A B
+
RO2C CH2 B OH B B B
N
H2C
H
87 A A A A
+
NH HN -ɋH2O NH HN
H2O
RO2C CO2R RO2C CO2R
A B
88
RO2C CH2OH
N
H
Pyrrylcarbinyl cations 87 are sufficiently stable due to the delocalization of positive

charge by the electron-rich pyrrole nucleus (Scheme 27); as the result, they are readily ob-
tained from various precursors, such as 89, by the SN1 mechanism. The broad diversity of
precursors and ease of formation of pyrrylcarbinyl cations makes it possible to obtain and
use them in various nucleophilic solvents, such as alcohols, carboxylic acid and even water.
The most stable α-acetoxymethylpyrroles give high yields of dipyrrolylmethanes in boiling
in ethanol or methanol containing 1% hydrochloric acid [185– 187] or in boiling in aqueous
acetic acid [188].
Scheme 28
A B A B A B A B
+
+
R CH2X -X
- R .. CH2 R + CH2 R .. CH2
N N N N
H H H H
87 89
+ +
X = -Br, -Cl, -OCH3, -OH, -OCOCH3, -NR3, -NHR2 etc. R = CO2Et(Bz)
The most widespread synthesis of 5,15-diphenylporphyrins 4 consists in condensation

of α,α′-unsubstituted dipyrrolylmethanes 78 with aldehydes in the presence of acid (route
A) and is similar to that of meso-tetraphenylporphins.
The condensation reaction of dipyrrolylmethane 78 (R = Et) with arylaldehydes in
benzene containing a catalytic amount of trifluoroacetic acid and oxidation by oxygen of
the air yielded trans-substituted porphyrins 4 with 30–40% yields and with only traces of
monoarylporphyrins 3; however, in boiling in propionic acid containing zinc acetate, only
monoarylporphyrins 3 with the yield of 15–25% were formed [189].
It follows from these data that oligomeric polymethylenepyrroles and cyclic porphy-
rinogens initially formed in condensation via route A exist in acidified solutions as an equi-
librium mixture with respective pyrrylcarbinyl cations, which acquire stability owing to the
delocalization of charge on the pyrrole ring. Pyrrylcarbinyl cations, e.g., 90, can be incul-
cated via the pyrrole– methylene carbon link, followed by the release of the alternative pyr-
rylcarbinyl cation 91, which can lead to the removal of the aryl substituent from meso-
position and isomerization of β-alkyl substituents in porphyrins obtained in the subsequent
oxidation of porphyrinogens (Scheme 29).
Scheme 29
A H Ar
B
NH + A B B A
H H
H + H
H Ar CH2 + Ar
NH N N
B H H
90
A H Ar
B H H
A
NH + A B B A
H H
H + H
Ar H CH + H
NH N Ar N
A H H
91
B H H
B H H B H H B H H
A
+90 A B A + -91
A
NH NH H NH
H H
Ar Ar
Ar N NH H NH
NH H
A A B
B H H B H H A H Ar
Initially, the synthesis was conducted under conditions similar to the production of tet-
raphenylporphins by a one-stage method in solvents containing acid [26]; however, further
studies [190] showed the best results to be provided for by the use of a two-stage method
with intermediate formation of porphyrinogen 93 from 92 under the action of acid, and the
subsequent oxidation of 93 to porphyrin 94 by benzoquinone derivatives (Scheme 30).
Condensation of 5,5′-unsubstituted tetraalkyldipyrrolylmethanes 92 (A, B = Me, Et)
with benzaldehydes in methanol in the presence of strong inorganic or organic acid in an
Scheme 30
A A H R A A R A
B B B B B
NH + NH HN NH N
H [O]
+ RCHO
NH NH HN N HN
B B B B B
A A H R A A R A
92 93 94
inert atmosphere followed by isolation and oxidation of the formed porphyrinogen 93 by

benzoquinone derivatives yields 5,15-diphenylporphyrins 94 with a high yield (up to 60%)
[190– 194].
Studies of the conditions for carrying out this reaction in methanol with organic acid
additions [193] showed the yield of diphenylporphyrins not to depend in practice on acid
used (CF3COOH or CCl3COOH). However, the formation rate of porphyrinogen decreases
significantly in passing from trifluoroacetic acid to benzoic acid.
The reaction of condensation and oxidation can be conducted similar to the Lindsey
method for meso-tetraphenylporphins without isolating intermediate porphyrinogen in me-
thylene chloride or chloroform. In this case, we can use sufficiently high concentrations of
the reacting components (in contrast to the Lindsey method). Studies have shown that the
reaction is best run in chloroform with an addition of chloro- or trichloroacetic acid fol-
lowed by the oxidation of the reaction mass by o-chloranil, p-CA or DDQ [195].
The effect of the length of alkyl substituents in initial dipyrrolylmethanes 92 on the
yield of 5,15-diphenyloctaalkylporphyrins 94 was studied [183]. The occurrence of
small-size substituents in 3,3′-positions of dipyrrolylmethane (B = H, Me, Et; A = H, Me)
has an insignificant effect on the yield of porphyrins; however, the presence of more bulky
groups (B = propyl, butyl, amyl, hexyl and especially strongly benzyl; A = Me) decreases
it considerably. These facts indicate that the occurrence of bulky substituents in 3,3′-posi-
tions of dipyrrolylmethane prevents the formation of conformation 92a, which is required
for the condensation reaction to be performed. In this case, they are mainly in the energet-
ically more advantageous transoid form 92b.
B H H
B H H B NH
A
A A NH
NH HN B A
92a 92b
Introduction of the methyl group in 4,4′-positions of dipyrrolylmethane 92 (A = Me)

little affects the yield of porphyrin 94 (it even slightly increases); however, its substitution
by the ethyl group (A = Et) leads to a decrease of the yield due to steric hindrances of the
condensation reaction.
In passing from formaldehyde to acetic aldehyde, the yield of porphyrins 94 increases
significantly. Transition to benzaldehyde little affects the yield. The electronic nature of
substituents and their position in initial benzaldehydes have a small effect on the yield of
porphyrins 94, but the occurrence of two substituents in ortho-positions of the phenyl ring
leads to a strong decrease of the yield [183].
This two-stage method is at present used sufficiently broadly also for synthesis of
strapped 95 [196-199] and cyclophane dimeric porphyrins 96 [198, 200, 201], as well as
compound systems containing porphyrin fragments [202–207].
Interestingly, strapped porphyrins 95 are formed in condensation of dimeric benzalde-
hydes with a certain, larger-than-minimal, length of the linking group [ortho-; R =
–(CH2)5-]; yields are sufficiently high. At the length of the substituting group close to a
minimum, strapped porphyrins possess a strained distorted dome-like structure, which de-
termines their main physicochemical properties. At the length of the substituting group less
than a minimum [ortho-, meta-; n = 3, 4], cyclophane dimeric porphyrins 96 with a low
yield are formed. These porphyrins possess an unstrained structure, unlike cyclophane
B B
A A
R NH N
B B O N HN O
A A
O A A O
NH N (CH2)n B B
(CH2)n
B B
N HN
A A A A
O NH N O
B B
95 N HN
A A
B B
96
dimeric porphyrins with four bridges 19; however, they exist in the form of three fixed at-
ropisomers with different distances between the reaction centres of the porphyrin nuclei
α,α,α,α-, α,α,α,β- and α,β,α,β- 97. Other possible isomers are not formed due to too large
strains in bridge groups [200, 201].
NH N NH N N HN
N HN N HN NH N
NH N NH N N HN
N HN N HN NH N
Į,Į,Į,Į-97 Į,Į,Į,ȕ-97 Į,ȕ,ȕ,Į-97
Condensation of bis(3-formylbenzyl)sulfide with 4,4′-dimethyl-3,3′-diethyldipyrro-

lylmethane in acetonitrile in the presence of trichloroacetic acid, with subsequent oxidation
by p-CA [208] yields mainly 5,15-porphodimethene 98 (yield, 65%) and only a small
amount of dimeric porphyrin 99 (yield, 3%).
In the process of condensation, use of a mixture of two aldehydes leads to three por-
phyrins (Scheme 31); herewith, the required porphyrin 100 by statistical considerations is
obtained in the amount of 50% of the porphyrin mixture, which is easier separated than the
NH N
S
N HN
CH2 CH2 H2C CH2
S S
NH N
H2C CH2
H N HN H NH N
N HN
98 99
mixture of six porphyrins formed in condensation of two aldehydes and pyrrole (Scheme
10), so this method is also used sufficiently widely [207, 209–214]. It should be noted that
this method sometimes makes use of a large excess of one, more accessible aldehyde, so
that the other aldehyde is used more completely [215, 216]. Other examples include the syn-
thesis of rotoxanes based on porphyrins [217–219] and oligoporphyrins [207, 220].
Scheme 31
C C H A C C A C
D D D D D
NH A-CHO H + NH HN NH N
[O]
+ + A2 + B2
B-CHO
NH NH HN N HN
D D D D D
C C H B C C B C
92 100 (AB)
As it was pointed out earlier, the reaction (route A) is complicated by that it forms, to
a greater or smaller extent, impurities of monophenylporphyrins due to partial regrouping
in an acidic medium of dipyrrolylmethanes or intermediate tetrapyrroles [190] (Scheme
29). In some cases, monophenylporphyrins become the main or even sole reaction products,
especially at elevated temperature [183, 189, 190], however, their yield is lower than that
of diphenylporphyrins.
To rule out regrouping processes, of great interest is the use of a method similar to Rot-
mund’s synthesis of diphenylporphyrins 94. The method consists in the condensation of
dipyrrolylmethanes 101 with aldehydes in a medium of high-boiling heterocyclic solvents
(pyridine, quinoline, collidine) in the presence of coordination agents (zinc acetate)
(Scheme 32) [221]. In the case of using pyridine, the process is run at 180°C under pressure
using nitrobenzene as an oxidant, and the application of higher-boiling quinoline makes it
possible to conduct the reaction at boiling in the presence of air. The yields of phenyl-sub-
stituted porphyrins in this reaction are 10–20%; however, on the one hand, the method rules
out regroupings proceeding in an acidic medium, and on the other it enables using more ac-
cessible α,α′-dicarboxydipyrrolylmethanes 101 (X = COOH) stable to oxidation. Besides,
the method is of interest in condensation with aldehydes unstable in an acidic medium.
Another, the most known, method of synthesis of 5,15-diphenylporphyrins 94, first
used by Baldwin [222], consists in condensation of α,α′-unsubstituted or α,α′-dicarboxy-
meso-substituted dipyrrolylmethanes 102 with trimethoxy(ethoxy)methane in the presence
of trichloroacetic acid or trifluoroacetic acid in chloroform or methylene chloride [152, 172,
Scheme 32
A A R A
X
B B B
NH N N
Zn(OAc)2
+ RCHO Zn
pyridine,
NH nitrobenzene N N
B B B
X
A X=H, COOH A R A
101
Scheme 33
B B B B B
X
A A A A A
H NH H+ NH N NH N
H R [O]
+ HC(OMe)3 R R
R NH R H
N HN N HN
A A A A A
X
B X=H, COOH B B B B
102 103 94
ɛ / / ɮ
221, 223– 226] (Scheme 33) (route B). An oxidant of intermediate porphodimethene 103
can be oxygen of the air or 1,4-benzoquinone added at the end of the synthesis [223, 225,
226] or DDQ [152]. The reaction proceeds via formylation of 102, condensation of formyl-
dipyrrolylmethenes to porphodimethene 103 and its subsequent oxidation to porphyrin 94,
being, in this way, a simplified variety of routes C or D.
In [225], the yield of porphyrin 94 was shown to strongly depend on the extent of dry-
ing of initial α,α′-dicarboxylic acid of dipyrrolylmethane, whereas the use of α,α′-unsub-
stituted dipyrrolylmethane makes this reaction less dependent on its conditions. Application
of the benzoquinone oxidant at the last stage increases the yield of porphyrins as compared
with the use of air oxidation.
Method B is widely used for synthesis of belted porphyrins 95 [179, 222, 227]
(Scheme 33) and cyclophane dimeric porphyrins 88 [200] (Scheme 34). In this case, self-
condensation of bis-dipyrrolylmethane 104 or 105 is performed.
Scheme 33
R R
B B B B
COOH HOOC
O A A O O A A O
NH HN H+ NH N
+ HC(OMe)3
NH HN [O] N HN
A A A A
COOH HOOC
B B B B
104 95
Scheme 34
B B B
COOH
A A A
NH NH N
O NH O N HN O
A A
COOH
B H+ B B
(CH2)n B + HC(OMe)3 (CH2)n (CH2)n
[O] B B
COOH
A A A
O NH O NH N O
NH N HN
A A A
COOH
B B B
105 96
Route D makes it possible to obtain trans-disubstituted porphyrins 108 having various

meso-substituents; however, this route requires two different dipyrrolylmethanes 106 and
107, each of which can not self-condense [228] (Scheme 35).
Scheme 35
B C B C
OHC
A D A D
NH HN NH N
R H H +
+ R R1
H NH R1 [O]
HN N HN
A D A D
OHC
B C B C
106 107 108
Asymmetrically substituted diphenylporphyrins 108, besides, can be obtained by con-

densation of 1,19-unsubstituted 10-substituted biladienes-a,c 109 with aldehydes. An ex-
ample of this kind of synthesis is presented in Scheme 36 [229]. Initial biladienes 109 can
be obtained by condensation of meso-substituted dipyrrolylmethanes 106 with
2-formylpyrroles 110.
Scheme 36
B B C B C
A A D A D
+
R NH C D + NH HN NH N
H R R 1-CHO
+ +
R R1
H NH OHC H +
H
N NH HN N HN
A H A D A D
B B C B C
106 109 110 108
3 Synthesis of meso(5)-Phenyl-β-octaalkylporphyrins
There are several methods of synthesis of 5-phenyloctaalkylporphyrins 3. Earlier, it has
been shown [189, 190] that monophenylporphyrins are formed, owing to regroupings in an
acidic medium (Scheme 29), as an impurity in condensation of α-unsubstituted dipyrrolyl-
methanes with aldehydes; herewith, in some cases they are the main products of this reac-
tion, though their yield is lower than that of respective diphenylporphyrins.
The most currently widespread method of synthesis of 5-substituted porphyrins 113
consists in condensation of meso-substituted α-unsubstituted dipyrrolylmethanes 106 with
5,5′-diformyldipyrrolylmethanes 111 in alcohols or methylene chloride under the action of
strong acids (hydriodic, perchloric or p-toluenesulfo); the oxidant of intermediate por-
phodimethene 112 is oxygen of the air or benzoquinone derivatives (Scheme 37) [207, 225,
230–240].
5,5′-Diformyldipyrrolylmethanes 111 are produced by formylation of α-unsubstituted
dipyrrolylmethanes 92 by a mixture of POCl3 –DMFA according to Wilsmeyer [173] or a
mixture of ortho-formic ester and trifluoroacetic acid [241].
Scheme 37
A C A C A C
OHC
B D B D B D
NH HN + NH N NH N
H H H [O]
+ R
R NH HN R N HN
N HN
B D B D B D
OHC
A C A C A C
106 111 112 113
Another, similar, method is condensation of α-unsubstituted meso-phenyldipyrrolyl-

methanes 106 with 5,5′-dimethoxymethyldipyrrolylmethenes 114 in boiling benzene fol-
lowed by oxidation by benzoquinone derivatives [180] (Scheme 38). A lower yield of
porphyrins is compensated for by the accessibility of 5,5′-dimethoxymethyldipyrrolyl-
methenes 114, which are obtained by solvolysis of pyrroles 115 in a hot solution of hydro-
bromic acid (48%) and formic acid (88-90%), which leads with high yields (80– 90%) to
dipyrrolylmethenes 116 symmetric relative to the meso-bridge carbon. Dipyrrolylmethenes
116 are further brominated in a methanol solution to 114 [116, 242, 243] (Scheme 39). The
method is complicated by that, along with 5-phenylporphyrins 113, it forms a sufficiently
large amount of octaalkylporphyrins unsubstituted in meso-positions. However, the perfor-
mance of this synthesis in the absence of acid catalysts excludes the possibility of regroup-
ings of β- and meso-substituents [180].
Scheme 38
A C A C
MeOH2C
B D B D
NH HN NH N
H
+ Br- R
R +
NH HN N HN
B D B D
MeOH2C
A C A C
106 114 113
Scheme 39
C C
CH3 CH2OMe
D D
C D
NH NH
HBr 1. Br2
Br- Br-
H3C CO2R HCOOH + 2. MeOH +
N NH NH
H
D D
R = H, Et, tBu CH3 CH2OMe
C C
115 116 114
Scheme 40
B B B R B
A A A A
+
NH HN + - NH N
H or OH
2Br- + RCHO
+
NH HN N HN
C C C C
D D D D
117 113
Scheme 41
C B B
X
D A A
B A +
NH + NH HN
H
2Br-
+ CHO +
NH N NH HN
D H C C
X X= H, COOH
C D D
101 109 117
Scheme 42
B B B B
A A A A
+
NH HN NH N
OH-
2Br-
+
NH HN NH HN
C C C C
D D D D
117 118
Yet another widespread method of synthesis of 5-phenylporphyrins 113 is condensa-

tion of benzaldehydes with 1,19-diunsubstituted biladienes-a,c 117 in alcohols at acid
catalysis [234, 238, 244] or base catalysis (Scheme 40). In this case, there are no limitations
on the symmetry of β-substituents in the porphyrin cycle, as their set is given by the initial
biladiene-a,c. Initial biladienes are sufficiently accessible and are obtained in acid-
catalyzed interaction of dipyrrolylmethanes 101 with formylpyrroles 109 (Scheme 41). An
interesting feature of this synthesis of 5-phenylporphyrins is that respective corrole 118 can
be formed besides the required 5-phenylporphyrin 113; herewith, the amount of the corrole
is determined by the porphyrine formation rate. Corroles 118 are the only products in
self-condensation of 1,19-diunsubstituted biladienes 117 in the presence of bases during the
action of weak oxidants on them [246] or in illumination [247] (Scheme 42).
4 Reactions of Introduction and Modification of Substituents in Phenyl

Rings of meso-Phenyl-substituted Porphyrins
4.1 Introduction of substituents into phenyl rings of meso-phenylporphyrins

As β-positions of the porphyrin macroring in reactions of electrophilic substitution are more
active than phenyl rings, only some examples of direct electrophilic substitution in meso-
aryl groups are known. In all such cases, the porphyrin cycle is deactivated to the electro-
philic attack by protonation via intracyclic atoms of nitrogen.
In 1962, Winkelman, in studies of the ability of porphyrins to be selectively accumu-
lated in tumour cells, first performed the sulfonation of tetraphenylporphin (H2TPP) and
obtained water-soluble meso-tetra(4-sulfophenyl)porphin 119 (yield, ~70%) (Scheme 43)
[248]. The reaction was performed at 100°C; concentrated sulfuric acid was used as a sul-
fonating agent. Subsequently, it was found that, along with 119, porphyrin with sulfo groups
was also formed in only three phenyl rings [249].
Scheme 43
HO3S SO3H
N N
H2SO4
NH HN NH HN
o
100 C
N N
HO3S SO3H
119
Isolation of compound 119 from the reaction mixture is associated with a number of
problems, however, is possible via its sodium or ammonium salts by way of multiple re-
sedimentation from a methanol– acetone solution [250], via the insoluble potassium salt
[251], as well as by dialysis [252].
To investigate the liquid-crystalline properties of H2TPP derivatives, the authors of
[253] conducted the sulfonation of covalently bound linear dimer of H2TPP to hexasul-
fophenyl derivative 120 with the yield of 90%. The reaction was carried out by heating por-
phyrin in concentrated sulfuric acid in a sealed ampoule on a boiling water bath for 4 h.
Similarly, sulfonation of mono-, di- and tetraphenyl-β-octamethylporphins was carried out
[254]. The results obtained show that the reaction is general irrespective of the number of
phenyl residues and the presence or absence of β-alkyl groups.
Electron-donor groups in phenyl rings facilitate sulfonation. Thus, in sulfonation of
β-octabromo-meso-tetramesitylporphin by oleum at 120°C all eight free positions of meso-
mesityl groups are sulfonated. This porphyrin was assumed to be used as a water-soluble
catalyst in oxygen transfer reactions [255, 256]. Production of sulfonated derivatives with
hydroxy- and methoxy- groups in ortho- and para-positions of phenyl rings [257– 259] and
meso-tetrakis(2-thienyl)porphin 121 was described [260]. These porphyrins were assumed
to be used in analytical purposes. It is interesting to note that sulfation of meso-tetrakis-
(2,6-dimethylphenyl)porphin proceeds in meta-positions of phenyl rings, forming 122 [43],

but not in para-positions as it was assumed earlier [261]. Sulfonation of mono-p-nitrophe-
nyltriphenylporphin by oleum leads to 5,10,15-tri(4-sulfonylphenyl)-20-(4-nitrophenyl)-
porphin, which was purified by chromatography on Sephadex G-10 [262].
HO3S OCH2CH2CH2CH3O SO3H
N N
NH HN NH HN
N N
HO3S SO3H HO3S SO3H

120
SO3H SO3H
HO3S S
Me Me
S SO3H
N
Me N Me
NH HN
NH HN
N
S Me N Me
S SO3H HO3S
HO3S Me Me
121 HO3S
122
The authors of [263], using the reaction of H2TPP to chlorosulfonic acid at room tem-
perature, obtained respective tetrachlorosulfonic derivatives. Chlorosulfonation of meso-
tetrakis(2,6-dichlorophenyl)porphin under the same conditions yields a mixture of mono-,
di- and tri-para-sulfonyl derivatives. Chlorosulfonation can be carried out with a high yield
at 100°C [263]; besides, tetrachlorosulfonic derivatives of H2TPP can be obtained by treat-
ing salts 119 with excess thionyl chloride in DMFA at 50°C for 3 h [264].
Chlorosulfonyl groups are easily converted to free sulfo acid, sulfamides or sulfo es-
ters at the nucleophilic attack with water, ammonia (amines) or alcohols, respectively [263,
264] (Scheme 44).
Scheme 44
ClO2S SO2Cl SO2OH
H2O
N
HNR2 SO2NR2
NH HN
N
HOR SO2OR
ClO2S SO2Cl
To obtain models of cytochrome P-450, porphyrin 123 was obtained by the treatment
of the sulfochloride derivative with respective amine [265, 266].
RO2S
Cl Cl
RO2S
Cl N Cl
Me
NH HN R = NH(CH2)3 Si OEt
OEt
Cl N Cl
SO2R
Cl Cl
SO2R
123
Of the other reactions of electrophilic substitution in phenyl rings of phenylporphyrins,

only the nitration of H2TPP by the action on it of excess fuming nitric acid in chloroform
is described [262]; herewith, the formed product is 5-(p-nitrophenyl)-10,15,20-triphenyl-
porphin 124 with the yield of 55% (Scheme 45). An increase of the nitration time and con-
centration of nitrating reagent leads to a mixture of mono- and di(nitrophenyl)porphyrins.
Porphyrin 124 can also be obtained with the yield of 80% by the treatment with a solution
of H2TPP in methylene chloride by acetic anhydride, trifluoroacetic acid and HNO3 in the
presence of montmorillonite K10 [267]. The work [268] studied the effect of the nature of
substituent on the nitration reaction. The most readily nitrated were found to be H2TPP de-
rivatives having electron-donor substituents in meta-positions of phenyl rings (nitration pro-
ceeds in para-position). Occurrence of an electron-donor substituent in para-positions
leads, under conditions of the reaction, mainly to oxidative destruction of the porphyrin cy-
cle. Electron-acceptor substituents in meta-positions hinder the nitration reaction, and in
their occurrence in para-positions the reaction does not proceed.
Scheme 45
NO2
N N
HNO3
NH HN 0 NH HN
CHCl 3, 0-5 C
N N
9
124
4.2 Modification of functional groups in phenyl rings of meso-phenylporphyrins

Many derivatives of phenyl-substituted porphyrins are difficult to synthesize by condensa-
tion of pyrrole or its linear derivatives with respective benzaldehydes. In this connection,
of interest is to consider methods, which make use of a modification of substituents in
benzene rings of porphyrins obtained with high yields. Of great interest are porphyrins con-
taining in their phenyl rings active substituents, such as oxy or amino groups, as they are
capable of various chemical conversions. By direct synthesis, these porphyrins are obtained
with a low yield or are not formed at all.
4.2.1 meso-Oxyphenylporphyrins and their modification

meso-Oxyphenylporphyrins are formed in condensation reactions with low yields and large
amounts of hard-to-separate impurities. Protection of the oxy group in respective oxyben-
zaldehydes by acylation [127] or sulfonylation [269] leads to a significant increase of the
yields of respective porphyrins, which, by way of hydrolysis in an alkaline medium, can
sufficiently easily yield required meso-oxyphenylporphyrins. However, a more promising
method is hydrolysis of readily available meso-methoxyphenylporphyrins (Scheme 46).
Scheme 46
OCOR
-
OH
-
OH RHal
OSO2R OH OR
K2CO 3
BBr3
OCH3
Hydrolysis of methoxyphenylporphyrins by the general method (by 48% hydrobromic

acid) showed this method to be little efficient, in view of its long duration and incomplete
conversion. The use of anhydrous aluminium chloride in boiling chlorobenzene as deme-
thylating agents gives no satisfactory results, either, as under the stringent conditions char-
acteristic of this method, porphyrins are subjected to a significant breakdown.
Good results are obtained using pyridine or aniline hydrochlorides in boiling as dem-
ethylating agents [270]. This method, however, is only applicable in the synthesis of the
most stable meta- and para-oxysubstituted tetraphenylporphyrins, whereas ortho-oxysub-
stituted tetraphenylporphyrins, as well as monooxyphenyl- and dioxyphenylporphyrins are
subjected to a significant destruction under conditions of this reaction.
A more suitable demethylating agent for synthesis of oxyphenylporphyrins is 60% hy-
drobromic acid (boiling in inert atmosphere), the use of which makes it possible to signif-
icantly increase their yield [270].
At present, a mild demethylating agent – boron tribromide in methylene chloride at
–20°C in inert atmosphere – is used for hydrolysis of methoxyphenylporphyrins
[271– 276]. However, it has been shown [277, 278] that the demethylation reaction can,
without decreasing the yield, be conducted at room temperature in the air in chloroform.
The reaction of alkylation of oxyphenylporphyrins by haloalkanes is of certain interest,
as it enables one-stage one-precursor production of porphyrins possessing diverse physic-
ochemical properties or having active groups at the periphery of the molecule. These active
groups can interact with the active central part of the porphyrin macrocycle or serve for at-
taching the molecule to various substrates.
Alkylation of oxyphenylporphins is performed at present mainly in dimethylforma-

mide in the presence of the basic agent potassium carbonate [279– 281]. The reaction pro-
ceeds at room temperature for labile oxyphenylporphyrins [279, 282] or during the boiling
for stable derivatives of tetraphenylporphin, which significantly decreases the interaction
time [281]. The yield of alkoxy derivatives of porphyrins is on average 80–95%. Alcohols
are not in practice used for this reaction, as both initial porphyrins and interaction products
solve in them poorly.
The discussed method was used to synthesize various alkoxy-substituted porphyrins,
well soluble in nonpolar organic solvents [281, 282], as well as tetra(carboethoxymethyle-
neoxyphenyl)porphyrins 125, whose hydrolysis forms tetra(carboxymethyleneoxyphenyl)-
porphyrins 126 soluble in alkaline solutions [283] (Scheme 47).
Scheme 47
OH OCH2CO2Et OCH2CO2H
ClCH2CO2Et 1. OH-
K2CO3 2. H+
125 126
Metalloporphyrins in nature function in chromoproteins, whose protein groups strong-

ly affect the properties of metal complexes [284]. Therefore, of great importance is the syn-
thesis and study of porphyrins, which at the periphery of the molecule have active
functional groups capable of interacting with the central reaction centre of the porphyrin
macrocycle.
Synthesis of such compounds based on modifications of natural porphyrins is rather
complicated and includes a large number of stages [223]. Bonds formed after attaching res-
idues with active groups are not too strong (they are mainly amide ones or ester ones) [223,
285–287].
For synthesis of such compounds, it is of interest to use synthetic porphyrins with oxy-
phenyl groups capable of forming stable ether bonds. The amount of active groups per one
porphyrin molecule rarely exceeds one, so the most appropriate oxyphenylporphyrins for
these purposes are monooxyphenylporphyrins: oxyphenyltriarylporphyrins 127a, which
are rather easily obtained by condensation of pyrrole with a mixture of oxybenzaldehyde
and benzaldehyde [47, 126, 127], or meso-oxyphenyl-β-alkylporphyrins 128a closer to nat-
ural phorphrins.
R A R A A R A
OH
B B B B aR=
NH N NH N NH N
Ar Ar NO2
N HN N HN bR =
N HN
C C B B
NH2
Ar D D R
A A ɫR=
127 128 129
A generalized scheme of the synthesis of porphyrins with active groups by the alkyla-
tion reaction is presented in Scheme 48.
As is seen in the scheme, the attachment of residues with active groups to porphyrin
Scheme 48
O(CH2)nX
Br(CH 2)nX
OH A
HX
B O(CH2)nBr
Br(CH 2)nBr
ɏ - residue with an active group
can proceed via two routes: alkylation of oxyphenylporphyrin by haloalkane with an active
group (route A), or preliminary alkylation of oxyporphyrin by excess α,ω-dibromoalkane
to form ω-bromoalkoxyphenylporphyrin with its subsequent interaction under similar con-
ditions with a compound having an active group (route B).
Route A is more preferable, as it is shorter and provides a higher yield of the end prod-
uct (with respect to initial oxyphenylporphyrin). However, it is not always possible to ob-
tain and purify haloalkane having an active functional group. In this connection, route B is
mainly used [279, 288].
4.2.2 meso-Aminoporphyrins and their modification

High yields of aminophenylporphyrins in condensation reactions of aminobenzaldehydes
with pyrrole and its derivatives are hard to expect. Aminobenzaldehydes are known to be
very unstable; e.g., p-aminobenzaldehyde polymerizes in an acidic medium and m-amino-
benzaldehyde exists only in diluted solutions or as a complex with tin dichloride dehydrate.
The literature gives a mention of only the direct synthesis of tetrakis(4-aminophenyl)-
porphin [26] with the nonreproducible yield of about 1%. Protection of the amino group by
acylation leads to a significant (up to 10%) increase of the yield of porphyrin; however, oth-
er isomeric aminophenylporphyrins could not be obtained in this way.
Proceeding from the above-said, a more promising approach is production of ami-
nophenylporphyrins by reducing respective nitrophenylporphyrins, which are obtained
with sufficiently high yields by the condensation of nitrobenzaldehydes with pyrrole and
its derivatives. As the sole reducer, this reaction makes use at present of tin dichloride di-
hydrate in hydrochloric acid or in polar solvents with its addition. This method was pro-
posed by Collman [289] and found wide use.
For tetra(nitrophenyl)porphyrins, reduction is done at a 1.5-fold excess of tin dichlo-
ride dihydrate in concentrated hydrochloric acid at a temperature of 70–80°C, which en-
ables an almost qualitative yield of tetra(aminophenyl)porphyrins [290– 293].
Mono- 127b, 128b and disubstituted 129b nitrophenylporphyrins require more mild
conditions of reduction, so in this case the reduction reaction is conducted at room temper-
ature [294, 295] and in a methanol medium, which transfers the obtained aminophenylpor-
phyrins into a solution and contributes to the complete reduction of initial nitrophenyl-
porphyrins.
The amino group of aminophenylporphyrins is very active and reacts to many re-
agents. Of special interest is the diazotization reaction, which is widely used for synthesis
of aromatic compounds with various functional groups. This reaction has advantages in that
only a small amount of groups prevent it, in contrast with reactions using Grignard reagents.
Application of the diazotization reaction enables production of an interesting and important
class of organic compounds – azo dyes.
Tetra(aminophenyl)porphins are readily diazotized by sodium nitrite in aqueous solu-
tions of mineral acids [296–298]. The obtained diazonium salts are rather stable. Their no-
ticeable decomposition with evolution of nitrogen is observed only at a temperature higher
than room temperature. The porphyrin cycle under conditions of the diazotization reaction
is stable; however, diazotized tetra(2-aminophenyl)porphin during the heating yields com-
pounds of nonporphyrin character. Owing to this, ortho-substituted tetraphenylporphins are
formed with low yields and a large amount of hard-to-separate impurities. However, the
azocoupling reaction of diazotized tetra(2-aminophenyl)porphin to phenol successfully
proceeds on the cold to form tetra(2-oxyphenylazophenyl)porphin. Diazotized tetra(3- and
4-aminophenyl)porphins during the heating yield tetra(oxyphenyl)porphins as well as enter
into the Sandmeyer reaction and the azocoupling reaction, which makes it possible to obtain
tetra(halogenophenyl)porphins and azo dyes based on porphyrins, which are impossible to
obtain in any other way (Scheme 49).
Scheme 49
F
I
HBF4
Cu KI
+
NO2 SnCl 2 NH2 HNO N2 Cu 2X2 X
2
HCl H+ X=Cl,Br,CN
t
HR
OH
N=NR
Diazotization of mono- 127c, 128c and disubstituted 129c aminophenylporphyrins

followed by conversion of the obtained diazonium salts makes it possible to synthesize sub-
stituted porphyrins, which are inaccessible using other routes of synthesis. The use of so-
dium nitrite as a diazotizing agent in aqueous solutions of the acids in this case does not
lead to positive results, as such aminophenylporphyrins are not soluble under these condi-
tions. However, aminophenylporphyrins are diazotized by amylnitrite in a mixture of chlo-
roform and acetic acid [298]. The obtained solutions of diazonium salts enter into the
Sandmeyer reaction and the azocoupling reaction; the required basicity of the medium (pH
= 8–9) was created in this case by triethylamine.
Methylation of aminoporphyrins by iodomethane in DMFA in the presence of 2,6-
lutidine makes it possible to obtain cationic trimethylaminophenylporphyrins with the yield
of about 70% [299]. Tetrakis(para- and meta-trimethylaminophenyl)porphins are soluble
in water within a wide range of pH. Using potassium carbonate as bases, the reaction is
stopped at the stage of formation of dimethylaminophenylporphyrins [299] (yield, about
80%) (Scheme 50).
Thus, combining the methods of synthesis of meso-phenylporphyrins and modifica-
tion of substituents in their phenyl rings makes it possible to obtain compounds with prac-
tically any predetermined physicochemical properties and a required set of substituents.
Scheme 50
N(CH3)2
CH3I
NH2 K2CO 3
CH3I +
N(CH3)3
2,6-lutidine
The chemistry of porphyrins and their analogues is rapidly developing, so this review
does not pretend to be a complete coverage of the literature on the subject. It can be recom-
mended as a strategy for the synthesis and modification of required meso-substituted por-
phyrins based on the methods developed at present the most.
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- ссылки 7 и 14 одинаковые
- упоминается табл. 6 (a таблиц всего 5)
3 The Mechanism of Catalytic

Action of the Coordination
Centres of Catalase
Synthetic Models
T.N. Lomova1, M.E. Klyueva2 and M.V. Klyuev3

1Institute
of Solution Chemistry, Russian Academy of
Sciences, 1 Akademicheskaya Street, Ivanovo, 153045,
Russia; email: tnl@isc-ras.ru
2Ivanovo State University of Chemistry and Technology,
7 F. Engels Prospect, Ivanovo, 153000, Russia; email:
tnl@isc-ras.ru
3Ivanovo State University, 39 Ermak Street, Ivanovo, 153025,
Russia; email: klyuev@ivanovo.ac.ru
This chapter presents the results of studying the catalytic activity of manganese(III)
and copper(II) porphyrins alkyl- and phenyl-substituted at β- and meso-macrocycle
positions in hydrogen peroxide decomposition in DMFA–KOH–H2O. The
ion-molecular mechanism of the process with kinetically significant stages of two-
electron metalloporphyrin oxidation and further partial reduction as well as acid-
base peroxide equilibrium reactions is stated. Effective catalysis of hydrogen per-
oxide decomposition is determined by the degree of porphyrin binding, mixed
coordination sphere structure and mutual influence of the ligands. The compounds
studied are shown to act similar to naturally occurring catalases.
Keywords: porphyrins, complexes, hydrogen peroxide, disproportionation reaction, catal-

ysis, mechanism, catalase models
Introduction
Studies of chelate complexes of copper as catalysts of the hydrogen peroxide decomposi-
tion reaction [1] showed the common factors for individual reaction mechanisms to be
94 T.N. Lomova, M.E. Klyueva and M.V. Klyuev
coordination of peroxide particles on the metal atom and availability of two free coordina-
tion sites for the catalytic action of copper(II) compound to be realized. Copper(II) porphy-
rins (CuP), as polychelate macrocyclic complexes, are of interest in both aspects. First, they
contain a coordinatively unsaturated copper atom of the 3d 9 electron configuration in the
environment of the cyclic aromatic ligand; this copper atom forms direct N→Cu dative
π-bonds along with coordination σ-interaction. A similar π-interaction is also possible in
the case of addition of axial ligands containing unshared electron pairs. Second, CuP satisfy
the requirement of the two-side access of reagents to the coordination core. Prospectivity
of using metalloporphyrins with a two-charge metal cation (MP) for catalyzing the dispro-
portionation reaction of hydrogen peroxide is evident at present [2, 3]. However, the re-
quired coordinative unsaturation of metal in metalloporphyrins is the greater, the higher the
formal charge of the metal atom is [4, 5]. For this reason, studies of acidoporphyrin metal
complexes of the oxidation degree greater than two (X)n–2MP is also rather promising. A
suitable model in the given case are manganese(III) porphyrins. Thus, in [3], monomeric
and covalently bound dimeric manganese porphyrinates were found to exhibit a catalase ac-
tivity in the presence of the basic nitrogen of imidazole under conditions of phase-transfer
catalysis.
However, no rigorous theory has been developed to predict the reactivity of new co-
ordination compounds, porphyrin complexes of copper and manganese including, and to
explain the differences in their catalytic activity with respect to hydrogen peroxide decom-
position. Several attempts have only been made to study the effect of the ligand environ-
ment of the central ion on its catalytic activity and the H2O2 degradation mechanism [6, 7].
In this situation, of special significance is to obtain new information on the mechanism of
the catalytic action of complex compounds with regulated coordination saturation of the
central atom. The latter is realized in the case of metalloporphyrins by targeted functional
substitution of porphin in the complexes and rational choice of acidoligands X – in mixed
(X)n–2MP complexes.
The results obtained by the authors of this chapter in studies of copper(II) porphyrin
complexes with various degrees of substitution by β- and meso-positions of the macrocycle
(formulas 1–4) as catalysts of the hydrogen peroxide decomposition reaction in the
DMFA–KOH–H2O system (reaction 1) are considered in [8, 9]. This chapter presents the
major regularities and conclusions pertaining to the catalytic properties of copper porphy-
rins with respect to reaction (1). For manganese complexes, new data are presented on the
catalysis of the disproportionation reaction (1) of manganese(III) porphyrins with a regu-
larly changing structure (formulas 5–9) [10]. The catalytic reactions involving manga-
nese(III) tetraphenylporphyrins 10 and 11 were studied earlier [8, 9].
2H2O2 = 2H2O + O2↑ . (1)
Octaethylporphin H2OEP and meso-tetraphenylporphin H2TPP were synthesized by

the known methods [11, 12]. meso-Phenyl-substituted octaethylporphyrins monophenyl-
octaethylporphin H2MPOEP, 5,10-diphenyloctaethylporphin H25,10DPOEP, 5,15-di-
phenyloctaethylporphin H25,15DPOEP and 5,10,15,20-tetraphenyloctaethylporphin
H2TPOEP were synthesized by A.S. Semeykin (Ivanovo State University of Chemistry and
Technology) [12].
Complexes of porphyrins with manganese(III) were obtained by the reaction of respec-
tive porphyrin with metal salt according to Adler’s method [13]. As complex formers, we
used MnCl2 ·4H2O (AR grade), Mn(AcO)2 ·4H2O (AR grade). (SCN)MnOEP 7 was
The Mechanism of Catalytic Action of the Coordination Centres of Catalase Synthetic Models 95
R R1 R CuOEP, R1 = R2 = R3 = R4 = H, R = C2H5 (1)

Cu5,10DPOEP, R1 = R2 = C6H5, R3 = R4 = H, R = C2H5 (2)
R R CuTPOEP, R1 - R4 = C6H5, R = C2H5 (3)
N N
CuTPP, R1 = R2 = R3 = R4 = C6H5, R = H (4)
R4 Cu R2
N N
R R
R R3 R
X (Cl)MnOEP, R1 = C2H5, R2 = R3 = R4 = R5 = H, X = Cl (5)

R1 (AcO)MnOEP, R1 = C2H5, R2 = R3 = R4 = R5 = H, X = AcO (6)
R5 R1
(SCN)MnOEP, R1 = C2H5, R2 = R3 = R4 = R5 = H, X = SCN (7)
R1 R2 (Cl)MnMPOEP, R1 = C2H5, R2 = C6H5, R3 = R4 = R5 = H, X = Cl (8)
Mn N
R1 N R1 (Cl)Mn5,15DPOEP, R1 = C2H5, R2 = R4 = C6H5, R3 = R5 = H, X = Cl (9)
N (Cl)MnTPP, R1 = H, R2 = R3 = R4 = R5 = C6H5 (10)
R4 N
R1 (AcO)MnTPP, R1 = H, R2 = R3 = R4 = R5 = C6H5 (11)
R1 R3
R1
obtained by treating a solution of (AcO)MnOEP 6 in DMFA with an excess aqueous solu-

tion of NaSCN.
1 Substituted Copper(II) Porphyrins as Catalysts of the Hydrogen Peroxide

Disproportionation Reaction
To carry out reaction (1), an aqueous solution of hydrogen peroxide was added to a prepared
solution of the copper complex and KOH in DMFA. The concentrations of the complexes,
KOH and H2O2 were varied within the limits of 10 –6 –10 –4, (0.18–3.54)·10 –2, 2.98–5.97
mol/l, respectively. The rate of reaction (1) was determined volumetrically by measuring
the volume of evolving oxygen.
For copper porphyrin complexes studied, the catalysis of H2O2 decomposition is ob-
served not within all catalyst concentration ranges used. Thus, for CuDPOEP, as for CuCl2
and Cu(AcO)2, the catalytic effect of the rate increase (W) as compared with the noncata-
lyzed reaction is observed only at concentrations greater than 1·10 –5 mol/l and increases
at an increase of CCuP. For reactions catalyzed by CuOEP and CuTPOEP the observed order
of the catalyst concentration is close to zero: W is practically independent of CCuP. Among
metalloporphyrins studied, significant catalytic activities in the H2O2 decomposition reac-
tions are manifested by CuTPP and CuDPOEP. Their presence in the reaction mixture in-
creases the reaction rate and decreases the activation energy three- to fourfold as compared
with the noncatalytic process. The activity of CuOEP and CuTPOEP complexes is compa-
rable with copper salts.
The complete kinetic equations (2) and (3) for complexes 2 and 4, CuCl2, Cu(AcO)2
and for complexes 1 and 3, respectively, were determined:
dCO2 / dτ = k ⋅ CH2O2 ⋅ CKOH ⋅ CCuP ⋅ (CO2 )0 = k ⋅ CH2O2 ⋅ CKOH ⋅ CCuP , (2)
dCO2 / dτ = k ′ ⋅ CH2O2 ⋅ CKOH ⋅ (CCuP )0 ⋅ (CO2 )0 = k ′ ⋅ CH2O2 ⋅ CKOH . (3)
Using the UV-visible spectroscopy method, π-cation-radical forms of metalloporphy-

rins 1 and 3 are identified as end products of the reaction with H2O2 in the presence of KOH.
fast
fast
slow
H2O + O2
Scheme 1 A scheme of elementary reactions in the copper(II) porphyrin–hydrogen peroxide–KOH

system.
For the reaction catalyzed by Cu5,10DPOEP (compound 2), the spectral characteristics in
the course of conversion are similar to complexes 1 and 3. In reaction mixtures, where
CuTPP is present as a catalyst, attempts to record the electronic absorption spectra were un-
successful due to the vigorous evolution of oxygen. However, after CuTPP was isolated
from the reaction mixture to CHCl3 the electronic absorption spectra reflected the absorp-
I I
tion of initial CuTPP (λmax = 538.5 nm) and its π-cation-radical form (λmax = 700 nm and
II
λmax = 664 nm).
According to the spectral data, CuTPP and Cu5,10DPOEP (complexes 2 and 4) in the
absence of alkali do not enter into a chemical interaction with H2O2, unlike 1 and 3, as well
as in contrast with similar manganese(III) complexes considered below. If the reaction with
hydrogen peroxide is assumed to run to form axial complexes with the catalyst (Scheme 1),
as is the case in the functioning of natural catalase [1, p. 466], which is in complete agree-
ment with the experimental kinetic equations (2, 3), the above difference in reactivity can
be explained by different stabilities of the above axial complexes.
The catalytic activity of metalloporphyrins in the H2O2 decomposition reaction, the
rate of which, according to Scheme 1, is W = k4 ·K, is explained by the ability of copper
cation in the complex to coordinate ligands additionally to the fifth coordination site and
by low ionization potentials of the coordinated porphyrin macrocycle. Inertness of com-
plexes 2 and 4 in the reaction with H2O2 in the absence of KOH is due to their poor capa-
bility of axial coordination: a weak holding of OH· ligands in (·OH)CuP+· eliminated in the
course of reduction at the slow stage (Scheme 1) leads to a sharp decrease of the activation
energy in the case of CuTPP (27±3 kJ/mol) and CuDPOEP (17±2 kJ/mol) as compared
with other CuP, for which E changes within the limits of (39±3)–(73±7) kJ/mol [9].
2 Kinetic Regularities and Mechanisms of Peroxide Decomposition

Reactions in the Presence of Acido Complexes of Highly Substituted
Manganese Porphyrins
As it was already noted, considering the role of axial coordination processes, one expects
an increased catalytic activity in passing to manganese complexes with porphyrins, where
the efficient charge of the central atom is not completely compensated for in coordination
and strong binding of the aromatic macrocyclic ligand. After elucidating the potential of
manganese complex compounds as catalysts of redox processes and understanding the role
of these compounds in medicine, agriculture and other fields, interest of researchers in them
permanently rises. Consideration of the role of manganese in enzyme as well as model sys-
tems simulating the functional properties of various enzymes [14] showed the significance
of compounds of this element in vital processes. Of especial importance are complexes of
manganese with porphyrins, as in their presence many reactions proceed under milder con-
ditions and in media, where simpler derivatives of manganese are inert or nonspecific [14].
Using complexes of manganese with various derivatives of porphin and various coordina-
tion centres, varying solvents and temperature, it is possible to achieve selective processes
of hydrogen peroxide decomposition, thus, modelling the action of catalases. The most sig-
nificant results in this direction were obtained in [15–20].
In the course of hydrogen peroxide decomposition catalyzed by metalloporphyrins and
metallophthalocyanines, the catalyst is observed to be partially (up to 30–50%) degraded
or its activity to be inhibited by an excess of bases added into the system, such as imidazole
[3, 21]. The data of the previous section suggest redox processes of the catalysis in the
course of the H2O2 decomposition reaction. For this reason, studies of the reactivity of por-
phyrin catalysts at the action of hydrogen peroxide on them are topical. In [22, 23] published
well back in 1960s, metallophthalocyanines have been shown to be subjected to oxidative
destruction under the action of H2O2 in an acidic medium. Pheophytin and its complexes
with metals are also oxidized but much slower. Since that time, conversion processes of
metalloporphyrins directly at the action of peroxide have not been studied systematically.
In studies of the kinetics of H2O2 decomposition in the presence of manganese(III) tet-
ra(2,6-dimethyl-3-sulfonatophenyl)porphin at pH 7.6–12.1, a conclusion was made of the
equilibrium coordination of the hydrogen peroxide molecule by the catalyst and the subse-
quent slow formation of the π-cation-radical form of oxomanganese(IV) porphyrin or ox-
omanganese(V) porphyrin [15]. Formation of oxo complexes (O)MnIVP, [(O)MnIVP]+·,
(O)MnVP, readily passing into one another, is also noted when using manganese
porphyrin–peroxide systems for oxidation and epoxidation of aliphatic and aromatic com-
pounds [16–20]. Thus, interaction of peroxyacetic acid with tetra(2,6-dichloro-4-R-phe-
nyl)porphyrin (R = CH3O, H, Br, Cl, NO2) complexes of manganese(III) in an
acetonitrile–acetic acid mixture leads to the formation of an intermediate
“catalyst–oxidant” complex [18]. In catalytic reactions of oxidation of cis-stilbene and
naphthalene, one observes an irreversible conversion of an intermediate complex
[(HOAc)MnRTDCPP](X) formed in the substitution of anion X by a molecule of HOAc in
the inner coordination sphere, into a mixture of two high-valent oxomanganese complexes.
The complexes are assumed to have the composition (X)(O)MnVRTDCPP and
[(O)MnIVRTDCPP]+· (X) [19]. The first complex is responsible for epoxidation of cis-stil-
bene; the second, for hydroxylation of naphthalene. In the medium of CH3CN, formation
of a stable complex is registered by the appearance of an absorption at 415 nm in the reac-
tion of (Cl)MnTDCPP with peroxyacetic acid, whereas in a medium of CH2Cl2 such a com-
plex proves to be short-lived and rapidly breaks down to destroy the macrocycle [20].
Conclusions on the conversions of catalysts in the cited works were mainly made
based on the analysis of the conversions of reagents or special compounds – traps – as in
[15], while the state of the catalyst itself – metalloporphyrin – was not studied. In this re-
view, we analyze the results of studies of the direct reaction of manganese porphyrins with
the oxidant H2O2. Using the spectrophotometric and kinetic methods, we studied the reac-
tions of (chloro)manganese(III) octaethylporphin (Cl)MnOEP (compound 5) with hydro-

gen peroxide in water–DMFA media at a temperature of 288–308 K.
2.1 Kinetics and Reaction Mechanism of Oxidation of Manganese(III) Porphyrins by

Hydrogen Peroxide
The kinetics of the reactions of mixed acidotetraphenylporphin complexes of manga-
nese(III) 10 and 11 with hydrogen peroxide in DMFA was studied earlier [8, 9, 24]. The
crucial role in the coordination of H2O2 by the porphyrin complex (equation 4), from which
the redox process begins, was shown to belong to the coordination unsaturation of the man-
ganese atom, which is mainly regulated by the state of the links with the coordinated mac-
rocycle. The structure of the macrocycle and extent of its aromaticity can be considered as
a means of controlling the oxidation process. Data accumulation on the porphyrin
structure–reactivity relationship with respect to the oxidation of hydrogen peroxide is at the
very starting point.
(Х)MnTPP + H2O2 (Х)(H2O2)MnTPP . (4)
According to the spectrophotometry data [8, 9], the end products of the reaction of
complexes 10 and 11 with hydrogen peroxide are not the same depending on the peroxide
concentration ranges, which are 0.017–0.5, 0.67–1.0 and 1.26–3.98 mol/l. The products
are identified as an oxidized form with the localization of electron deficit at the aromatic
macrocycle – the π-cation-radical of manganese(III) tetraphenylporphin in the first interval
of H2O2 concentrations and the form oxidized at Mn atom – manganese(IV) tetraphe-
nylporphin – at higher concentrations of peroxide. In all cases, kinetic measurements re-
vealed the first order by initial metalloporphyrin. Experimentally found complete kinetic
equations for the H2O2 concentration range of 0.017–0.1 and 1.26–3.32 mol/l are written
down as equations (5) and (6):
− dC(Cl)Mn III TPP / dτ = k v1 ⋅ C(Cl)Mn III TPP ⋅ CH 2O2 , (5)
−1/ 2 . (6)
− dC(Cl)Mn III TPP / dτ = k v3 ⋅ C(Cl)Mn IIITPP ⋅ CH O
2 2
Here kv1 and kv3 are the true rate constants of the reaction with hydrogen peroxide respec-
tively for the first and third concentration ranges of peroxide.
For the overall reaction described by kinetic equation (5), the following sequence of
elementary stages is proposed, with account for which the rate constant in equation (5) is
equal to kv1 = k2 ·K1:
K1
(Cl)MnIIITPP + H2O2 (Cl)(H2O2)MnIIITPP , (7)
(Cl)(H2O2)MnIIITPP ⎯⎯→
2 k
О=MnIII(TPP+ ?) + Н2О + Cl- .
. (8)
The concentration constant of equilibrium (7) at 298 K, found by the method of

log I
a b
log CH2O2
CH2O2, mol/l
Figure 1 A titration curve (a) and a dependence of log I on log CH2O2 (b) (R2 = 0.98) at the titration
of the complex of (Cl)MnIIITPP in DMFA by hydrogen peroxide.
spectrophotometric titration in this work (Fig. 1) and calculated by formula (9), is K1 =

(20±3) l·mol –1. Hence, the rate constant k2 of an elementary reaction (8) at 298 K is equal
to 1.95·10 –3 s –1.
(Ap − A0 )/(A∞ − A0 ) 1
K = ⋅ , (9)
1 − (Ap − A0 )/( A∞ − A0 ) ( С H O − C (Cl)M nTPP ⋅ (Ap − A0 )/( A∞ − A0 ))
0
2 2
where A0, A∞, Ap are the optical densities of the solutions of initial (Cl)MnTPP, of the com-
plex (Cl)(H2O2)MnIIITPP and of an equilibrium mixture at the working wavelength of
468 nm.
The scheme of the reactions conforming to kinetic equation (6) is more complex due
to the appearance of perhydroxyl HO2– in kinetically significant amounts at the increase of
the concentration of hydrogen peroxide:
K1
−
H2O2 H+ + HO 2 , (10)
− k −
(Cl)MnIIITPP + HO 2 ⎯⎯
2
→ (Cl)(HO 2 )MnIIITPP fast , (11)
K3
−
(Cl)(HO 2 )MnIIITPP (Сl)O=MnIV(TPP
+•
(TPP+·) ) + ОН- , (12)
+• − k
(TPP+·) ) + HO 2 ⎯⎯→
(Сl)O=MnIV(TPP 4
O=MnIVTPP + HCl + O2 slow , (13)
K5
−
ОН- + H2O2 HO 2 + H2O . (14)
It has been experimentally found that, due to the low concentration of manganese por-
phyrin in the form of (Cl)O=MnIV(TPP+·), there is no gas evolution in the experiment, and
the O2 formed is, evidently, in solution. The rate constant in equation (6) includes the equi-
librium constants (10), (12) and (14):
k3 = k4 ⋅ K3 ⋅ K5 ⋅ K1−1/ 2 . (15)
The numerical value of K1 is known: 2.4·10 –2 at 298 K [25]. The values of kv1 and
kv3, found by optimizing the dependences of the efficient rate constant of the reaction of
(Cl)MnIIITPP with H2O2 on the concentration of the latter, are given in Table 1 [8].
Table 1 True rate constants, E and ∆S≠, of the reaction of (Cl)MnTPP with H2O2.
CH2O2 range, mol/l T, K kv ·102 (a), E, ∆S≠,

s–1 ·mol –1 ·l kJ/mol J/(mol·K)
(Cl)MnTPP
0.017–0.10 288 1.41 70±1 –44±3
298 3.90
308 9.45
1.26–3.32 288 0.72 46±1 –133
298 1.34
308 2.51
(AcO)MnTPP
0.013–0.052 288 7 54±1 –87±3
298 15
308 31
(a)k
v1 and kv 3 for the CH2O2 ranges of 0.017–0.10 and 1.26–3.32, respectively.
The overall reaction of (Cl)MnTPP with H2O2 at all concentrations of the latter can be
presented as Scheme 2. At low concentrations of H2O2 the reaction is limited by the stage
of two-electron oxidation of manganese porphyrin and H2O2 reduction on metal in
(Cl)(H2O2)MnTPP (k2′); at high concentrations, by the stage of two-electron reduction of
π-cation radical (Cl)O=MnIV(TPP+·) up to O=MnIVTPP and Cl – and oxidation of HO–2
(k3′) with evolution of O2 and H+.
HO
Scheme 2 An overall scheme of the reaction of (Cl)MnTPP with H2O2.

Under conditions of H2O2 excess, the complex O=MnIVTPP oxidized by the central
atom of manganese is stable in solution in time.
It proved that the acetate complex (AcO)MnIIITPP (compound 11) reacts with H2O2
much faster than the chloride complex (Cl)MnIIITPP (Table 3). Already at CH2O2 equal to
1.0 mol/l the reaction is instantaneous, the reaction product in this case being O=MnIVTPP.
The order for [H2O2] for this reaction at low CH2O2 (0.013–0.052 mol/l) is close to unity
(0.72), which indicates the common character of its mechanism with that for (Cl)MnIIITPP
(Scheme 2, route K1′ → k2′). The rise of reactivity in the case of (AcO)MnIIITPP is explain-
able by a stronger binding of acidoligand AcO – as compared with Cl – . In the medium of
DMFA–H2O, where the reaction is run, in the initial complex at the coordination of H2O2
there occurs the substitution of the DMFA molecule coordinated to the sixth coordination
site. Owing to the effect of trans-influence of AcO – , substitution of DMFA is easier in the
(AcO)MnIIITPP complex than in (Cl)MnIIITPP. Besides, owing to the possibility of trans-
ferring the electronic effect of acidoligand via the transition metal with a partially filled
d-shell to the macrocycle, the macrocycle donates the electron easier to form π-cation rad-
ical in (AcO)MnIIITPP than in (Cl)MnIIITPP.
Manganese(III) porphyrins studied possess a catalase activity in an alkaline medium.
The results presented above show the ease of forming oxidized forms of complexes and the
absence of destruction of the macrocycle in the interaction with H2O2. This makes manga-
nese(III) porphyrins rather promising for their use as models of natural catalases.
New data on the spectral and kinetic studies of the reaction of (chloro)manganese(III)
octaethylporphin (Cl)MnOEP with hydrogen peroxide in water–DMFA media show that
what was said above regarding catalase models also pertains to the complex of manga-
nese(III) with another porphin derivative octaethylporphin.
The kinetics of the reaction of (Cl)MnOEP with H2O2 in an H2O–DMFA medium was
studied using a spectrophotometric method within the temperature range of 288–308 K.
The optical density of the solutions in the course of the reaction was measured at a wave-
length of 458 nm. The initial concentration of H2O2 in water was 17.4 ± 0.1 mol/l.
Interaction of (Cl)MnOEP with H2O2 at temperatures close to room temperature is ac-
companied with the change of its electronic absorption spectra. Herewith, the isobestic
points at 394, 446, 478, 523 and 582 nm are preserved (Fig. 2), which indicates the mutual
conversion of two stable stained compounds, one of which is the initial (Cl)MnOEP, the
form of existence of which differs at different concentrations of peroxide, as is seen from
the further discussion.
The intensity of the absorption bands of initial (Cl)MnOEP at 367, 458 and 541 nm
decreases, and at 434, 517 and 726 nm goes up. The end product was identified using liter-
ature data [26–28]. The spectrum of the end product corresponds to π-cation radical of man-
ganese(III) octaethylporphin (Fig. 2). The pattern of spectral changes presented in Fig. 2 is
characteristic of the entire range of H2O2 concentrations studied, which distinguishes the
octaethylporphin complex from above-considered (Cl)MnTPP, for which the similar reac-
tion was observed to yield various end products at different CH2O2. The first order of the
reaction of (Cl)MnOEP with H2O2 with respect to metalloporphyrin is demonstrated by the
data of Fig. 3. The efficient rate constants of the reaction are given in Table 2.
The order of the reaction with respect to [H2O2] is variable. Experimentally, we found
three ranges of peroxide concentrations: 0.01–0.07, 0.2–0.52 and 1.00–2.22 mol/l, each of
which was observed to have its own order for hydrogen peroxide (n) (Figs. 4 and 5): by the
T
slope of the lines in the coordinates log kef – log CH2O2, n is, respectively, 1/2, 0 and –1/2.
The zero order is evident due to the absence of the dependence of kef on CH2O2 (Table 2).
λ, nm
Figure 2 Electronic absorption spectra of (Cl)MnOEP in the course of the reaction with H2O2.
298 K, CH2O2 = 1.0 mol/l.
ln
τ, s
Figure 3 A dependence of ln(C0 /Cτ) – τ for the reaction of (Cl)MnOEP with H2O2. C(Cl)MnIIIOEP
= 2·10 –5 mol/l. CH2O2 = 0.33 (1), 0.49 (2), 0.05 (3), 1.35 (4), 0.03 (5), 0.01 (6) mol/l. T, K: 308 (1),
303 (2), 298 (3), 293 (4, 5, 6). R2 = 0.99.
The considered results make it possible to write down the experimentally found com-
plete kinetic equations for the first, second and third reacting sequences, respectively:
−dC(Cl)Mn III OEP / dτ = kv1 ⋅ C(Cl)Mn IIIOEP ⋅ C1/2

Н О
, (16)
2 2
−dC(Cl)Mn III OEP / dτ = k v2 ⋅ C(Cl)Mn III OEP , (17)
−1/2 . (18)
− dC(Cl)Mn IIIOEP / dτ = k v3 ⋅ C(Cl)Mn IIIOEP ⋅ CН О2 2
Table 2 Efficient kinetic parameters of the reaction of (Cl)MnOEP with H2O2 in DMFA.
CH2O2, kef ·104, s –1 E′, kJ/mol ∆S≠ ′,

mol/l J/(mol·K)
293 K 298 K 303 K 308 K
0.01 2.9±0.2 4.6±0.1 7.2±0.3 11.0±0.4 67±1 –91±3

0.03 5.0±0.1 8.1±0.3 12.3±0.2 17.8±0.3 63±2 –100±7
0.05 6.1±0.2 10.0± 0.4 14.6±0.5 22.9±0.4 65±2 –91±7
0.07 8.6±0.5 12.2±0.4 18.8±0.8 25.7±1.0 56±2 –120±7
0.20 10.2±0.4 16.4±0.7 25.0±1.0 32.6±0.9 59±4 –107±13
0.33 10.3±0.6 16.4±0.6 24.0±1.3 31.3±0.8 56±4 –118±13
0.49 9.8±0.5 15.8±0.4 20.7±0.6 31.7±0.6 57±4 –115±13
0.52 10.0±0.7 15.9±0.8 22.5±1.1 30.3±1.0 55±3 –121±10
1.00 9.5±0.2 13.1±0.5 17.8±0.5 25.8±0.5 50±2 –140±7
1.35 8.5±0.3 10.9±0.3 16.3±0.6 22.3±0.5 49±3 –144±10
1.72 7.3±0.2 9.9±0.2 14.9±0.5 19.7±0.6 51±2 –138±7
2.22 6.1±0.3 8.1±0.2 11.4±0.5 16.6±0.5 50±2 –143±7
log kTe
-log CH2O2
Figure 4 A dependence of log kef T on –log C

H2O2 for the reaction of (Cl)MnOEP with H2O2 within
the H2O2 concentration range of 0.01–0.07 mol/l. T, K: 308 (1), 303 (2), 298 (3), 293 (4). R2 = 0.98.
log kTe
log CH2O2
Figure 5 A dependence of log kefT on log CH2O2 for the reaction of (Cl)MnOEP with H2O2 within
the H2O2 concentration range of 1.00–2.22 mol/l. T, K: 308 (1), 303 (2), 298 (3), 293 (4). R2 = 0.98
(1, 3, 4), 0.92 (2).
The numerical values of the true rate constants kv1 and kv3, found by optimizing the
dependences presented in Figs. 4 and 5, and the reaction activation parameters are given in
Table 3.
Table 3 True rate constants, activation energy and entropy of the reaction of (Cl)MnOEP with
H2O2 in DMFA.
CH2O2 range, T, K kv ·103, E, kJ/mol ∆S≠,

mol/l s–1 ·mol –1 ·l J/(mol·K)
0.01–0.07 293 3.0±0.1

298 4.6± 0.1 63±1 –85±3
303 7.2±0.1
308 10.4±0.4
1.00–2.22 293 0.95±0.02
298 1.27±0.02 50±2 –140±6
303 1.8±0.1
308 2.55±0.02
Considering the spectral data (Fig. 2) and taking into account equation (16), the fol-
lowing scheme of elementary stages can be proposed for the first range of H2O2 concentra-
tions. The scheme is supported by comparing experimental equation (16) with the
theoretical equation deduced for the system of equations (19–21).
K1
−
H2O2 Н+ + НО 2 , (19)
K2
− −
(Cl)MnIIIOEP + НО 2 (Cl)(НО 2 )MnIIIOEP , (20)
−
(Cl)(НО 2 )MnIIIOEP
k3
⎯⎯→
.
O=MnIII(OEP + • ) + ОН- + Cl- slow . (21)
Perhydroxyl anion present in the system at equilibrium concentrations in accordance

with the equilibrium (19) is added reversibly (reaction (20)) to the coordinatively unsatur-
ated Mn atom, thus forming the donor–acceptor bonds –O – → Mn. Hydrogen peroxide is
coordinated by manganese(III) octaethylporphin not in the form of the H2O2 molecule as
in the case of (X)MnTPP complexes, but as a stronger nucleophile HO–2 . The cause is, ap-
parently, a decrease of the positive charge δ+ at the Mn atom in transition from the tetraphe-
nylporphin complex to the octaethylporphin complex due to the electron–donor action of
eight β-alkyl substituents and the elimination of the electron–acceptor effect of the meso-
phenyl groups. The perhydroxyl ion activated owing to the coordination at Mn donates the
oxygen atom to metalloporphyrin with abstraction of OH – in the slow reaction (21) of
two-electron oxidoreduction. (Cl)MnOEP passes into the oxidized form O=MnIII(OEP+·),
which is well identifiable by the electronic absorption spectra (Fig. 2). Due to the low con-
centration of HO–2 , no interaction of the oxidized form of the complex with the second mol-
ecule of peroxide (as is the case in all catalytic processes of H2O2 disproportionation) is
observed.
Coordination of HO–2 (reaction (20)) evidently occurs via the sixth coordination site;
CH2O2, mol/l
Figure 6 A curve of the titration of the complex (Cl)MnIIIOEP by hydrogen peroxide in DMFA.
acidoligand Cl remains to stay in the coordination sphere. This is confirmed by the example
of (X)MnTPP, where the rate of the process was found to strongly depend on the nature of
X (see above).
The rate equation for the limiting stage has the form:
− dC(Cl)(HO − )Mn IIIОЕP / dτ = k3 ⋅ C(Cl)(HO− )Mn IIIОЕP . (22)

2 2
After expressing C(Cl)HO–2)MnIIIOEP via the equilibrium constant K2 and then CHO–2 via
the equilibrium constant K1, we obtain equation (23):
− dC(Cl)(HO − )Mn IIIОЕP / dτ = − dC(Cl)Mn IIIOEP / dτ =

2
k3 ⋅ K 2 ⋅ C(Cl)Mn IIIОЕP ⋅ C1/2

H O
⋅ K11/ 2 . (23)
2 2
The first equality in equation (23) is written with account for equilibrium (20). Thus,
from the comparison of equations (20) and (23), we have
k ν1 = k3 ⋅ K11/2 ⋅ K 2 , (24)
where K1 = 2.4·10 –12 at 298 K [25].

To determine the numerical value of K2, we carried out the spectrophotometric titra-
tion of the solution of (Cl)MnIIIOEP in DMFA by hydrogen peroxide (Fig. 6).
The titration curve has one stage. From the slope of the curve in the coordinates “the
logarithm of the ratio of the equilibrium concentration of (Cl)(HO–2)MnIIIOEP to the equi-
librium concentration of (Cl)MnIIIOEP (indicator ratio) vs the logarithm of CH2O2 to the ab-
scissa axis”, the number of added H2O2 is 1/2. The concentration constant of the
equilibrium of coordination of molecular peroxide, which can be equated to the thermody-
namic constant with account for large dilutions in the reaction system, is, at 298 K,
(2.2±0.3)·10 –5 mol –1/2 ·l1/2. Hence, the equilibrium constant (equation 20) K2 =
(9±1)·106 mol –3/2 ·l3/2 provided that for the above constant K1 the value is given in molar
scale, which does not explicitly follow from the results of [25]. The rate constant of the el-
ementary reaction (21) at 298 K is k3 = kv1/(K1/2 –4 –1
1 · K2) = (3.3±0.3)·10 s . Note that the
dimensionality of the constant k3 corresponds to equations (21) and (22), which confirms
the condition put forward above.
The third range of H2O2 concentrations with experimental kinetic equation (18) needs
the account for the following five reactions:
K1
−
H2O2 Н+ + НО 2 , (25)
− k2 −
(Cl)MnIIIOEP + НО 2 ⎯⎯→ (Cl)(НО 2 )MnIIIOEP fast , (26)
−
(Cl)(НО 2 )MnIIIOEP
K3 .
(Cl)O=MnIV(OEP + • ) + ОН- , (27)
K4 −
ОН- + Н2О2 НО 2 + Н2О , (28)
2(Cl)O=MnIV(OEP+ .) + HO2-⎯⎯→
k5 .
2O=MnIII(OEP + • ) + HCl + O2 + Cl- slow . (29)
Due to the low concentration of HO–2 , no gas evolution is observed in the experiment,
and O2 formed is, evidently, in solution.
The first stage involving manganese porphyrin (26) – coordination of hydrogen per-
oxide in the form of HO–2 to the sixth coordination position of the manganese atom – occurs
at higher concentrations of H2O2 rapidly and irreversibly. The transfer of two electrons
from the readily polarized porphyrin macrocycle and metal cation to the coordinated mol-
ecule of peroxide proceeds in the equilibrium stage (27). Probably, this reaction is made
equilibrium by the stability of π-cation radical of (Cl)O=MnIV(OEP+·) decreased as com-
pared with O=MnIII(OEP+·) (equation (21)), and the presence of higher concentrations of
negative ions HO–2 and OH –, which prevent the ionization of acidoligand Cl – . The latter
remains in the coordination sphere of π-cation radical (Scheme 3). Finally, the reaction with
the second H2O2 (in the form of HO–2) becomes possible; the reaction proceeds slower than
reaction (26) and limits the process. The reduction of π-cation radical of manganese(IV)
porphyrin does not proceed to the end, that is to the relatively stable O=MnIVOEP, but stops
at the one-electron reduced form O=MnIII(OEP+·), which is registered in the experiment as
a relatively stable end product. This is one more distinction from the (Cl)MnIIITPP–H2O2
system, where the end product is a two-electron reduced form.
We used the word “relatively” when assessing the stability. The cation-radical form
O=MnIII(OEP+·) upon isolation into chloroform after the reaction of (Cl)MnOEP with
H2O2 and washing off excess H2O2 still passes into the MnIV complex: bands at 394, 450
(arm), 514, 546 and 667 nm appeared in the spectrum, which is indicative of the formation
of O=MnIVOEP. In turn, this compound, if kept still in solution for 24 h, slowly passes into
manganese(III) porphyrin with a characteristic band in the region of 473 nm (the fact known
in the chemistry of manganese porphyrins [26]).
The rate equation for the limiting stage has the form:
− dC(Cl)O = Mn IV (OEP+ .• ) / dτ = k5 ⋅ C(Cl)O = Mn IV (OEP+.• ) ⋅ CHO − . (30)

2
After successively expressing C(Cl)=MnIV(OEP+·) via the equilibrium constants K3

(equation 31) and K4 (equation 32), we obtain equation (33):
Scheme 3 Assumed structure of π-cation radical of (Cl)O=MnIV(OEP+·).
−1
C(Cl)O= Mn IV (OEP +•. ) = K3 ⋅ C(Сl)(HO− )Mn IIIOEP ⋅ CОН −, (31)
2
2 −1
CОН − = СHO − ⋅ K4 , (32)
2
−2
C(Cl)O= Mn IV (OEP +•.) = K3 ⋅ C(Сl)(HO− )Mn IIIOEP ⋅ K4 ⋅ CHO − . (33)
2 2
With account for the equilibrium (25), we express CHO2–:
CHO− = K11/ 2 ⋅ C1/2

H O
. (34)
2 2 2
Upon substitution of the expressions for concentrations, (33) and (34), into equation
(30), we obtain equation (35), which is consistent with the found experimental equation
(18):
− dC(Cl)O = Mn IV (ОЕP +•. ) / d τ = − dC(Cl)Mn III OEP / d τ =
k5 ⋅ K 3 ⋅ K 4 ⋅ C (Cl)(HO − )Mn III OEP ⋅ C −1 − = k5 ⋅ K 3 ⋅ K 4 ⋅ K11/ 2 ⋅ C (Cl)Mn III OEP ⋅ C H−1/2

O . (35)
2 HO 2 2 2
Thus,
k ν3 = k5 ⋅ K3 ⋅ K 4 ⋅ K11/ 2 , (36)
where K1 = 2.4·10 –12 at 298 K.

Change of details of the mechanism of the reaction of the octaethylporphin complex
with H2O2 as compared with complexes with H2TPP is due to the difference in the elec-
tronic state of the aromatic ligand. The proof is the result of comparing the quantitative
characteristics of the reactions. The (Cl)MnTPP reacts with H2O2 much faster than
(Cl)MnOEP. The true rate constant is by an order higher in the case of (Cl)MnTPP both at
low and high concentrations of H2O2, and the reaction of (Cl)MnTPP with H2O2 has more
negative values of ∆S≠ (Tables 3 and 5).
In the case of (Cl)MnTPP, the π-cation radical form is formed only at low concentra-
tions of H2O2, and at CH2O2 > 0.1 mol/l the end product of the reaction is manganese(IV)
tetraphenylporphin – the form oxidized at Mn atom. In the case of (Cl)MnOEP, the end
product of the reaction is π-cation radical of manganese(III) octaethylporphin within the en-
tire range of H2O2 concentrations. Hence, it follows that, by changing the electronic state
of the macrocycle and of Mn–N bonds, we can achieve optimal parameters for the coordi-
nation reaction of H2O2 molecules and HO2– ions at various stages of the reaction with per-
oxide and then use them in the development of synthetic catalases.
2.2 Kinetics of peroxide disproportionation in the presence of manganese(III)

porphyrins
This section presents the results of studies of the kinetics of H2O2 homogeneous decompo-
sition reaction by the volumetric and spectrophotometric methods in the presence of man-
ganese(III) complexes with octaethylporphin unsubstituted in meso-positions and meso-
phenyloctaethylporphins and acidoligands Cl – , AcO – , SCN – (compounds 5 – 9).
The reaction was carried out in a DMFA–KOH–H2O medium at temperatures of
343–363 K in a reactor equipped with a jacket for thermostatting and a magnetic stirrer,
under conditions of intensive stirring. An aqueous solution of hydrogen peroxide was added
to a prepared solution of the manganese complex and KOH in DMFA. The concentrations
of the complex, KOH and H2O2 were varied within the range of 10 –6 –10 –5, 0.73–3.65 and
2.61–4.79 mol/l, respectively. The initial concentration of H2O2 in water (17.43±0.06
mol/l) was determined by iodometric titration.
Table 4 presents the experimental values of the rate constants for the hydrogen perox-
ide decomposition reaction of formal zero order, found as a slope of the linear portion of
the dependence of VO2 – τ to the positive direction of the abscissa (Fig. 7). The figure also
presents the catalytic activity (A) obtained by dividing the value of W on the known con-
centration of the catalyst – the manganese complex.
Table 5 presents activation energies determined by the slope of the curves in the coor-
dinates log W – 1/T, and activation entropies calculated by formula (37):
ml
τ, min
Figure 7 A dependence of the volume of evolving O2 on time in the course of the H2O2 decompo-
sition reaction. C(Cl)MnMPOEP = 1.08·10 –5 (1), C(Cl)MnOEP = 2.77·10 –5 (2), C(AcO)MnOEP =
2.25·10 –5 (3), C(Cl)Mn5,15DPOEP = 3.87·10 –5 mol/l (4), CH2O2 = 3.49 mol/l, CKOH = 1.5·10 –2 mol/l.
T = 343 K.
Table 4 Decomposition rates of H2O2 (W) and catalytic activity (A) of metalloporphyrins at 343 K.
Catalyst CH2O2, mol/l(a) W, ml O2/min A, s –1
(Cl)MnOEP 2.61 0.45±0.02 0.28±0.01

3.49 0.83±0.02 0.52±0.01
4.36 1.26±0.03 0.78±0.02
4.79 1.54±0.08 0.96±0.05
(AcO)MnOEP 2.61 0.40±0.01 0.67±0.02
3.49 0.74±0.01 1.25±0.02
3.87 1.08±0.05 1.82±0.08
4.36 1.35±0.08 2.3±0.1
4.79 1.45±0.07 2.45±0.11
(SCN)MnOEP 2.69 0.44±0.01 0.57±0.01
3.14 0.55±0.02 0.72±0.02
3.58 0.74±0.02 0.97±0.02
4.03 1.07±0.03 1.40±0.04
4.48 1.31±0.07 1.72±0.09
(Cl)MnMPOEP 0.45 0.97±0.03 12.7±0.4
0.72 1.45±0.04 12.0±0.3
1.08 1.86±0.06 10.2±0.3
(Cl)Mn5,15DPOEP 2.58 0.33±0.01 0.20±0.01
3.44 0.73±0.02 0.45±0.01
3.87 0.83±0.02 0.51±0.01
4.30 1.12±0.04 0.69±0.02
4.73 1.31±0.04 0.80±0.04
Catalyst CKOH ·10 –2, mol/l(b) W, ml O2/min A, s –1
(Cl)MnOEP 0.73 0.45±0.02 0.28±0.01

1.46 0.83±0.02 0.52±0.01
2.19 1.28±0.03 0.79±0.02
2.56 1.42±0.05 0.88±0.02
2.92 1.68±0.09 1.04±0.05
3.65 2.0±0.1 1.26±0.06
(AcO)MnOEP 0.73 0.47±0.01 0.79±0.02
1.46 0.74±0.02 1.25±0.02
2.19 1.09±0.06 1.8±0.1
2.56 1.51±0.08 2.6±0.1
2.92 1.62±0.08 2.7±0.1
3.65 2.14±0.09 3.61±0.15
(Cl)Mn5,15DPOEP 0.78 0.40±0.01 0.24±0.01
1.55 0.73±0.02 0.45±0.01
2.33 1.21±0.06 0.74±0.03
2.71 1.4±0.1 0.86±0.06
3.10 1.52±0.08 0.93±0.05
3.88 2.0±0.1 1.25±0.06
Table 4 Decomposition
(continued) rates of H2O2 (W) and catalytic activity (A) of metalloporphyrins at 343 K.
Catalyst Ccatalyst ·105, mol/l(c) W, ml O2/min A, s –1
– – 0.62±0.02 –
(Cl)MnOEP 0.095 0.76±0.01 47±1
0.19 0.83±0.02 26±1
0.38 0.84±0.06 13±0.6
2.77 1.58±0.06 3.3±0.1
5.11 2.10±0.06 2.4±0.1
(AcO)MnOEP 0.07 0.74±0.01 63±1
0.14 0.74±0.04 31±2
0.35 0.73±0.03 12±0.5
2.25 1.29±0.04 3.4±0.1
(SCN)MnOEP 0.09 0.73±0.03 48±2
0.18 0.73±0.02 24±0.6
0.32 0.75±0.03 14±0.5
0.45 0.78±0.05 10.3±0.6
2.71 1.48±0.05 3.2±0.1
(Cl)MnMPOEP 0.45 0.97±0.03 12.7±0.4
0.72 1.45±0.04 12.0±0.3
1.08 1.86±0.06 10.2±0.3
(Cl)Mn 5,15DPOEP 0.097 0.76±0.02 46±1
0.48 0.73±0.03 9.0±0.4
0.97 0.76±0.02 4.6±0.1
3.87 1.05±0.02 1.61±0.03
8.53 1.52±0.03 1.06±0.02
MnCl2 0.60 1.26±0.04 12.4±0.4
1.43 1.3±0.1 5.5±0.5
5.95 1.4±0.3 1.5±0.3
11.90 1.7±0.2 0.8±0.1
(a)C –2 –5
KOH = 1.5·10 mol/l, C(X)MnP = 0.2·10 mol/l,
(b)C –5
H2O2 = 3.49 mol/l, C(X)MnP = 0.2·10 mol/l,
(c)CH2O2 = 3.49 mol/l, CKOH = 1.5·10 –2 mol/l.
Table 5 Catalytic H2O2 decomposition reaction rates at various temperatures and activation
parameters. C(X)MnP = 0.2·10 –5 mol/l, CH2O2 = 3.49 mol/l, CKOH = 1.5·10 –2 mol/l.
Catalyst T, K W, ml O2/min A, s –1 E, kJ/mol ∆S≠,

J/(mol·K)
(Cl)MnOEP 343 0.83±0.02 0.52±0.01 64±4 –67±13

348 1.16±0.08 0.71±0.05
353 1.74±0.12 1.05±0.07
358 2.27±0.15 1.35±0.08
363 2.75±0.20 1.61±0.11
Table 5 Catalytic H2O2 decomposition reaction rates at various temperatures and activation
C(X)MnP = 0.2·10 –5 mol/l, CH2O2 = 3.49 mol/l, CKOH = 1.5·10 –2 mol/l.
Table 5 (continued)
parameters.
(AcO)MnOEP 343 0.74±0.01 1.25±0.02 63±2 –71±7

348 0.95±0.03 1.58±0.05
353 1.38±0.08 2.26±0.13
358 1.86±0.10 3.01±0.16
363 2.44±0.09 3.90±0.15
(SCN)MnOEP 343 0.74±0.02 0.97±0.02 65±3 –65±10
348 0.91±0.06 1.17±0.07
353 1.27±0.04 1.62±0.05
358 1.80±0.15 2.26±0.18
363 2.46±0.18 3.05±0.22
(Cl)Mn5,15DPOEP 343 0.73±0.01 0.45±0.01 82±3 –16±10
348 1.00±0.05 0.60±0.03
353 1.61±0.10 0.96±0.06
358 2.23±0.15 1.31±0.09
363 3.53±0.20 2.04±0.11
E ± ∆E
∆S ≠ = 19.1 ⋅ log W + − 19.1 ⋅ log T − 205 . (37)
T
It is seen from Table 4 that the decomposition rate of hydrogen peroxide increases with
the rise of the concentrations of the manganese(III) complex, KOH and H2O2 in the system
on condition of a significant excess of the other reagents as compared with the complex.
There is a satisfactory correlation between W and the catalyst concentration, which
points to the first order of the reaction with respect to this component (Fig. 8). A similar
dependence for CH2O2 is not linear. Figure 9 demonstrates a correlation in the logarithmic
coordinates log W – log CH2O2 with the slopes of the curves close to 2 (n = 2.01 for
(Cl)MnOEP, n = 2.26 for (Cl)Mn5,15DPOEP). A similar dependence for COH – is also linear,
with the slope α close to unity (Fig. 10). Thus, according to the data of the experiment, we
can write down the equation for the reaction rate:
W, ml O2 / min
Ccatalyst, 10-5, mol/l
Figure 8 A dependence of the H2O2 decomposition rate on the concentration of (Cl)MnMPOEP

(1), (Cl)MnOEP (2), (AcO)MnOEP (3), (Cl)Mn5,15DPOEP (4). CH2O2 = 3.49 mol/l, CKOH =
1.5·10 –2 mol/l, T = 343 K. R2 = 0.97–0.99.
logW logW
logCKOH
logCH2O2
Fig. 9 Fig. 10
Figure 9 A dependence of log W on log CH2O2 for the reaction of H2O2 decomposition in the pres-
ence of (Cl)MnOEP (1) and (Cl)Mn5,15DPOEP (2). C(X)MnP = 0.2·10 –5 mol/l, CH2O2 = 3.49 mol/l,
CKOH = 1.5·10 –2 mol/l, T = 343 K. R2 = 0.99.
Figure 10 A dependence of the logarithms of the rates of H2O2 decomposition in the presence of
(Cl)MnOEP (1) and (Cl)Mn5,15DPOEP (2) on the logarithms of KOH concentration. C(X)MnP =
0.2·10 –5 mol/l, CH2O2 = 3.49 mol/l, CKOH = 1.5·10 –2 mol/l, T = 343 K. R2 = 0.99.
2
dCO2 / dτ = k ⋅ Ccatalyst ⋅ СН 2О 2
⋅ COH - ⋅ CO0 . (38)
2
The numerical values of the true rate constants k are calculated as arithmetic means of
three true constants obtained in optimizing the dependences in coordinates W–Ccatalyst,
log W–log CKOH and log W–log CH2O2 at integer values n, respectively, 1, 1 and 2. The con-
stants k for (Cl)MnOEP, (AcO)MnOEP and (Cl)Mn5,15DPOEP are, respectively,
(1.73±0.06), (1.64±0.17) and (1.49±0.07) l4 ·mol –3 ·s –1.
A special spectrophotometric study of the interaction of (Cl)MnOEP with H2O2 in a
DMFA–H2O medium in the absence of alkali, the results of which are considered in the pre-
vious section, made it possible to reveal in the entire range of H2O2 studied (0.01–3.74
mol/l) a manganese(III) porphyrin → π-cation radical of manganese(III) porphyrin transi-
tion (Fig. 2).
With account for the data considered, we can write down the assumed scheme of con-
versions in the course of the catalytic reaction of H2O2 decomposition (equations
(39)–(43)).
K1
(Х)MnIIIOEP + H2O2 (Х)(H2O2)MnIIIOEP , (39)
(Х)(H2O2)MnIIIOEP
k2
⎯⎯→
.
O=MnIII(OEP + • ) + H2O + Х- fast , (40)
K3 −
H2O2 + OH- HO 2 + H2O, (41)
.
O=MnIII(OEP + • ) + HO 2−
K4
(HO 2 )O=MnIII(OEP + • ),
− . (42)
(HO 2 )O=MnIII(OEP + • )
− . k5
⎯⎯→ [MnIIIOEP]+ + H2O + O2 slow. (43)
For the limiting stage (43), we can write down the kinetic equation:
−dC(HO- )O=Mn III (OEP+•.) / dτ = dCO2 / dτ = k5 ⋅ C(HO- )O=Mn III (OEP+•.) . (44)
2 2
With account for the equilibrium (42), we express C(HO2-)O=MnIII(OEP+·) :
C(HO- )O=Mn III (OEP+•.) = K4 ⋅ CO=Mn III (OEP+•.) ⋅ CHO- . (45)

2 2
We write down equalities (46) –(48) for reactions (41), (40) and (39), respectively:
СHO- = K3 ⋅ CH 2O2 ⋅ COH - . (46)

2
СO=Mn III (OEP+•.) = C(Х)(H O )Mn IIIОЕP . (47)

2 2
C(Х)(H O )Mn IIIОЕP = K1 ⋅ C(Х)Mn IIIOEP ⋅ CH 2O2 . (48)

2 2
Let us perform the substitution of the expressions for CHO-2 and CO=MnIII(OEP+·) also
into equation (45):
2
C(HO- )O=Mn III (OEP+•.) = K 4 ⋅ K1 ⋅ K3 ⋅ C(Х)Mn III OEP ⋅ CH 2 O2
⋅ COH - (49)
2
After substituting the expression for C(HO-2 )O =MnIII(OEP+·) (equation (49)) into (44), we
obtain equation (50), which is consistent with experimentally found equation (38):
2
dCO2 / dτ = k5 ⋅ K 4 ⋅ K1 ⋅ K3 ⋅ C(Х)Mn IIIOEP ⋅ CH 2 O2
⋅ COH - . (50)
From the comparison of equations (38) and (50), it follows that k = K1 · K3· K4· k5.
In accordance with equations (39)–(43), the catalytic reaction of H2O2 decomposition
proceeds with the participation of the catalyst, which changes its redox states, and two per-
oxide units (in the form of the H2O2 molecule and perhydroxyl anion HO2– ). At the first
stage, H2O2 is coordinated to the sixth coordination site (X)MnIIIOEP; as the result, the
O–O bond, which is not too strong as it is (200 kJ/mol), is activated [25]). The coordinated
molecule of H2O2 breaks down to evolve H2O and to eliminate two electrons from metal-
loporphyrin. The latter passes into a π-cation radical form, being oxidized via the macro-
cycle and metal, which remains triple-charged only formally (O=MnIIIOEP+·) R
[O=MnIVOEP]+), as, for instance, in the known stable oxidized forms of diphthalocyanines
of lanthanides LnIIIPc2· [29]. The second molecule of peroxide is added as a more nucleo-
philic particle HO–2. The formed π-cation radical of the complex, being in a highly coordi-
nated state, slowly reduces with abstraction of molecular O2. It should be emphasized that
no manifestations of the radical mechanism of H2O2 decomposition have been revealed in
support of the considered mechanism, in particular, no slow stages of reaction initiation.
Variation of the ligand, which is in the fifth coordination position, with the view to
confirm the provisions of the mechanism of the catalytic action of manganese(III) porphy-
rins, gave the following results. The catalytic activity of manganese(III) octaethylporphins
(Table 6) is essentially not dependent on the nature of acidoligand (Cl, AcO, SCN), which
experimentally confirms the proposed stage-wise ion-molecular mechanism (equations

(39)–(43)). The rate constants of the limiting stage, k5, do not depend on the nature of X
(equation (43)), and the constant K1 is present in the expression for the rate (50) as a con-
stant value, apparently differing for different X insignificantly.
In earlier works [9], we proposed another form of recording the intermediate oxidized
state of the catalyst formed from (X)(H2O2)MnIIIP (P, dianion of H2TPP). The one-electron
oxidized form of (Cl)(OH·)MnIII(TPP+·) formed in abstraction of OH – from coordinated
H2O2 was assumed. Evidently, this particle is unstable and should pass into a form with bi-
dentate coordinated oxygen O=MnIII(TPP+·).
Among the manganese(III) porphyrins studied, the highest catalytic activity in the
H2O2 decomposition reaction is manifested by (Cl)MnMPOEP; the lowest, by the diphe-
nyl-substituted complex. The activity of (Cl)Mn5,15DPOEP is comparable with that of
MnCl2 (Table 6).
Thus, introduction of one meso-phenyl group into (Cl)MnOEP leads to a significant
activation of the catalase ability of the complex. This result is expectable within the frame-
work of the expounded mechanism: a decrease of electronic density in the macrocycle at
the substitution by phenyl group contributes to an increase of the positive charge at man-
ganese, which leads to the reduction of (HO–2)O=MnIII(OEP+·) in the limiting stage (43).
However, introduction of the second phenyl group (transition (Cl)MnMPOEP →
(Cl)Mn5,15DPOEP) not only fails to increase the catalytic activity, but decreases it to the
level of manganese salts (Table 6). This could be due only to the worsening of the coordi-
nation conditions of the second molecule of peroxide in trans-position with respect to O2–
owing to the distortions, which, apparently, the (Cl)Mn5,15DPOEP molecule experiences
like similar copper(II) complexes. According to [30, 31], the coordination centre of the cop-
per complex of octaethylporphin remains planar at the introduction of one meso-phenyl
substituent and is significantly distorted in Cu5,15DPOEP.
Thus, the catalase activity of manganese(III) porphyrins can be both enhanced and re-
duced to zero depending on the structure of the macrocycle. Probably, understanding of the
mechanism of the process would facilitate prediction of this property for other metallopor-
phyrins and catalytic systems.
3 Conclusion
The catalytic-action mechanism of manganese(III) porphyrins expounded above repeats in
many ways the mechanism of action of natural catalases. The general regularities of con-
versions involving the enzymes hemoproteins have been discussed already in the mono-
graph [25]. The enzyme catalase, which has a prosthetic group of iron protoporphyrin, is
bound to the protein by means of one coordination link (one coordination site of Fe) and
via propionic acid residues. H2O2 in the amount of several micromoles per litre is either
decomposed or reacts with other particles. With iron protoporphyrin, hydrogen peroxide
forms, apparently owing to coordination to the sixth coordination position, complexes of
three types: Fe·H2O2, Fe·OOH and Fe·OOH –, the last of which is active as catalase. This
complex further reacts with another molecule of H2O2 or other active particles. Thus, the
expounded conclusions regarding the catalytic-action mechanism of manganese(III) por-
phyrins obtained in our work basically correspond to the data for the mechanism of natural
catalases. Complexes of natural catalases and peroxidases have not been isolated individu-
ally due to their instability. The use of simpler models, as shown by our study, makes it pos-
sible to describe intermediate complexes spectrally. Besides, a possibility appears to
regulate and, which is more important, increase the catalytic activity of catalases by modi-
fying the aromatic moiety of their molecules.
The work was supported by grants from the Programme for Basic Research of the Rus-
sian Academy of Sciences “Elaboration of Methods for Producing Chemical Substances
and Development of Novel Materials” (No 8, 2006); from the Analytical Departmental Tar-
get-oriented Programme “Development of Higher-School Scientific Potential”
(2006–2008). Activity 2: Scientific Methodological Provision of the Development of High-
er-school Science Infrastructure”; from RNP 2.2.1.1.7181 “Development of the Mecha-
nisms of Integrating Ivanovo State University and Institute of Problems of Chemical
Physics RAS”; and from the Russian Foundation for Basic Research 06-03-96343-r.
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4 Complexation of Porphyrins
with Ions and Organic
Molecules
N.Zh. Mamardashvili1, V.V. Borovkov2,

G.M. Mamardashvili1, Y. Inoue2 and O.I. Koifman1
1Institute
of Solution Chemistry, Russian Academy of
Sciences, 1 Akademicheskaya Street, Ivanovo, 153045, Russia
ngm@isc-ras.ru
2Entropy Control Project, ICORP, Japan Science and
Technology Agency, Kamishinden 4-6-3, Toyonaka-shi,
Osaka 560-0085, Japan
victrb@inoue.jst.go.jp, inoue@chem.eng.osaka-u.ac.jp
Among the great diversity of host–guest interactions, a special place is occupied

by the complexation processes of porphyrins and related chromophores with ions
and organic molecules of various nature. This chapter considers the major
principles, driving forces and factors, which enable an efficient control of
supramolecular systems formed in this process.
Keywords: porphyrin, complexation, conformational correspondence, conformity, multi-

point binding, chirality, circular dichroism
Introduction
At present, the complex-forming properties of porphyrins are understood to be both the abil-
ity of porphyrins to enter into complexation reactions with metal cations to form metal-
loporphyrins and the ability of porphyrins to form supramolecular complexes. While
metalloporphyrin formation processes have been fruitfully studied for many decades, works
on supramolecular complexes of porphyrins having no less than two binding points, ap-
peared in the literature only in the recent 10 to 15 years. Presented material generalizes and
118 N.Zh. Mamardashvili, V.V. Borovkov, G.M. Mamardashvili, Y. Inoue and O.I. Koifman
analyzes the literature data and own results by the authors on supramolecular complexes of
porphyrins, showing the main trends of the development of the modern supramolecular
chemistry of porphyrins.
1 Complexation of Porphyrins with Ions

One of the essential coordination properties of porphyrins and their analogues is their ability
to coordinate virtually with all ions of metals in the periodic system of elements to form
especially strong intracomplex salts. Complexation of porphyrins with metal cations, ac-
cording to [1], occurs as the result of bimolecular collision of the ligand molecule and sol-
vated salt: activation of solvation complexes of octahedral structure as the determining
factor includes a partial decomposition of the first coordination sphere of metal, i.e., for-
mation of two coordination vacancies in cis-position of the octahedral complex. The reac-
tion, as a rule, is of the first kinetic order with respect to porphyrin and metal salt. Salts of
two-charge ions of d-metals interact the most readily with porphyrins in a medium of weak-
ly coordinating organic solvents – carboxylic acids and lower alcohols. The rate of the re-
action very little depends on the anionic composition of a salt. Thus, very strong complexes,
in which anion is bonded by an ionic-covalent bond (PdCl2, HgI2 etc) react with porphyrins
at rates characteristic of almost all ionic salts ( (AgNO3, Cu(NO3)2, FeCl2 etc). This indi-
cates that not anions but molecules of a solvating solvent go from the coordination sphere
of a salt in transition state. Anion of the salt leaves the coordination sphere beyond the peak
of the potential barrier.
It is seen in Table 1 that small and acyclic molecules (CH3OH, C2H5OH, CH3COOH,
acetone) are eliminated more readily and, as a rule, with lower energy and entropy of acti-
vation; large and cyclic molecules (pentanol, ethyl acetate, dioxane, pyridine) are hard to
eliminate. In solvents of the first group, metalloporphyrins are formed much faster and eas-
ier than in those of the second group. Optimal solvents for complexation are monocarbox-
ylic acids. At a high temperature, a good medium to form metalloporphyrins is pyridine.
Structural peculiarities of porphyrin molecules also have a strong effect on the rate of
the process. At the stage of activation of the reaction, the porphyrin structure undergoes
very significant changes, in particular, loses two central hydrogen atoms as protons, which
Table 1 Rate constants (k) and activation parameters (E and ∆S≠ ) of the complexation of
copper(II) cation with chlorophyllic acid in organic solvents at 298 K [1].
Solvent k, E, ∆S ≠,
l/(mol·s) kJ/mol J/(mol·K)
Methanol 4.45 58 –17

Ethanol 1.65 44 –96
n-Pentanol 0.55 113 126
Acetone 0.90 46 –67
Dioxane 0.05 72 –25
Acetic acid 5.30 44 –79
DMFA 0.04 79 –8
Pyridine 0.01 96 37
Quinoline 0.03 100 63
Ethyl acetate 0.51 61 –63
under the action of an electric field of the cation pass into solution and bind with solvent
molecules. Therefore, both the rate of formation of metalloporphyrin and activation param-
eters of the process depend on the extent of change of the N-H bond covalence and electron
density at atoms of nitrogen of the porphyrin coordination core under the influence of sub-
stituents in the molecule. Table 2 shows how great that influence is. The formation rates of
metal complexes, depending on the structure of porphyrin, change hundreds of times; and
activation energies, from 41.2 up to 115 kJ/mol.
Table 2 Rate constants and activation parameters of the complexation reaction of porphyrins with
copper(II) cation in ethanol at 298 K [1].
Porphyrin k, E, ∆S ≠,
l/(mol·s) kJ/mol J/(mol·K)
Chlorophyll 0.35 60.7 –50

Porphin 0.55 55.3 –67
Ethioporphyrin I 1.94 56.1 –50
Ethiochlorin II 2.08 47.3 –75
Tetraphenylporphin 2.05 58.6 –42
Protoporphyrin 2.43 78.7 –63
Mesoporphyrin 5.44 69.5 –4
Deuteroporphyrin 3.88 60.7 –37
Hematoporphyrin 2.26 75.7 –8
Phylloporphyrin 13.5 41.2 –75
Rhodoporphyrin 0.25 76.0 –11
Tetrabenzoporphin (a) 3.92 58.7 –84
Tetrazaporphin (a) 1000 115 118
(a)Solvent:
pyridine
It should also be noted that, as compared with complex-formation processes with open
(not macrocyclic) ligands, the formation rates of metalloporphyrins are extremely low. This
is due to the rigid planar structure of the ligand, to the shielding of the coordination core by
the adjacent atoms and π-electronic cloud. This spatial shielding of the reaction site is typ-
ical of the formation processes of rigid aromatic macrocycles and forms the basis of the
macrocyclic effect. It is the rigidity of the ligand, determined by the aromaticity of porphy-
rin, that does not enable it to take on the conformation suitable for rapid coordination. In
the course of interaction, it is not the ligand that has to adjust to the coordination sphere of
the salt, but, the other way round, the salt to the ligand, and the reagent is primarily the salt.
Upon introduction of an alkyl group, the porphyrin molecule is strongly polarized,
which contributes to the solvation of the molecule as a whole. As the result, the transition
state of meso-alkyl-substituted porphyrin becomes more probable [1]. The coordination rate
of phylloporphyrin 1 by Cu(II) cations in ethanol is three times as large as that of pyrropor-
phyrin 2. meso-Ethyl substitution of octaethylporphyrin (compound 3) increases the rate of
the reaction with zinc cation in acetonitrile 6.6-fold (Table 3). Introduction of a hexyl group
into hexamethyldiethylporphyrin 4 increases the rate of coordination of the molecule by
copper(II) and zinc(II) cations in propanol by approximately an order [2]. That is, with the
length of alkyl in meso-position (Alk = CH 3, C2H5, C6H13) increasing, the rate of the pro-
cess rises 3- to 10-fold.
Table 3 Kinetic parameters of the coordination reaction of octalkylporphyrins (1–6) with various
substituents in meso-position, copper and zinc cations in alcohols at 298 K [3].
Porphyrin meso- c°salt·103, k, E, ∆S ≠,

Substituent mol·l–1 l·mol–1 ·s–1 kJ/mol J/(mol·K)
1 CH3 0.92(Cu(OAc)2) 48.5 10.0 –18.7

2 H 2.61(Cu(OAc)2) 15.5 7.0 –32.3
5 C6H13 0.35(Cu(OAc)2) 26.1 44.2 –81.9
6 C6H5 0.37(Cu(OAc)2) 30.2 48.7 –64.9
3 (a) C2H5 1.15(Zn(OAc)2) 25.0 35.0 –10.4
4 C6H13 0.33(Zn(OAc)2) 23.4 42.1 94.6
5 C6H13 0.24(Zn(OAc)2) 29.0 34.4 –111.1
6 C6H5 0.20(Zn(OAc)2) 55.0 38.2 –94.7
(a) Data are given for acetonitrile.
A single meso-phenyl substitution (compound 6) with fixed position of the phenyl

group leads to an almost the same effect as hexyl substitution (compound 5). What is more,
in the case of Zn(II) the reactivity of the porphyrin molecule strongly changes (~ 2-fold) as
compared with the similar process involving Cu(II).
The works [4–7] studied the change of the complex-forming ability of 3,7,13,17-tetra-
methyl-2,8,12,18-tetrabutylporphyrin (7) during its substitution at two meso-positions: 1)
by phenyl fragments (compound 8); 2) by phenyl fragments containing electron-donor
(CH3O) and electron-acceptor (NO2) substituents (compounds 9–12); by alkyl groups of
various lengths (compounds 13, 14).
The complexation kinetics of disubstituted octalkylporphyrins with copper(II) and
zinc(II) cations was studied in ethanol, acetic acid, acetonitrile and pyridine. The complex-
ation reaction of porphyrins (7–14) with copper and zinc cations in these solvents is de-
scribed by the first-order kinetic equation with respect to porphyrin, usual for monomeric
porphyrins. As the order of the reaction with respect to copper acetate in acetic acid [8],
ethanol and dimethylformamide [9] is equal to 0.5, the total order of the reaction with re-
spect to the reagents is 1.5. Tables 4–9 present the 1.5-order rate constants calculated using
equation (1):
(1) k
1.5 = kef / cCu (OAc)
2
(1)
R4 R5
R3
1 : R1 =CH2CH2COOH, R2= R3= R5 = R7 = R9 = Me, R4= R6=Et,
R6
R8=H,
NH N
2 : R1 =CH2CH2COOH, R2= R3= R5 = R7 = Me, R4= R6=Et,
R8=R9 =H
3 : R1 = R2= R3= R4 = R5= R6= R7 = R8= R9=Et
N HN
4 : R1 = R2= R3= R6 = R 7 =R 8 =Me, R4 = R5=Et, R9 =C 6H13
R2
R7
5 : R1 = R4 = R5 =R 8 = Me, R2= R3 =R 6=R 7 =Et, R9 =C6H13
R1 R9 R8 6 : R1 = R4 = R5 = R8 = Me, R2= R3 =R6 =R7=Et, R9 =Ph
1-6
R R 7 : R = Bu, R' = H
8 : R = Bu, R' = Ph
H3C
CH3
NH N 9 : R = Bu, R' = PhOMe-p
R' R' 10 : R = Bu, R' = PhOMe-m
N HN 11 : R = Bu, R' = PhOMe-o

H3C
CH3 12 : R = Bu, R' = PhNO2-p
R R 13 : R = Me, R' = Et
7-14 14 : R = Me, R' = C3H7
In the complexation reaction with copper(II) acetate the substituents of benzene

fragments little affect the reactivity of porphyrins 9–12. The presence of methyl groups in
β-positions of the macrocycle excludes the possibility of rotation of benzene fragments,
which, being located in the plane perpendicular to the plane of porphyrin, do not enter into
conjugation with the macrocyclic π-electronic system. Nevertheless, there is a tendency to
an increase of the complexation rate under the influence of electron-donor substituents of
benzene residues. Apparently, the substituents, by changing the electon density in a benzene
fragment, affect the electronegativity of carbon atom of porphyrin macrocycle’s methine
bridge. Further on, the effect of the substituent is passed by means of σ-bonds C–C and
C–N to the reaction centre. Introduction of electron-donor substituents (para-CH3O 9,
meta-CH3O 10) into benzene fragments of 5,15-diphenylporphyrin 8 insignificantly accel-
erated the reaction (Tables 4, 5).
Table 4 Kinetic parameters of the complexation reaction of 5,15-diarylporphyrins (8–12) with
copper(II) cation in acetic acid at 298 K: cCu(OAc)2 = 3.047·10–3 mol/l; cºH2P = 4.021·10–5 mol/l.
No meso-Substituent k1.5 ·102, E, ∆S ≠,

l0.5 ·mol–0.5 ·s–1 kJ/mol J/(mol·K)
8 C6H5 5.1±0.1 97±2 70±6

9 C6H5(n-OCH3) 5.2 ±0.2 89±1 43±4
10 C6H5(m-OCH3) 5.3±0.15 87±2 37±7
11 C6H5(o-OCH3) 8.4±0.1 89±1 43.5±4
12 C6H5(n-NO2) 11.0±0.2 77±0.5 3.5±3

copper(II) cation in acetonitrile at 298 K: cCu(OAc)2 = 3.760·10–3 mol/l; cºH2P = 3.521·10–5 mol/l.

7 H 17.1±0.05 46±1 –113±3

8 C6H5 9.2 ±0.05 77±2 87±6
9 C6H5(n-OCH3) 10.3±0.05 78±1 –10±3
10 C6H5(m-OCH3) 10.1±0.05 71±1.5 –34±5
11 C6H5(o-OCH3) 5.1±0.05 85±0.5 7±3
12 C6H5(n-NO2) 5.8±0.05 76±2 –21±7

copper(II) cation in pyridine at 298 K: cCu(OAc)2 = 4.956·10–3 mol/l; cºH2P = 2.04·10–5 mol/l.
7 H 0.16 78±1.0 3±3

8 C6H5 0.08 98±4 70±10
9 C6H5(n-OCH3) 0.09 98±1.5 70±5
10 C6H5(m-OCH3) 0.08 94.5±2 58±8
11 C6H5(o-OCH3) 0.09 94±2 57±6
12 C6H5(n-NO2) 0.11 86.5±1.0 32±4
Table 7 Kinetic parameters of the complexation reaction of 5,15-diarylporphyrins (7, 8, 10, 11)
with copper(II) cation in ethanol at 298 K: cCu(OAc)2 = 1.211·10–3 mol/l; cºH2P = 4.520·10–5 mol/l.
7 H 219 ± 2 37±3 –59±8

8 C6H5 117±1.2 44±3 –103±8
10 C6H5(m-OCH3) 130±1.4 45±1 –101±3
11 C6H5(o-OCH3) 128±2 43±4 –108±4
The electron-acceptor substituent (para-NO2, 12) slightly slows down the reaction.
Against the background of pronounced effects of benzene fragments’ substituents in por-
phyrins 7--12, the role of solvation effects increases. Thus, in pyridine (Table 6), diphe-
nylporphyrin 8 and its ortho-methoxy derivative 11 have almost the same reactivity
(α,α-atropisomer was studied, in which both methoxy groups are on one side of the por-
phyrine macrocycle), whereas in acetonitrile (Table 5) the derivative 11 reacts with cop-
per(II) cation two times as slow as the initial compound 8.
In passing from meso-unsubstituted porphyrin 7 to dialkylporphyrins 13, 14, the reac-
tivity of the macrocycle increases (Tables 8, 9). The rate constant in ethanol changes from
219·10 –2 l/mol·s (compound 7) to 271·10 –2 l/mol·s (compound 13). Probably, alkyl sub-
stituents, possessing a positive induction effect, increase the electron density at tertiary at-
oms of nitrogen and facilitate the interaction of metal cation with porphyrin in transition
state. A decrease of the activation energy (~ by 7.0 kJ/(mol·K)) in meso-alkyl substitution
of porphyrin in ethanol is due to an increase of solvation of the salt--porphyrin system’s
transition state during the emergence of alkyl substituents in the molecule. It is known that
introduction of two ethyl groups in meso-position of octamethylporphyrin causes an in-
crease of solubility of the molecule in benzene by two orders of magnitude. The effect of
substituents is more pronounced in acetic acid (Tables 4, 9), which specifically solvates the
reaction centre of porphyrin and decreases its reactivity in the interaction with copper(II)
cation. The strength of the hydrogen bonds between nitrogen atoms of porphyrin and hy-
drogen atoms of acid is reduced at the introduction of acceptor substituents into 8. This
causes an increase of the reaction rate (Table 4). At 298 K, the rate constant increases from
5.1·10 –2 (8) to 11·10 –2 l/(mol·s) (para-nitroderivative 12). As compared with diphenylpor-
phyrin, the activation energy of the nitroderivative decreases by ~20 kJ/mol; and activation
entropy, by 67 J/(mol·K). Introduction of electron-donor alkyl groups leads to a decrease
of reactivity of the macrocycle (Table 9).
Table 8 Kinetic parameters of the complexation reaction of porphyrins (7,13, 14) with copper(II)
and zinc(II) cations in ethanol at 298 K.
No meso- c°salt ·105, mol/l k1.5 ·102, E, ∆S ≠,

Substituent l0.5 ·mol–0.5 ·s–1 kJ/mol J/(mol·K)
7 H 1.21 (Cu(OAc)2) 219±6 40±1.5 –59±9

13 C2H5 2.80 (Cu(OAc)2) 271±6 37±3 –121±5
14 C3H7 5.81 (Zn(OAc)2) 208±2 48±3 –89±12
The rate constant in the case of zinc complexes of porphyrins is of the second order (kv, l/mol · s).
Table 9 Kinetic parameters of the complexation reaction of 5,15-diarylporphyrins (7, 13, 14) with
copper(II) cation in acetic acid at 298 K: cCu(OAc)2 = 2.410·10–3 mol/l; cºH2P = 6.721·10–5 mol/l.

0.5
l ·mol–0.5 ·s–1 kJ/mol J/(mol·K)
7 H 3.1±0.15 60±2 –54±6

13 C2H5 0.7±0.05 52±4 –82±13
14 C6H13 0.4±0.05 55±2 –73±7
The reactivity of porphyrins in complexation with metal cations strongly depends on

the nature of a solvent. Low rates of formation of metal complexes in pyridine (Table 6) are
determined by a high strength of the solvate coordination sphere of [Cu(OAc)2(Py)4]. The
maximal values of k1.5 are observed in running the reaction in ethanol (Tables 7, 8), which
forms a solvated complex [Cu(OAc)2(C2H5OH)4] unstable as compared with pyridine.
Owing to the increase polarity of ethanol, the polar transition state of the salt–porphyrin
system is subjected to efficient additional solvation in alcohol. For this reason, in ethyl alco-
hol as compared with pyridine and acetic acid, the values of energy and entropy of the co-
ordination reaction have minimal values. A decrease of reactivity of porphyrins in acetic
acid shows that the prevalent effect on the rate of the process is rended by blocking the por-
phyrin’s reaction centre due to the occurrence of hydrogen bonds. A significant increase of
energy and entropy of activation in acetic acid as compared with ethanol has been noted
(Tables 7–9).
Despite the low donor strength of acetonitrile [10--12] forming labile solvated com-
plexes of transition metals [13--16], the reaction rate of copper(II) cation with porphyrins
is lower than in ethanol (Tables 7, 8). Probably, this is due to the decreased basicity of ac-
etonitrile, incapable of efficient solvation of protons leaving the porphyrin’s reaction centre
in transition state.
Of special interest are porphyrins containing complex-forming cavities of various na-
ture (e.g., calix[4]arenes). The presence of additional complex-forming fragments in such
molecules enables their use in studies of complexation processes occurring without the di-
rect participation of tetrapyrrole macrocycles [17, 18].
The works [19, 20] investigated porphyrin-calix[4]arenes 15, 16 with “activated”
amide substituents, which are capable of interacting with anions by forming hydrogen
bonds by NH groups of bridges. Analysis of the binding constants of porphyrin-calixarene
conjugates 17--19 with ions I – , Br – , Cl – and NO3– (Table 10) shows that, irrespective of
the conformation of the calix[4]arene moiety of the molecule, compounds 17--19 efficiently
bind the spherical anions of small size. The bridge groups linking the calix[4]arene and
O
NH N
NH
NH
N NH
R
R anion
R
R
X
15, 16
O
NH N
15 : X = H , 16 : X=
NH
NH
N NH
P
P P
NH
P
NH P P
O NH
NH NH
O O
NH O NH
NH O
NH O
NH NH
OPrOPr NH
OPr OPr
OPr OPr OPr OPr
OPr OPr OPr OPr
17 18 19
N
P= NH
HN
N
porphyrin fragments possess a sufficient flexibility for placing ions I – and Br – , irrespective
of their location [cis- or trans-], on the upper rim of calix[4]arene. What is more, the pro-
poxy groups located between porphyrin fragments of 1,3-alternate do not prevent the com-
polexation with anion. At the same time, significant differences in the binding constants for
larger anions (I – and NO3– ) point to certain spatial restrictions for complexation. With the
diameter of anion increasing, the binding constant of anions decreases [for porphy-
rin-calix[4]arene 19 KCl (6.9·105 l/mol) > KBr (6.9·104 l/mol) > KI (2.4·103 l/mol)]. Here-
with, compound 19 exhibits a better complexing ability with respect to Cl – as compared
with the system having a similar structure of the calix[4]arene platform, but instead of tet-
rapyrrole fragments containing phenyl groups (KCl 4.6·103 l/mol) [20].
Table 10 Constants of binding Ka [l/mol] by porphyrin-calix[4]arenes (17–19) of various-nature

anions in dichloromethane at 24ºC (creceptors ~ 1.5·10–6 mol/l).
Anion 17 18 19
Cl– 6.3×10 3 5.8 × 10 5 6.9×10 5

Br– 1.2×10 3 7.8×10 4 6.9×10 4
I– 240 8×10 3 2.4×10 3
NO–3 820 3×10 4 1.3×10 4
2 Complexation of Porphyrins with Organic Molecules:

The Thermodynamic Aspect
Axial coordination (extracoordination) of ligands is inherent in many complexes with a pla-
nar structure of the coordination node ML4, but on metalloporphyrins it is especially pro-
nounced and peculiar [1, 21–23]. Formation of porphyrin--ligand extra complexes (their
structure, composition, stability) depends on the nature of metal. In the case of Zn(II) prone
to sp3 or d 2sp2 hybridization, but in a planar porphyrin molecule forced to take on a dsp2
configuration, electron-deficient 4pz and 4dz2 orbitals are formed, which enter into a donor-
acceptor interaction with electron-donor ligands.
Zn-porphyrins are readily bind nitrogen-containing ligands and much weaker coordi-
nate oxygen- and sulfur-containing ligands. But although oxygen coordination on zinc is
less weak than nitrogen coordination, it also can be used as a nonselective anchor point in
multipoint recognition [24, 25]. According to [26, 27], monomeric Zn-porphyrins are ca-
pable of binding only one axial ligand, thus forming a five-coordinated complex. At the
same time, the authors of [25, 28, 29] note that in the case of weaker bases Zn-porphyrins
can add two extra ligands, but with different strength.
In coordination of oxygen-, sulfur- and nitrogen-containing ligands on monomeric
Zn-porphyrin, the binding constant of Zn-porphyrin--ligand complexes (Ka) (association
constant, extracoordination constant, stability constant -- there are different terms in the lit-
erature) very strongly depends on the basicity of this ligand. In a general case, the larger
the basicity of a ligand, the stronger it is coordinated on metalloporphyrin. In the sequence
ethanol < pyrrole < DMSO < pyridine < imidazole < piperidine
the binding constant of Zn-tetraphenylporphin (Zn-TPP) in toluene increases by four orders
[27]. This dependence is disturbed, e.g., in the case when the free electron pair of the ligand,
taking part in coordination, is delocalized on adjacent substituents [30]. As seen from Table
11, the effect of the solvent and of the alkyl- and arylporphyrin structure on the process of
axial coordination is much smaller than the effect of the nature of the axial ligand itself. The
extracoordination process on metalloporphyrins is usually studied in a medium of “inert”
solvents (benzene, toluene, dichloromethane). Works by Viugin et al. [31, 32] investigated
extracoordination of molecules of various solvents for a large number of metalloporphyrins
by the calorimetric method. Coordination enthalpies of the above “inert” solvents on
Zn-porphyrins are close to zero. According to the data of Table 11, the binding constants
of Zn-porphyrin--ligand complexes in passing from one “inert” solvent to another change
no more than fourfold. The largest values of Ka are observed in aromatic solvents of the
type of benzene and toluene.
The effect of alkyl and aryl substitution on the binding constants in porphyrins 20--31
is levelled out even more (no more than twofold), as the electronic influence of phenyl frag-
ments and alkyl groups on the state of the central zinc ion is insignificant. At the same time,
Table 11 Ka of some alkyl- and aryl-substituted monomeric Zn-porphyrins in monodentate ligands

at 298 K.
Zn-porphyrin Monodentate ligand Solvent Ka, mol–1 ·l Reference
20 Piperidine Toluene 809000 1

Imidazole Toluene 24000 1
Quinoline Toluene 16900 1
Pyridine Toluene 5800 1
Pyridine Toluene 4790 33
Pyrrole Toluene 200 1
Methanol Toluene 4.6 1
Pyridine Benzene 3750 34
Pyridine CHCl3 2525 35
4-Me-Pyridine CHCl3 1600 37
Pyrazole CHCl3 1500 37
Methylimidazole o-Xylene 110000 38
Imidazole o-Xylene 15000 38
Pyridine o-Xylene 1800 38
DMFA o-Xylene 560 38
21 Pyrazole CH2Cl2 2600 39(a)
22 Imidazole Benzene 51920 1
23 Pyridine CH2Cl2 2890 40(b)
24 Pyrazole CH2Cl2 800 39
25 Pyridine Benzene 10300 35
Pyridine Toluene-methanol (2:1) 1380 41
26 Pyridine Toluene 5000 42
4-Me-Pyridine Toluene 11000 42
4-But-Pyridine Toluene 14000 42
27 Pyridine CH2Cl2 2800 43(c)
28 Pyridine CH2Cl2 5200 43
(a) Data
are given at 295 K; (b) temperature not indicated; (c) data are given at 303 K.
in the sequence of metalloporphyrins 32--49 with different deformation degrees of the tet-
rapyrrole macrocycle the binding constant of monodentate basic nitrogens changes, de-
pending on the nature of this macrocycle, by several orders (Table 12).
The main factors determining the axial coordination process in these porphyrins are:
a) schielding of the reaction centre of the tetrapyrrole macrocycle (on one or both sides); b)
change of electron density on nitrogen atoms and of charge on Zn(II) cation as the result of
Table 12 Ka of sterically hindered monomeric and dimeric Zn-porphyrins and monodentate

ligands at 298 K.
Zn-porphyrin Monodentate ligand Solvent Ka, mol–1 ·l Reference

Isoquinoline Toluene 4.6×105 46
Isoquinoline Toluene 9.6 ×105 46
36 2-Methyl-imidazole o-Xylene 460 38
Imidazole o-Xylene 930 38
Pyridine o-Xylene 240 38
DMFA o-Xylene 42 38
Imidazole CH2Cl2 1259000 48
2-Methyl-imidazole CH2Cl2 195000000 48
2-Phenyl-imidazole CH2Cl2 2550000 48
42 Pyridine CHCl3 14330 49
43 Pyridine CHCl3 91855 49
4-Methyl-pyridine Toluene 62000 42
4But-Pyridine Toluene 730000 42
53 Pyridine Toluene-methanol (2:1) 2700 41
54 Pyridine Toluene-methanol (2:1) 3200 41
R8 R1
R
R7 R2
N N
Zn
N N
N N R6 R3
Zn
N N
R R R5 R4
22, 23
22 : R1=R2=R3=R4=R5=R6=R7=R8=H
R 23 : R1=R3=R5=R7=CH3, R2=R6=C2H5,
20, 21 R4=R8=(CH2)2COOCH3
20 : R = H, 21 : R = CO2CH3-m
R1 R1 R
R
R2 R2 N N
N N
Zn
Zn
N N
N N
R1 R1 CH3OOCCH2CH2 CH2CH2COOCH3
24-28 29-31
24 : R1=C2H5, R2= COOCH3-m 29 : R= CH(OH)CH3

25 : R1 = C4H9, R2 = OCH3-o 30 : R = CH2CH3
O
26 : R1 = , R2= C CH-m 31 : R = CH=CH2
O
27 : R = C2H5, R2= C CH-m

28 : R = (CH2)2COOCH3 , R2= C CH-m
deformation of the macrocycle; c) correspondence of the size of the cavity formed by ster-
ically hindered porphyrin macrocycle to the size of extra ligand; d) emergence of additional
interactions of the type of π−π, CH−π and others between metalloporphyrin (host molecule)
and ligand (guest molecule).
Coordination of axial ligands on picket-fence, strapped and dimeric porphyrins has its
own features. The term “picket-fence” is used for porphyrins with peripheral bulky substit-
uents. In most cases, these are derivatives of tetraphenylporphin containing an ortho-sub-
stituent in each benzene nucleus. Depending on the orientation of substituents relative to
the plane of the tetrapyrrole macrocycle (α, on one side; β, on the other side), four atropi-
somers are distinguished: α,α,α,α- (α4), α,β,α,α-, α,α,β,β- (cis-α2) and α,β,α,β
(trans-α2).
Works by Imai and Kyuno investigated the axial coordination on picket-fence metallo-
porphyrins (32, 33) of pyridine, piperidine and quinoline in toluene and other noncoordi-
O t-Bu t-Bu O
NH O t-Bu
t-Bu NH
NH
O N
O t-Bu NH N
N Zn
N
N N
NH N Zn O t-Bu NH
H
N
NH O
R
NH
R O
33
32
Et O R1
Et R1 HO HO
O
N N OH
Zn
N N OH
O OH
Et
OH Et O OH
O
O
O O O R1
34
R1 O O
OH
X
O
Bu Bu
O
N N O RO RO O
Zn Et Et
N N O O
N N
Zn
Bu N N
Bu
Et Et
35, 36
37,38
35 : X =
OCH3 37 : R = COCF3 38 : R = H
36 : X = CH2 CH2
OCH3
nating solvents [46, 49]. The results obtained showed trans-α2 atropisomers to have a high-
er affinity to pyridine and quinoline than α2 atropisomers.
As in the case of trans-α2 atropisomers both sides of the reaction centre of porphyrin
are equally accessible for coordination, an increase of affinity to pyridine and quinoline is
O O
N N
O
O Et O O Et
O O O
O N
N N N
O Zn Zn
N N N N
O
Et Et
39 40
C5H11
C5H11 C5H11
C5H11 C5H11
C5H11 C5H11
C5H11
O O
O O
O O
O O O
O R
O O R
R O O
O O
R N
R NR N Zn N
N Zn N N
N
43, 44
41, 42
O
41 : n = 1, 42 : n = 5
R= NH (CH2) n 43 : n = 1, 44 : n = 5
O
Et Et 45 : X= O
O
O
N N
Zn O
N N
46 : X= O
X
Et Et Et O
Et
O
X
N N
47 : X= O
Zn
N N
O
O O
Et Et O
O N N
48 : X=
45-48 O
O O O
determined by the emergence of weak CH-π interactions in the system considered, which
is the cause of higher binding constants. In α4 atropisomers 32, both sides of the reaction
centre of porphyrin are inadequate. Four ortho-substituents shield the reaction centre almost
completely, and the ligands are coordinated only by the “open” side of the porphyrin mac-
rocycle, without forming any additional bonds. .
R R .
N N
N N Zn
Zn N N
N N
RO
R OR
( )n R RO
R R ( )n OR
N N N N
Zn Zn
N N N N
R R
49 - 51 52, 53
O
49 : R= , n=2 52 : R = H, 53 : R = COCF3
O
50 : R = (CH2)2COOCH3 , n = 2
51 : R = (CH2)2COOCH3 , n = 4
Strapped porphyrins include compounds, in which two diametrically opposite periph-

eral positions of the macrocycle are linked by bridges of various nature [51, 52]. The aim
of the synthesis of belted porphyrins is to develop a diversity of structures modelling en-
zyme systems. The binding constant of strapped porphyrins is first of all determined by the
shielding of the porphyrin site (on one or two sides) and by the correspondence of the size
of the intramolecular cavity of metalloporphyrine to the size of coordinated ligand. If the
bulky substituents do not shield the reaction centre (as in the case of 34), then Ka of such a
porphyrin does not in practice differ from the respective value of a sterically not shielded
porphyrin (Table 12) [53]. If the bulky substituent shields the reaction centre on one side,
the binding constant depends primarily on the correspondence of the size of the intramo-
lecular cavity of metalloporphyrin to the size of coordinated ligand. If there is no such a
correspondence, the ligand is coordinated by only the “open” side of the strapped macro-
cycle. Herewitth, the binding constants of metalloporphyrins with various ligands can be
lower than those of porphyrins without steric shielding, which is observed, e.g., in the case
of porphyrins 34--40 (Table 12). The authors of [54--58] give no unambiguous explanation
to these data.
All investigated metalloporphyrins (35--40) with lower binding constants, occurring
in the linking bridge, have aromatoc fragments, which, in our opinion, by entering into
π – π-interaction with porphyrin macrocycles, lead to a decrease of charge at the Zn cation
of the reaction centre. This is one of the main causes of lower Ka in porphyrins (35--40) as
compared with respective metalloporphyrins without “shielding” substituents.
If the size of the ligand corresponds to that of the intramolecular cavity of sterically
shielded porphyrin, i.e., the ligand, by being coordinated at the Zn cation, can incorporate
into the intramolecular cavity, in this case the binding constants increase significantly.
Complexation studies of Zn-porphyrin-calix[4]arene conjugates 41--44 with organic
molecules of various size and nature by the method of spectrophotometric titration showed
complex 41 with the least distance between the porphyrin and calix[4]arene fragments to
possess the greatest capability of axially binding N-methylimidazole and pyridine (Table
13) [34]. Complex 41 has four binding links, which, due to the strengthening of the mole-
cule structure impart the hydrophobic cavity of the calix[4]arene fragment with a fixed
shape, and organic molecules are bound depending on their size.
Large-size molecules (nicotine, nicotinamides and 4-phenylpyridine) are bound on the
ooo
Table 13 Ka (·103 l/mol) of nitrogen-containing ligands by porphyrin-calix[4]arenes (41–44) and

Zn-tetraphenylporphyrin (Zn-TPP) in chloroform (cZn-porphyrins ~ 5·10–6 mol/l).
Ligand 41 42 43 44 Zn-TPP
147 15 18 6.8 0.9
233 46 80 20 1.6
N
0.4 10 52 13 1.4
N
1077 140 290 79 1.5
N
O
0.7 0.3 62 19 0.3
NH 2
O
N H 2 0.4 13 15 1.6 0.3
0.2 16 20 41 1.4
N
outer side of the receptor with the binding constant equal to or even smaller than in Zn-TPP.
Binding of smaller-size molecules (N-methylimidazole, pyridine, picoline) strongly de-
pends on the shielding effect of the calix[4]arene cover and on the electron density at the
donor atom of nitrogen. The presence of long and flexible binding links in receptor 42 leads
to the formation of a large-size cavity, and most molecules of the host are bound on the inner
side of the receptor (excluding nicotine). Simultaneously, the shielding effect of the
calix[4]arene “cover” is attenuated. The decrease of the number of binding links in complex
43 to two leads to an increase of the molecule’s flexibility and, as a consequence, to an in-
crease of the distance between the porphyrin and calix[4]arene fragments. This enables the
molecule of 4-phenylpyridine to penetrate into the inner cavity of the receptor. Porphy-
rin-calix[4]arene 43 binds nicotinamide 200 times stronger as compared with Zn-TPP and
is the best receptor in the sequence of complexes 41--44 for binding nicotinamide, 4-phe-
nylpyridine and isonicotinamide.
An increase of the length of the binding links in compound 44 leads to a decrease of
the shielding effect of the calixarene cover as compared with complex 43. In compound 44,
the flexibility of the binding links enables the molecule to “fold” the binding links with a
shift of the porphyrin and calix[4]arene fragments relative to one another. This decrease of-
the inner cavity leads to a decrease of the complexing ability of receptor 44 with small mol-
ecules as compared with complexes 41--43. An exception is nicotine, whose efficient
binding can, probably, be explained by the additional formation of hydrogen bonds with
amide groups of the binding links.
In cyclophane dimers 45--53, each monomeric porphyrin fragment is also shielded on
one side owing to the other porphyrin fragment, as the result of which Ka of the ligands by
dimers 45, 47, 48 and 51 is lower than of their monomeric analogues. In coordination by
47, 49 and 50 of monodentate ligands to form complexes of composition 1:1, the binding
constants are observed to increase as compared with monomeric analogues (in the case of
the coordination of pyridine, Ka increases two- to threefold). Some authors associate this
with electronic effects [34, 54].
In their opinion, pyridine, along with the formation of a σ-bond Zn--N, interacts with
metal by the π-type with the transfer of electron density from dxz, dyz orbitals of Zn(II) to
vacant π*-molecular orbitals of pyridine. In passing from monomeric porphyrin to dimeric
one, the probability of this interaction increases due to the enhancement of the electron-do-
nor properties of the deformed porphyrin macrocycle, which ultimately is accompanied
with an increase of Ka of the pyridinate axial complex of cyclophane dimer.
According to the opinion of other authors [59], an increase of the binding constant of
one ligand by dimer as compared with monomer, as in the cases of spatially hindered por-
phyrins, is determined by the correspondence between the size of the inner cavity of dimeric
porphyrin and that of the ligand. The better this correspondence, the greater the binding con-
stant is. In the sequence of para-substituted pyridins, the best correspondence has para-tert-
butylpyridine (Ka increases 50-fold). The second extra ligand is attached only from the out-
er side and its binding constant is much weaker.
In coordination by cyclophane dimeric porphyrins of bidentate ligands (with two ni-
trogen atoms), if the geometric requirements of the interplane space correspond to the size
of the ligand, a dimeric porphyrin--bidentate ligand complex of composition 1:1 is formed,
in which the bidentate ligand is located in the inner cavity of dimeric porphyrin and had two
binding points.
Conditions of forming various-composition complexes, their stability constants and
other regularities are described in works by Sanders by example of such ligands as pyrazine,
4,4′-dipyridyl and DABCO (1,4-diazo-bicyclo[2,2,2]octane) [53, 56, 60]. Zn-porphyrin
with monodentate ligand forms a complex of composition 1:1 with the binding constant Ka
(Scheme 1). In the case of bidentate ligand, a 1:1 complex is formed with the binding con-
stant Ka,1 ≈ 2Ka (Scheme 2). Addition of the second ligand to form a 2:1 complex occurs
Scheme
Scheme1 1
L
N Ka
N N
Zn + L Zn
N
N N N N
Scheme 2
Scheme 2
N N
Zn
L Ka,1 N N
N N
Zn
N N + L
L
L
Scheme 3
Scheme 3
N N
N N Zn
Zn N N
N N
N N Ka,2 L
N
Zn L
N
+
L
L
N N
Zn
N N
with Ka,2, which is related to Ka,1 as 4Ka,2 = aKa,1, where a is the “interaction” parameter,
being a measure of interactin between two reaction centres (Scheme 3).
In the case of bifunctional ligand and dimeric porphyrin, the situation becomes more
complicated. Complexes can be formed both on the inner (Ka (in)) and outer ((Ka (out)) side
of the dimer. The strongest complexes are formed in the case, when bifunctional ligand is
coordinated by the inner cavity of dimer. Formation energy of this complex in the general
Table 14 Ka of Zn-porphyrins with bidentate ligands.
Zn-porphyrin Ligand Solvent Ka, mol–1 ·l
Zn-monomeric porphyrins without steric shielding

(binding constants of the complexes of composition 1:1)
20 Ethylene-diamine CHCl3 18270
23 DABCO CH2Cl2 240000
24 DABCO CH2Cl2 16000
BiPy CH2Cl2 5400
25 DABCO Toluene-methanol 5400
(2:1)
Pyrazine Toluene-methanol 500
(2:1)
26 Ethylene-diamine CHCl3 18970
Table 14 (continued)
Ka of Zn-porphyrins with bidentate ligands.
Zn-porphyrin Ligand Solvent Ka, mol–1 ·l
27 BiPy CH2Cl2 4700

28 BiPy CH2Cl2 8400
Sterically shielded Zn-monomeric porphyrins
(binding constants of the complexes of composition 1:1)
37 BiPy CH2Cl2 970
38 BiPy CH2Cl2 2.6×103
39 DABCO CH2Cl2 4.9×104
40 DABCO CH2Cl2 2.3×104
Zn-dimeric porphyrins
(binding constants of the inner complexes of composition 1:1)
45 DABCO CH2Cl2 7.4×107
46 DABCO CH2Cl2 1×107
47 DABCO CH2Cl2 1.3×106
BiPy CH2Cl2 3800
Pyrazine CH2Cl2 600
48 DABCO CH2Cl2 2.4×105
BiPy CH2Cl2 1800
Pyrazine CH2Cl2 690
52 DABCO CH2Cl2 2.0×105
BiPy CH2Cl2 1.7×104
53 DABCO CH2Cl2 8.0×107
BiPy CH2Cl2 2.5×106
case is not the doubled energy of forming a complex with monomer, owing to a whole range
of factors: the chelate effect, interaction between reaction centres of bidentate ligand, con-
formation changes, as well as new interactions, which can emerge in the system. These
components of the total binding energy of the dimeric porphyrin--bidentate ligand complex
depend on the nature and conformations of linking bridges in dimeric porphyrin and can be
rather significant. Depending on the nature of a linking bridge, the constant of binding of
dimeric porphyrin with bidentate ligand by the two-site scheme can be higher, as compared
with the monomeric analogues, from 2 up to 5,000 times (Table 14).
Coordination of bidentate ligands by linear dimeric porphyrins depends on the flexi-
bility of the linking bridge. In the case of dimer 54 with the rigid aromatic bridge, porphyrin
fragments are fixed in space at a large distance one from another, and complexation with
DABCO in CH2Cl2 proceeds by the type of 1:2 interaction with Ka ≈ 105 l/mol (each
metalloporphyrin fragment of the dimer binds one molecule of bidentate ligand) [54, 61].
In dimer 55 with a more flexible linking bridge, complexation with DABCO proceeds to
form an “internal” complex of composition 1:1 with Ka ≈ 107 l/mol (two metalloporphyrin
fragments of the dimer bind one molecule of bidentate ligand). “Internal” complexes with
DABCO are also formed by dimer 56 (Ka = 6·105 l/mol) [59, 62]. The authors of [55] note
a high affinity to diamine in “flexible” dimer 57 with a crown-ether linking bridge; in the
presence of barium perchlorate, the affinity increases.
The recognizing ability of calixarene-porphyrin conjugates 58, 59 with respect to
DABCO was studied in [19, 56]. It is noted there that compound 59 forms an internal 1:1
OH
N N
N Zn N N Zn N
N N
54
O O
O O
N N
N Zn N N Zn N
N N
55
Ph
N N
Zn Ph
N N
N N
Zn
N N
O Ph
O
O
O
N N
O O
Zn
N N
Ph
N N
Zn
N N Ph
56
57
Ph
complex, whereas 58 coordinates DABCO only on the outer side. A significant difference
of the complexing ability of the calix[4]arene derivative 58 from thiacalix[4]arene deriva-
tive 59 is explained by the authors by differences in the size of respective calixarene cavi-
ties. A larger cavity in conjugate 59 corresponds better to the geometry of bidentate ligand
containing two diametrically located nitrogen atoms, which leads to the formation
O
t N
Bu NH N
O Zn
N N
XX
t
Bu O R
t
Bu o R
X
O
X N N
t O NH Zn
Bu N N
58-59
58 : X=CH2, 59 : X=S
R R
N N
Zn
.
N N
R R
SiMe3
R R n
N N
Zn
N N
Me3Si
60-62 R
R
60 : R = CH2CH3, n=3 63
61 : R = (CH2)2COOCH3, n=3
62 : R = (CH2)2COOCH3, n=4 n=4, R=CH2CH2COOCH3
of an intramolecular 1:1 complex with the binding constant Ka = (1.0±0.1) ·107 l/mol in
chloroform at 294 K. Conjugate 58 containing the usual calix[4]arene fragment forms a
complex by way of independent coordination of DABCO molecules by two tetrapyrrole
macrocycles (each metalloporphyrin fragment coordinates a ligand molecule) to form a 2:1
complex.
A detailed investigation of binding oligopyridyl ligands by di-, tri- and tetrameric por-
phyrins was carried out in [59--62]. A good geometrical correspondence of the size of por-
phyrin cavity and the size of ligand is observed in dimeric porphyrin 50 with BiPy (6 ·106
l/mol), trimeric porphyrins 60, 61 with Py3P (9 ·109 and 4·1010 l/mol, respectively), tet-
rameric porphyrin 62 with H2Py4P (2 ·1010 l/mol), which is slightly larger than in linear
tetramer 63 with this ligand (7·109 l/mol) [43]. Dimeric, trimeric and tetrameric porphyrins
synthesized according to an improved method have even more capacious intromolecular
cavities than cyclophane dimers considered above. The distance between porphyrin frag-
ments in dimer 51 is approximately 1.5 nm. This enables the zinc complex to be the host
R R
N N
Zn
N N
R R
n
64
n=3, R= CH2CH2COOCH3
N N N N
Py2C2
BiPy N
N
N N
N
N
N N
N
Py3P N Py3C12H3
NH N
N N
N HN
H2 Py4P
for a guest of respective geometry -- bis(4-pyridyl)ethine (Py2C2) (7·106 l/mol). Trimer 64

has a good geometric correspondence with Py3C12H3 (2·108 l/mol) [59].
Of other metalloporphyrins, Ni- and Ru-porphyrins are used the most widely as por-
phyrin receptors to nitrogen-containing ligands. The binding strength of basic nitrogens by
the central metal atom decreases in the sequence Ru >> Zn >> Ni. The most strong com-
plexes are formed by Ru-porphyrins [63--66]. In the case of Zn-porphyrins, molecular com-
H2P H2P
H2P N Ru N H2P
H2P
H2P
65
+
N
N N
Zn
N N N N
+ +
N M N
R
N N
67-69
67 : R=H, 68 : R=OH, 69 : R=OCH3

+
N
66 : M=H2, Zn, Co
plexes are formed based on two or three (maximum four) porphyrins; based on
Ru-porphyrin complexes, compounds based on five, seven and nine porphyrins were pro-
duced. An example of such a compound is Ru-complex 65 [64].
In processes of molecular recognition of amino acids, sugars, quinones and purine
bases by porphyrins, of great significance are hydrogen bonds, van-der-Waals and disper-
sion interactions. Recognition of amino acids in aqueous media was studied in [67, 68] by
the example of tetrakis-(4-methylpyridyl)-porphyrin 66 and its zinc and cobalt complexes.
In the case of metalloporphyrins, the main driving force of amino acid recognition is
extracoordination of the amino group of amino acid to metalloporphyrin. Besides this, a
weighty contribution is introduced by the interaction between the molecule of porphyrin
and amino acid (ligand--ligand interaction) and dispersion interactions between carbonyl
anion of the guest and pyridinium cation of the host. The value of dispersion interactions
depends on pH [68]. Ligand--ligand interaction takes place in the case of amino acids with
aromatic substituents phenylalanine and triptophane. This π--π interaction between the por-
phyrin molecule and aromatic moiety of amino acid stabilizes the complex [69], thus sig-
nificantly increasing the binding constant (from 60 l/mol in glycine up to 1200 l/mol in
triptophane).
According to [70--74], in organic solvents recognition of amino acids is performed

owing to the coordination interaction via metal cation of the porphyrin complex with the
simultaneous formation of H bonds at the periphery of the macrocycle. Porphyrinates, in
which the hydro group participating in the formation of H bonds is in naphthalene fragments
possess a high recognizing ability with respect to methyl esters of α-amino acids [72].
Based on studies of the complexing ability of porphyrins 67--69 forming and not forming
H bonds, the authors assessed the contribution of amino acid derivatives into the total en-
ergy (∆G°total) of forming ZnP--amino acid methyl ester complexes: 1) contribution of axial
coordination energy (∆G°ac); 2) contribution of H bonds (∆G°hb). While ∆G°ac in CH2Cl2
for 68 is ~15 kJ/mol, ∆G°hb is, depending on the nature of amino acid, within the limits of
1--6 kJ/mol. Of esters of aliphatic amino acids, porphyrin has a preference to amino acids
with bulky groups. The largest value of ∆G°hb is in leucine (6.2 kJ/mol); the lowest, in gly-
cine (0.8 kJ/mol). Though Ka in aliphatic and aromatic amino acids are commensurable, the
high binding constants of aromatic amino acids are determined not by the formation of hy-
drogen bonds, but by additional π--π interaction (Table 15).
Table 15 Association constants observed in binding of esters of amino acids and porphyrinates
(68, 69) in chloroform at 288 K.
ME of amino acid Ka (68), l/mol Ka (69), l/mol Ka (68)/Ka (69)
Gly-OMe 3.46×103 9.15×102 3.8

Ala-OMe 2.23×103 3.29×102 6.8
Val-OMe 8.07×103 3.51×102 23.0
Leu-OMe 1.09×104 2.72×102 40.0
Complexation of porphyrinates 70–76 with amino acid esters was studied in [74]. For-
mation of an additional hydrogen bond between the hydroxy group of porphyrinate and oxy-
gen atom of amino acid (complex 77) can be judged by the ratio of the association constants
(Ka) of porphyrins with hydroxy and methoxy groups of analogous structure. Thus, for three
diphenylporphyrins 71, 73 and 75, the hydrogen bonds with all amino acids studied are
formed only when the hydroxy group is in para-position (Table 16). The association con-
stants (Ka′) of complexes with two binding points (donor-acceptor and hydrogen bonds) dif-
fer from the association constants (Ka′′) of complexes with one binding point (donor-
acceptor bond) two- to fourfold. As seen from the data of Table 17, the energy of axial co-
ordination for all systems studied is within the limits of 22–24 kJ/mol, whereas the largest
value of hydrogen bonds, which are formed in Zn-porphyrin–amino acid methyl ester
H R
H3C CH3
70 : X = H
H9C4 C4H9 71 : X = OH-p CO2CH3
N N 72 : X = OMe-p NH2
Zn 73 : X = OH-m HO OH
74 : X = OMe-m Zn
N N
H9C4 C4H9 75 : X = OH-o
76 : X = OMe-o
H3C CH3
77
X
70-76
complexes, are ~4 kJ/mol. The number of phenyl fragments with hydroxy groups also af-
fects the value of Ka, but to a lower degree. The largest differences are observed in cases
with the symmetric and nonsymmetric arrangement of phenyl rings.
Table 16 Association constants observed in binding of esters of amino acids and porphyrinates
(72--77) in toluene at 293 K.
Zn-porph. (1) Ka, l/mol Zn-porph. (2) Ka, l/mol Ka' /K''a
Gly-OMe
70 1750
71 6270 72 1420 Ka(71)/ Ka(72) 4.4
73 2730 74 1430 Ka(73)/ Ka(74) 1.9
75 1680 76 1400 Ka(75)/ Ka(76) 1.2
L-Ala-OMe
70 1020
71 2660 72 915 Ka(71)/ Ka(72) 2.9
73 1680 74 920 Ka(73)/ Ka(74) 1.8
75 1080 76 910 Ka(75)/ Ka(76) 1.2
L-Leu-OMe
70 1660
71 2850 72 1120 Ka(71)/ Ka(72) 2.5
73 2480 74 2040 Ka(73)/ Ka(74) 1.2
75 1390 76 1380 Ka(75)/ Ka(76) 1.0
Table 17 Energies of hydrogen bonds (∆Ghb), axial coordination (∆Gac) and the total free
association energy (∆Gtotal) of porphyrin–amino acid ME in toluene.
∆Gtotal, ∆Gac, ∆Ghb, ∆Gtotal, ∆Gac, ∆Ghb, ∆Gtotal, ∆Gac, ∆Ghb,

kJ/mol kJ/mol kJ/mol kJ/mol kJ/mol kJ/mol kJ/mol kJ/mol kJ/mol
Gly-OMe L-Ala-OMe L-Leu-OMe
72 –27.2 –23.2 –4.0 –25.1 –22.1 –3.0 –24.9 –22.6 –2.3

74 –24.7 –23.2 –1.6 –23.6 –22.2 –1.4 –24.5 –24.0 –0.5
76 –23.6 –23.1 –0.4 –22.5 –22.1 –0.4 –23.0 –23.0 0
∆Gtotal = −RT ln Ka (ppm−1 ) , (2)
∆Ghb = − RT ln( Ka′ / Ka′′ ) , (3)
∆Gac = ∆Gtotal − ∆Ghb . (4)
The effect of hydroxyl groups, which are at both sides of porphyrin receptor 78, on its
recognition properties to amino acides is considered in [75[. The largest binding constants
(6.9·104 l/mol) are observed in the case of aspartic acid dimethyl ester with two –COOCH3
groups, whereas in leucine methyl ester the binding constant is four times as small (1.8·104
l/mol).
R N
N N
N NH2
OH
OH OH
OH
N N
Zn
N N N N
Zn
OH N N
OH
NH2
N
78
N
N 79
N
R
The recognizing ability of porphyrin receptors with respect to purine bases was studied
by the example of Rh(III) and Zn(II) complexes. Rh(III) complexes exhibit a very strong
affinity to nucleobases, especially to adenine. For instance, meso-tetraphenylporphyrin
Rh(III)·Cl has Ka in CH2Cl2 with 9-ethyladenine of the order of 107 l/mol, which in practice
indicates the irreversibility of this process [76]. 9-Ethyladenine is coordinated on Rh(III)
of the metalloporphyrin fragment by N-1 atom, which is indicated by X-ray data [71]. Un-
like Rh(III)-porphyrins, the coordination process of nucleobases on Zn(II) porphyrins is a
reversible process. Ka of Zn(II)porphyrin--purine base complexes of composition 1:1 are
within the limits of 102 -104 l/mol [77].
Table 18 Constants of binding (Ka, l/mol) of sugars by porphyrinates.
Porphyrin Sugars Ka, l/mol Solvent T, K Reference
37 Octyl-β-D-mannoside 4.8 ×103 CH2Cl2 297 78

37 Octyl-α-D-mannoside 6.4 ×103 CH2Cl2 297 78
37 Octyl-β-D-glucoside 1.4 ×103 CH2Cl2 297 78
37 Octyl-α-D-glucoside 8.5 ×102 CH2Cl2 297 78
76 Octyl-β-D-mannoside 2.2 ×103 CHCl3 288 80
76 Octyl-α-D-mannoside 3.0 ×103 CHCl3 288 80
76 Octyl-β-D-glucoside 1.6 ×103 CHCl3 288 80
76 Octyl-α-D-glucoside 2.9 ×103 CHCl3 288 80
Porphyrins with two hydroxyl groups on one side of the macrocycle form complexes
79 with adenine; in these complexes, one of the adenine molecules is coordinated by cation
of the tetrapyrrole fragment, and the other by the hydroxyl groups of the naphthalene frag-
ments. Such porphyrins are called “bidentate”, and the binding constants of such complexes
are several orders higher [77].
Complexes of sugars with Zn-porphyrins containing two nitrogen atoms at the periph-
ery of macrocycle 80 were studied in [78--83]. The hydroxyl groups of sugars were found
to be involved in the formation of bonds with: 1) Zn cation of the porphyrin’s reaction cen-
tre; 2) nitrogen atoms of the aryl substituent. The binding constants of these complexes are
presented in Table 18.
Besides Zn-porphyrins, some porphyrin ligands also possess a recognizing ability with
OR −
δO
δ− O H
+ O
δH O
δ− δ+H O
H δ−
N N
N N
δ+Zn
N N
80
NH N
N HN Br
Br N N
X X X
X Y X N
N Y X N N
OR OR N
NH HN OR OR
X N X X N X
N
OR OR OR OR NH N
Br N HN Br
X = (CH2)6 , Y = CO(CH2) 4CO

81 82
O O
O O O O
N N S
S N N
M O
M
N N O
N N
O
O
83 : M= H2, Zn
respect to sugars [84-86]. Thus, Kral et al. made use for this purpose of such porphyrin
receptors as macrocycles with 1,1′-binaphthosubstitution 81 or cyclic cryptand-porphyrin
conjugates 82 [84--86]. The latter are more efficient in aqueous media.
Increased interest in triple acceptor--donor--acceptor systems for studies of particular
stages of photosynthesis led to a large number of works on quinone-substituted porphyrins.
Of special interest are model compounds with well-defined geometry. Thus, in [87] it was
found that the conformational arrangement of the quinone fragment in porphyrin 83 de-
pended on the presence or absence of Zn cation in the tetrapyrrole macrocycle. In the pres-
ence of Zn, the quinone fragment is immediately over the reaction centre of porphyrin and
enters into a donor-acceptor interaction with it.
Porphyrins with four hydroxyl groups possess a recognizing ability with respect to var-
ious quinone substituents,, irrespective of whether it is metalloporphyrin or a free base.
R1 R1 R1 R1
OH
OH
OH N
NH - + -
- N
N
HN SO3 SO3 - SO3
N 2+ SO
N3
OH Zn
N N N N+
+
R R
+
N
84 R
85 : R1= O
O ,
R=
Especially high values of binding constants are observed in the case of porphyrin 84 and
methoxy-substituted quinones. Ka for 2,3,5,6-tetramethoxy-4-benzoquinone is 3.5·104
l/mol [88].
As an example of supramolecular porphyrin-based complexes formed only at the ex-
pense of dispersion interactions, we can name complexes formed by tetracations of Zn-por-
phyrins and tetraanions of calix[4]arenes [89--90]. The high strength of complexes formed
is indicated by high binding constants -- Ka of complex 85 in CHCl3 is 1.4·107 l/mol [89].
3 Complexation of Porphyrins with Organic Molecules: The Chirality

Aspect
In the recent years, among the great diversity of host--guest interactions, special attention
is attracted to complexation processes of porphyrins and related chromophores with organic
molecules possessing a chirality [22, 91--116]. This section considers the main principles,
mechanisms, driving forces and factors, which enable efficient control over the supramo-
lecular systems formed.
3.1 Host–guest systems based on monomeric porphyrins

Two major types can be singled out among host--guest systems based on monomeric por-
phyrins: 1) those including achiral porphyrins or racemates (optically inactive compounds
of equimolar amounts of antipodes); 2) those including porphyrins possessing a chirality.
In the former case, there occurs an interaction between the host molecule of optically inac-
tive porphyrin and the optically active guest molecule, which can be registered by various
spectral methods of analysis. Despite the apparent simplicity of systems based on mono-
meric porphyrins, they can possess some very interesting properties. For instance, a number
of highly substituted porphyrins (86 – 88) having a saddled shape owing to steric repulsion
between adjacent substituents, exist as two enantiomers, which, being mutually converted,
form a racemate mixture [117, 118]. It has been found that this equilibrium can be shifted
towards one certain conformation (more than 98%) at the interaction with various enantio-
pure acids via the formation of hydrogen bonds between carboxy groups and nitrogen atoms
of pyrrole fragments of the porphyrin cycle to form a 1:2 complex (one porphyrin and two
R1
R3 Me
Me
Me Me
NH N
NH N
R R
R2 R
N N HN
HN
Me Me
Me R1 Me R1
86-88 89
86 : R = Ph(OMe)2 -o,o; R1=R2=R3=Ph
R=PhB(OH)2-o, R1 =Ph
87 : R = R2= Rh(OMe)2 -o,o; R1=R3=Ph
88 : R = R1=R2=Ph(OMe)2-o,o; R3=Ph
Et Et
Et Et
N N
N
Zn
N N
N
Et Et
Et Et
90
acids). These supramolecular complexes have a rather large signal of induced circular
dichroism (CD) in the region of the Soret band, whose sign correlates with the relative value
of substituents at the asymmetric centre and makes it possible to determine the absolute con-
figuration. Although the authors do not discuss the electronic nature of observed optical ac-
tivity, induced chirality exhibits the properties of memorizing the optical activity in the
replacement of chiral acid by achiral acid. Another interesting example is the porphyrin
sensor for D-lactulose, whose selectivity is determined by the correspondence in the ar-
rangement of the reaction centres of sugar considered and two fragments of boric acid in
cis-porphyrin 89, which leads to the formation of a 1:1 complex with a CD signal of one
sign in the Soret band region [119]. The works [79, 120, 121] developed and investigated
the complexation properties of porphyrin-containing sensors for saccharides 68, 78, 90, 91
and alkaloids 92.
We would especially like to mention host--guest systems, which contain amino acids
being natural building blocks. Thus, using trans-isomers of porphyrins 68 and 90, the au-
thors of [75] developed a receptor for amino acids with a high selectivity with respect to
aspartic acid methyl ester (the binding constant corresponds to 6.98·104 M –1). The confor-
mational correspondence in this case creates conditions for three-point binding (one coor-
dinational interaciton and two interactions to form hydrogen bonds). In all systems studied,
a negative-positive CD signal of small intensity was found in the region of the Soret band,
induced by electronic interaction through the space between porphyrin and carbonyl group
of amino acid, though the influence of other factors is not to be ruled out, either. To expand
Ph
O(CH2)3OH
Me O(CH2)3OH
N N N
HO(H2C)3
N N Ph Zn PhOCH2COR-o
M
N N
N N
Et Me
Et (CH2)3OH Ph
92 : R = OH, NH2
91 : M = Lu(NO3), Gd(NO3)(Cl)
R1
XOOC(H2C)n
93 : M = Zn, R = R1 = COOX
N N
R1 M R XOOC(H2C)n
N N
X = Me or K, n=1,4,10
R1
93-95
O
X
O O
94 : M = Gd(acac), R = R1 = Y
O O
X O
X = t-Bu, Me, OMe, F, Cl, Br; Y = H, OMe

NHCO
95 : M = Er(acac), Gd(acac), Yb(acac); R = , R1 = Ph
the potentialities of using receptors for amino acids, the work [122] proposes a number of
porphyrins (93), which in nonpolar organic solvents can be used as esters, and in aqueous
media as products of their hydrolysis. It has been found that two factors contribute to the
complexation: electrostatic interactions in coordination of the amino group of amino acid
and Zn ion of the porphyrin cycle in organic solvents (enthalpic forces) and host--guest dis-
persion interactions (enthalpic force) in combination with the desolvation-controlled
PhSO3H-p Br Ph PhOMe-o
Br
Br
N N
NH N
Ph Zn Ph
p-HO3SPh PhSO3H-p
N N
N HN
Br Br
Br Br
PhSO3H-p Ph
96 97
process of complexation (entropic force). The latter predominate in water. In [123], recog-
nition of the optical activity of zwitterionic amino acids was studied in a two-phase organic
solvent--water system. For extraction from the aqueous phase, use was made of porphyrins
94, which formed 1:1 complexes with amino acids. Their CD spectra have two well-re-
solved opposite-sign signals, which correspond to a split Soret band and absorption bands
in the visible range of the spectrum. The value of CD molar amplitude (A) strongly depends
on the nature of solvent and the nature of substituents of phenyl rings. The largest sensitivity
was found for aromatic solvents and substituent X=t-Bu, and the sign of induced CD is de-
termined by chirality of amino acids. This method is proposed by the authors for determin-
ing the absolute configuration of α-amino acids. The sensitivity of this method was
subsequently increased by including a crown ester fragment into porphyrin molecule 95
[123]. Simultaneous coordination of the CO2– 2 group of amino acid by the lanthanide reac-
tion centre of porpyrin and the NH3+ group by the crown ester fragment in combination with
the respective choice of metal significantly increases the extraction capacity and value of
A. In [125], recognition of amino acids was done using the method of solid-phase optical
detection. It is based on hypsochrome shifts in the visible spectrum of porphyrin 96 immo-
bilized as a monolayer on a cellulose film, which occur owing to the interaction with amino
acids. The extent of spectral changes depends on the structure of amino acids, which makes
possible quantitative assays of amino acid in solutions.
In the literature, there is an large body of information on supramolecular systems, in
which both the guest molecule and the host molecule possess a chirality. On the whole, the
effect of chiral recognition is based on the principle of three-dimensional correspondence
according to the lock--key principle, when the complexing centres of the guest and host well
correspond to each other, and usually requires the presence of three sites for respective in-
teractions. Ideally, of two enantiomeric forms of the guest molecule, only one form would
be efficiently bound by the respective host molecule to form a stable complex, whereas the
other enantiomeric form woudl be bound weaker. This difference in binding can be estab-
lished by various spectral methods.
For instance, two diastereomeric complexes of porphyrin 97 and (S)-2-pyrro-
lidinemethanol were easily identified in [126] by the method of 1H NMR as a consequence
of the stabilization of the optimal complex due to coordination and formation of hydrogen
bonds. In another case, a direct correspondence was established between enantioselectivity
in complexation and catalytic oxidation [127]. Thus, at the substitution of Fe by Al, 1,1′-bi-
naphthyl derivative of porphyrin 98 was found to have clear distinctions in the electronic
O
R N
H
O
NH
N N
Cl MeO
Fe R=
MeO
N N
HN
O
H R
N
98
Ph
N N
O O
Ph Zn R
P
N N
O
100 : R =
Ph
99-101
SiMe3
O O
O O
P P
O SiMe3 O
99 : R = 101 : R =
absorption spectra in antipodal binding of epoxides and alcohols; these distinctions occur
against the background of the linear dependence between respective differences in free
energy for complexation and catalytic activity, which indicates the similar stereoselective
mechanism for both processes. Binaphthyl fragments were also used to obtain optically
active phosphite-containing porphyrins 99--101. They, in combination with various phos-
phorus-containing chiral and achiral ligands, were applied as supramolecular bidentate
102 : M=2H, R=R1=R2=R3= L-ValOMe

or L-ThrOMe or L-TrpOMe
NHCOR3
103 : M=Ru(O2),
R=R1=R2=R3=C(OMe)(CF3)Ph
R2CONH
N N 105 : M =Zn, R=R1=Ph,
+
R2=R3=CH2N Me3
M
N N
NHCOR
106 : M = Zn, R=R1=R2=R3=CH(NHBoc)Me

R1OCHN
107 : M = Zn, R=R1=R2=R3=CH(NHBoc)Pro
108 : M = Zn, R=R1=R2=R3=CH(NHBz)Gln
102, 103, 105-108
NO2
O
NH
O
HN
N N
Zn
N N
NH
O
O 2N
HN
O
104
matrices in palladium-catalyzed asymmetric allyl alkylation of racemates of 1,3-diphe-

nyl-2-propenyl acetate using dimethylmalonate as nucleophile [128]. The enantioselectiv-
ity was found to be determined not only by the absolute configuration of porphyrins and
ligands, but also by such a structural feature of porphyrins as voluminosity (introduction of
SiMe3 groups into porphyrin 101) and by the location of the substituent (ortho- and meta-
in porphyrins 99 and 100). The work [129] investigated amino acid-modified derivatives
of 5,10,15,20-tetraphenylporphyn 102, yielding with sugars complexes of composition 1:2.
It is noted that these porphyrins exhibit no advantages in complexing ability as compared
with the achiral analogue. At the same time, it is reported [130, 131] that the tetraphenylpor-
phyrin derivative possessing a chirality (103) exhibits chiral recognition abilities with re-
spect to amino acid esters. The largest value of enantiomeric excess (66%) was found for
methyl ester of valine; what is more, predominantly the L-enantiomer is bound. The respec-
tive Zn-porphyrin complex (103) can also efficiently recognize the [S]-antipode of bulky
1-(1-naphthyl)ethylamine with a 2:4 enantiomeric binding ratio [132]. The further improve-
ment of selectivity up to 7.5 (also for valine methyl ester) is achieved in the case of porphy-
rin 104 by way of respective coordination and hydrogen bonds between host and guest
molecules [132]. The thermodynamic analysis revealed a significant change in the guest
molecule owing to the coordination process, whereas the enthalpy factor determined mainly
by the formation of hydrogen bonds indicates the stabilization of the complex as a whole.
Two enantiomers of chiral water-soluble porphyrin 105 were also used for chiral recogni-
tion of a number of amino acids and dipeptides in an alkaline aqueous solution. The stere-
oselectivity changing from 1.2 up to 3.3 is based on a combined action of the effect of
coordination, Coulomb and spatial interations [134]. The work [135] describes one of the
most efficient ways of recognizing amino acid esters with an enantioselectivity of 21.54 (for
methyl ester of phenylalanine) using amino acid-modified porphyrins 106--108 [135].
Enantioselectivity was shown to depend exponentially on temperature, and it was found that
diastereoisomeric complexes exhibit various CD properties . In particular, the CD spectrum
of porphyrin 106 in the presence of weakly bound methyl ester of L-analine is similar to the
spectrum of individual 106 and represents a single negative Cotton effect in the Soret band
region. On the other hand, strongly bound methyl ester of D-alanine gives a positive-neg-
ative signal determined by exciton interactions between porphyrin and the carbonyl group
of amino acids. The importance of the model of three-site interactions for chiral recognition
was additionally confirmed by low stereoselectivity (15%) of the Ru complex of porphyrin
109 in the binding of racemate isocyaanides (it is capable of forming respective coordina-
tion complexes, but has no additional interactions sites related to the formation of hydrogen
bonds) [136]. In [137--141], by the example of chiral porphyrins 110, 111, it was clearly
shown using the methods of X-ray diffraction analysis and 1H NMR that a combinaiton of
one coordination interactin and two interactions by the type of forming hydrogen bonds pro-
vides for efficient chiral recognition of amino alcohols and amino acid esters as in the case
of the Co complex of porphyrin 110. Moreover, this porphyrin is suitable for the determi-
nation of the enantiomeric excess in primary amines and aziridines by 1H NMR, yielding
a linear dependence for a large number of enantiomeric compositions with a relative error
of several percent. Porphyrin 110 was also found to feature the dependence of the confor-
Me
O Me COOMe
O
R R
O O Me Me COOMe
N N N N
M M
N N O N N
O MeOOC Me Me
R R
O O
MeOOC Me
Me
Me Me
110 : M =Co, Fe
109 : R = O O
Me Me Me Et
Me
A
HO Et
OH O O
Me Me CONH(CH2 )2 NHCOCH2
O
Et Et
NH HN
Me Et
HO
Me N N Me
H
N
R R3 111 : R=R3=Et, R1=Me, R2=CH2A
112 : R= A, R1=R2=Et, R3= CH2COPheOBu-t

R1 R2
113 : R= A, R1=R2=Et, R3=CH2COPheOH
111-113
mational state of the macrocycle on the nature of the metal’s central atom; the dependence
is exhibited in the conversion of α,α,α,α-conformation to α,β,α,β-conformation at the sub-
stitution of Zn by Ni.
Various porphyrin analogues possessing a chirality are also used for binding optically
active compounds. Thus, in [142] complex lasalocid-sapphirine conjugates 111--113 were
used to transfer aromatic amino acids from one water phase into another water phase via an
organic layer. The proposed reaction mechanism includes the formation of a stable su-
pramolecular ensemble between the conjugate and zwitterionic amino acid owing to the
electrostatic coordination of carboxylate anion to protonated sapphirine and the formation
of hydrogen bonds between the NH3+ group and polyester ionophore fragment of lasalocid.
The above-considered examples of host--guest systems based on monomeric porphy-
rins clearly show the significance and broad potential of their applications. However, in
many cases the sole monomeric porphyrin fragment can not efficiently funciutn without the
synergic support of another porphyrin fragment, or several porphyrins. This aspect promot-
ed the development of new chirogenic host--guest ensembles based on dimeric and oligo-
meric porphyrins, which shall be considered in the next section.
3.2 Host–guest systems based on dimeric and oligomeric porphyrins

Host--guest systems based on dimeric and oligomeric porphyrins are distinguished depend-
ing on the presence of chiral fragment, type of covalent bridge and number of porphyrin
macrocycles. First, we shall consider examples of ensembles consisting of achiral and race-
mate bis-porphyrin molecules of the host and chiral molecules of the guest.
This type of chirogenic systems was initially used to observe asymmetry in the inter-
acting chiral molecule of the guest. On the whole, the achiral molecule of bis-porphyrin
should possess at least two interacting sites and the sufficiently flexible covalent bridge to
be capable of taking on a stereospecific three-dimensional conformation induced by the
chiral guest molecule. In this case, a supramolecular ensemble possessing a chirality is
formed.
R1
R
R2 N.
R N .
N
N
Fe N.
N . R
R N
N
R1
R
Ce
O
R R
N R2
R N
N N
Fe N
N
N R R2 N
R1
R
114-115 116-121
114 : R = PhB(OH)2-m, 115 : R = PhB(OH)2 - p 116 : R =R1=R2= p-pyridyl

117 : R =R2= p-pyridyl, R1=Ph
118 : R =R2= p-pyridyl, R1=Ph(OMe)2-m,m
119 : R =R2 = p-(N-CH2PhB(OH)2-p)pyridyl, R1=PhOMe-p
120 : R = p-(N-CH2PhB(OH)2-p)pyridyl, R1=R2=PhOMe-p
121 : R = R2= p-(N-Me)pyridyl, R1=PhOMe-p
The signicance of these two factors is easily understood from the following examples.
Thus, double-decker bis-porphyrins 114--121 linked centre-to-centre were developed
based on µ-oxo-dimers and Ce complexes to observe the chirality of various guest mole-
cules [143--148]. The association mechanism includes the interaction of the bidentate guest
with two complexing groups of adjacent porphyrins, which makes these porphyrins rotate
aroung the central axis and, respectively, to turn macrocycles left or right depending on the
stereochemistry of the guest molecule. This asymmetric travel leads to the emergence of a
significant optical activity (the consequence of interporphyrin exciton interaction) in the re-
gion where porphyrin absorbs. Though the electronic nature of induced CD has not been
completely explained, these systems were successfully used to observe chirality of saccha-
rides (for 114, 115, 119 and 120), dicarboxylic acids (for 116--118), dianions (for 121) and
memorize chirality (for 118). The generated optical activity was excellently stored for three
days at 0°C and even for one year at --37°C. Host--guest complexation occurs with a high
degree of cooperativity, exhibiting a positive allosteric effect, and CD properties strongly
depend on the guest [143, 144] and solvent [148] molecule structure and pH [147]. It is in-
teresting to note that the effect of a change of optical activity controlled by the number of
saccharide links was established for porphyrin 119 in the case of maltooligosaccharide
guest molecules. An entropically unfavourable face-to-face arrangement of bis-porphyrins
can be also preserved owing to the diametrically located covalent bridges as in the case of
cryptand-containing porphyrins 82, 122 [86]. These host molecules are also capable of
binding various saccharides predominantly by encapsulation. An observed very weak in-
duction of CD in the region of Soret band is a consequence of the inability of considered
bis-porphyrins to follow the stereochemistry of the guest molecules, probably, because of
the structural features of the linking bridge. More flexible covalent bridges of various struc-
tures and lengths were used to ensure the face-to-face orientation in porphyrins 123--125
[149, 150]. bis-Porphyrins 123, 124 with a longer bridge showed a high correspondence to
R R2 R1 A
R1 R
NH N
(CH2)6 N
N HN
CO(CH2)4CO (CH2)6
R1 R1 N
R R2 R
A
A R R2 R1
(H2C)6 (CH2)6
N
R1 R N
NH N (CH2)6
N HN
R1 R1
R R2 R
R R1 R
122 R R
N N
122 : R=Pr, R1 = Me, R2=H M
R R1 R
122 R R
N N
122 : R=Pr, R1 = Me, R2=H M
N N
R R
R R1 R
A R1
A
R R
R R
N N
M
N N
R R
R R1 R
123 : M=Zn, R=H, R1=Mes, A = -CO(NHCMe2CO)9NH

-
123-125
124 : M=Zn, R=H, R1=Mes,
A= -CO(NHCMe2CO)3NHC(CHPh)CONHCH2CONH(CHPh)CO(NHCMe2CO)3NH-
125 : M=2H, R=Me, R1=Ph(OMe)2-o,o, A = -OCH2PhCH2O
-
bis-pyridyl substituted chiral guest molecules of respective length, which leads to the for-
mation of stable inclusion complexes of composition 1:1 with association constants of the
order of 2.3·106 M-1. It was found that the helicity of the guest molecule plays a key role
in transferring information of chirality and induction of a strong exciton interaction in the
reigon of Soret band. bis-Porphyrin 125 with a shorter bridge was used for complexation
with α-hydroxybenzeneacetic acid. Herewith, the activity of CD (A =260 cm – 1 M –1) was
observed to increase significantly (more than sevenfold) as compared with respective
monomer, owing to the conversion of chirality due to the nonplanar structure of the por-
phyrin ring to that due to the helicity of the ensemble as the whole. The stepwise increase
of the concentration of the guest molecule leads to complex changes of CD signals, which
reflect a multilevel equilibrium of the complexation process. Although the observed chiral
responses have not yet been totally explained, these chiroptical systems are of certain in-
terest as potential sensors of chirality in studies of the stereochemistry of interacting guest
molecules. Another structural type of bis-porphyrins is based on combining two chro-
mophore fragments via one covalent bridge. As the mutual arrangment of the porphyrin
R1 R R1 R
R R
R2
R2 R2 R2
N N N N
R1 M A M R1
N N N N
R2 R2 R2 R2
R R1 R R R1 R
126-133
126 : M =Yb(acac), Gd(acac), R=R2=H, R1=Ph 127 : M =Yb(acac), Gd(acac), R=R2=H, R1=Ph
A CH2O(CH2)2OCH2
A CH2OCH2
129 : M = Zn, R = R2 =H, R1 = Ph(t -Bu) -m,m

128 : M =Zn, R = Me, R1=H, R2=n-C6H13
MeO2C CO2Me
A
A
macrocycles becomes less predictable in this case due to the reduced rigidity of the system
as a whole, a number of additional factors, such as complexation with the bidentate guest
molecule or shortening of the bridge, can bring two porphyrin fragments closer to each oth-
er and fixe bis-porphyrin in face-to-face orientation or linear conformation. Thus, series of
lanthanide bis-porphyrins 126, 127 linked with ester bonds of various lengths were prepared
as potential host molecules for binding polyions of cystine [151]. These receptors exhibit
a significant selectivity to size in studies of the optical activity of guest molecules. Thus,
the Yb complex of porphyrin 126 with a short bridge efficiently extracts cystine from an
aqueous solution to organic phase, whereas cystathionine, homocystine and methionine are
extracted weakly. In contrast, porphyrin 127 preferably extracts a longer homocystine.
Complexation occurring via the formation of tweezer complexes of composition 1:1 leads
to a complex CD signal in the Soret band region, which is significantly increased as com-
pared with respective monomer. The authors of [152] report of a high selectivity to length
in binding of chiral diamines by porphyrins 128, 129 with rigid bridges. The dimensional
correspondence in the host--guest system is an important condition for efficient complex-
ation with the association constant of the order of 2.4·106 M –1 for the best pair. Induced
CD strongly depends on the volume and number of stereogenic sites, peripheral substituents
of the porphyrin macrocycle and temperature. For instance, the largest value of A (1340
cm –1 M –1) was increased 1.5-fold by decreasing the temperature to --45°C. The binding
link of the type of crown ester significantly enhances the potentialities of controlling the
conformation of bis-porphyrin via specific complexation with ions of alkaline metals
[153--155]. This effect is beautifully demonstrated by the example of an almost twofold in-
crease of the binding constant (from 2.6·105 M –1 to 4.5·105 M –1) upon addition of Na+
ions to porphyrin (130)/trans-1,2-diaminocyclohexane system of the tweezer type due to the
change of geometry of bis-porphyrin. Addition of Na+ ions significantly increases the value
of A of induced CD. A similar effect of binding and CD increase was found in [154] for
porphyrin 131 at the interaction with N-alkyl substituted 1,2-diaminocyclohexane. Howev-
130 : M =Zn, R = R2=H, R1=Ph 131 : M = Zn, R = R2 = H, R1 =Ph
A (OCH2CH2)4O
OMe MeO
A N O N
3
O O O
CONH NHCO
132 : M =Zn, Mg, R = R2 = H, R1 = Ph
A CO2(CH2)5OCO
133 : M =Zn, Zn/2H, R=R2=Et, R1=H, A = -CH2CH2
er, a larger-size K+ ion is required in this case for a larger crown ester cavity. It is noted that
porphyrin 131 is capable of recognizing the chirality of such potassium carboxylates as
camphorates and mandelates. What is more, induced chirality of porphyrin 57 in the twee-
zer conformation was efficiently saved in the substitution of chiral diamine for achiral one
in the presence of Ba ions. The authors of [155] note that in 24 h the intensity of CD in this
system decreases only insignificantly. Although the discussed systems of the tweezer type
exhibit a rather high optical activity due to the chiral spatial arrangement of two porphyrin
chromophores, no detailed explanation of electronic and structural factors involved in this
phenomenon has been given until now.
One of the first attempts to solve this issue in the case of bis-porphyrin systems was
made in [156--166] using a more flexible porphyrin 132, which is capable of taking on a
tweezer conformation in the complexation with bidentate ligands. It has been found that the
emergence of optical activity is based on the stereospecific differentiation of the relative
volume of substituents at the asymmetric carbon atom due to the bis-porphyrin molecule
taking on a spatially less hindered conformation. It should be also taken into account that
other factors, such as formation of hydrogen bonds; presence of heteroatoms, solvent etc.,
can also affect the geometry of the ensemble as a whole. All this leads to an oriented ar-
rangement of two tetrapyrrole fragments in porphyrin 132, which on the strength of exciton
interactions induces a CD signal, whose sign can be used as a tool for determining the ab-
solute configuration of various bidentate guests. In the cases when the relative size can not
be determined directly, the conformation analysis is made using calculations according to
the method of molecular mechanics with the view of explaining the observed contradiction
between the predicted chirality and the one obtained in some cases. Though it is well known
that metalloporphyrin chromophore contains two degenerated (or almost degenerated, due
to the asymmetry determined by meso-substitution) electronic transitions along the meso-
5-15 and 10-20 axes, only the exciton interaction of the pair of 5-15 oscillators was chosen
for explaining the induced optical activity in porphyrin 132. However, the contribution of
10-20 oscillators due to homo- and heteroconjugation can also play an important role in cer-
tain conformations. The works [156--166] also adduce some dependences of induced CD
on the nature of the guest molecule, the solvent and temperature.
To investigate various aspects of optical activity of supramolecular systems based on

bis-porphyrins and to explain the chirality induction mechanism, the works [98, 167--181]
studied bis-porphyrin 133, in which porphyrin fragments are linked by an ethane bridge. In
contrast with the above bis-porphyrin systems, this host molecule possesses an ability to
sense the chirality of not only bidentate but also monodentate guest molecules due to the
semi-flexibility / semi-rigidity of the bridge. This ability is provided for by the relatively
short but sufficiently flexible C2 chain. As shown by the analysis of the absorption and CD
spectra, porphyrin 133 forms with primary amines, by the cooperative mechanism, stable
extended guest--host complexes of composition 1:2 with free Gibbs energy within the range
of –6.7 to –8.4 kcal·mol –1. The specific chirogenic mechanism for porphyrin 133 in com-
plexation with monodentate guest molecules includes competitive repulsive interactions
between two most bulky substituents at the asymmetric centre of the ligand and ethyl groups
of an adjacent porphyrin fragment. The most bulky group makes adjacent macrocycles
move away one from another, thus generating a directed turn. The latter leads to the emer-
gence of an (average to strong) signal of CD exciton interaction in the Soret band region.
The chirality sign correlating with induced helicity makes possible the determination of the
absolute configuration of monodentate guest molecules. The authors thoroughly discussed
the mechanism of host-guest interactions, which enables a detailed investigation of various
external and internal factors affecting the chirality induction processes. For instance, it was
shown that the value of A linearly depended on the size of the largest substituent at the ste-
reogenic centre for homologous ligands, thus making it possible to predict induced chirality
[167, 168, 170]. The crucial role of solvent as an active part of the supramolecular system
on the whole was clearly explained through the thorough analysis of solution--solvent se-
lective interactions in near-boundary regions, where the distinctions between competing
chiral interactions are small and the relative volume of a substituent can be formed owing
to the interaction with the solvent [175, 176]. Temperature was found to be another impor-
tant factor for controlling chirogenic processes. According to [171--173], a decrease of tem-
perature, significanlty increasing the values of A, makes possible induction of chirality by
binding alcohols, though they are known to possess a low affinity to Zn-porphyrins. How-
ever, the binding of alcohols and the ability to generate optical activity were significantly
enhanced by the substitution of the central Zn ion in porphyrin by the Mg ion [174]. The
authors of [181] note the importance of the phase transition as a factor of controlling the
optical activity in porphyrin (133) / monodentate amine complexes, which leads to a sig-
nificant increase of the anisotropy factors (16--9.1-fold) and solid-phase “switchover” of
chirality owing to the formation of intermolecular aggregates of opposite helicity. Interac-
tion of porphyrin 133 with bidentate guest molecules leads to the formation of extremely
stable tweezer complexes of composition 1:1, with binding constants > 107 M –1. A further
increase of the concentration of the bidentate guest shifts the supramolecular equilibrium
towards the extended complex of composition 1:2, which makes it possible to study the re-
spective stoichiometric effect [177--180]. In the case of enantiomeric 1,2-diphenylethyl-
enediamine, an excellent chirality-switchover phenomenon was revealed, which is
controlled by the stoichiometry of the supramolecular system as the result of the opposite
mutual spatial arrangement of 1:1 and 1:2 complexes. A more detailed explanation of in-
duced optical activity in supramolecular systems based on bis-porphyrins was first achieved
using chiral 1,2-diaminocyclohexane. In particular, based on the obtained crystallographic
structure of the (133) / (R,R)-1,2-diaminocyclohexane complex and the Кuhn--Kirkwood
oscillatory interaction mechanism, CD was explained as a combination of two B||–B|| and
B⊥ –B⊥ electronic homo interactions, which are oriented counterclockwise and are exhib-
ited as an intensive negative--positive CD signal (A = –590 cm –1 M –1), whereas the con-

tribution of respective B|| –B⊥ hetero interactions is insignificant [180]. Thus, the high
sensitivity, complete understanding of the mechanism of the process and broad applicability
for various types of guest molecules show that porphyrin 133 can serve as a potent tool for
studies of various aspects of supramolecular chirogenesis and for using as a universal sensor
of chirality.
Another bis-porphyrin with one bridge (134) was proposed for selective binding of oli-
gosaccharides by the cooperative mechanism due to the use of advantages of the increased
rigidity of the system considered [147, 182]. In particular, a fixed porphyrin--porphyrin dis-
tance matches well the size of maltotetraose, which leads to the formation of a stable 1:2
complex. This is reflected as a complex chiroptical response in the Soret band region, which
response consists of three nonequivalent and asymmetric Cotton effects, probably, due to
the superimposition of two or more CD signals. The further increase of the number of por-
phyrins in respective meso--meso linked oligomers (135) led to a significant increase of in-
duced chirality [183]. These oligomers form heliciform structures owing to the fixation of
meso-aryl substituents by way of forming hydrogen bonds between carboxyl groups of por-
phyrin and cyclic molecules of urea. The direction of the helix can be controlled by com-
plexation with the chiral diamino derivative of 1,1′-binaphthyl, which leads to induced CD.
The intensity of CD was increased by increasing the number of porphyrin links from 2 up
to 8. An interesting effect of chirality memorizing was registered for porphyrin 135 (n = 8).
Upon addition of the antipode to the helix formed by the opposite enantiomer, the sign of
induced chirality decreased, but not disappeared completely.
Another type of two-dimensional multiporphyrin arrangement was studied in [184,
185] for studies of their chirogenic properties. It was found that hexamers 136 induced a
strong CD signal in the Soret band region in the presence of enantiopure (1-naphthyl)ethy-
lamines, whereas other chiral amines lead only to weak Cotton effects. An even stronger
chiroptical selectivity was found for a series of dendrimeric porphyrins 136--140 in com-
plexation with bidentate chiral guest molecules. In particular, induced CD strongly depends
on the number of porphyrin fragments, and the maximal value of A (2693 cm –1 M –1) is ob-
served for 138. However, it should be noted that this effect has not be comprehensively
studied to date.
R R R
N N N N N N
Zn Zn Zn
N N N N N N
R R R
n-2
134, 135
CO2H
O
134 : n = 2, R = B 135 : n = 2, 3, 4, 8, R = OC12H25
O CO2H
R
A
R R N N
136 : R = Zn B
N N
R R
A
A = B = Ph(t-Bu)2-m,m
R or
A = H, B = Ph(OC8H17)2-m,m
136-140
O2C X
O2C X
O2C X
137 : R = O2C
O2C
O2C X
138 : R = O2C
O2C
O2C X
O2C X
Further modification of the geometry of porphyrin was done in [186] with the view of
developing supermolecular systems for chiral recognition and enantioselective catalysis
based on heterometallic trimer 141. At the first stage, various chiral carboxylic acids were
totally linked to Sn porphyrins 141 by the type of hexacoordination to form a 1:4 ensemble
possessing a chiral cavity. However, subsequent attempts of optical separation of the race-
mate mixture of chirally substituted pyridines and catalytic asymmetrical epoxidation were
unsuccessful, apparently, due to the loss of rigidity inside the chiral cavity.
A broader application in various fields proved to be chirogenic host--guest systems, in
which both component parts possess a chiral activity due to a number of specific properties.
In particular, chiral recognition is the most appropriate application of various derivatives
of chiral bis-porphyrins. Thus, a series of bis-porphyrins 142--144 was prepared in [187]
for recognition of dicarboxylate anions via a combination of Coulomb attraction and for-
mation of hydrogen bonds. It was found that porphyrins 142 and 143 formed strong com-
plexes with N-carbobenzoxyaspartic acid (N-Cbz-Asp) and N-carbobenzoxyglutamic acid
(N-Cbz-Glu) with association constants within the limits of 104--105 M –1 and exhibit a
preference towards glutamic acid as compared with aspartic acid. Porphyrin 143 shows an
insignificant enantiomeric selectivity, while cyclic and more rigid porphyrin 144 exhibits
a low affinity to guest molecules considered, but shows a significant chiral recognition. Ap-
parently, the latter occurs from a good dimensional and geometric correspondence between
the host and guest as the result of fixation of two porphyrin fragments by two covalent
bridges, however, the detailed mechanism of recognition still requires additional studies.
O2 C X
O2 C X
O2C X
O2 C X
140 : R =
139 : R = O2C O2C X
O2 C
O2C X
O2 C
O2 C
O2 C X
O2 C X
O2 C X
N N OCH2 Ph(OMe)2-m,m
X = O2C Zn
N N
OCH2Ph(OMe)2-m,m
MeO2C(CH2)2 (CH2)2CO2Me
Me Me
N N
Ru(CO)(Py)
N N
Me Me
Me MeO2C(CH2)2 (CH2)2CO2Me
(CH2)2CO2Me
MeO2C(CH2)2 Me
Me
N Me
N
N
Sn(OH)2 (CH2)2CO2Me
MeO2C(CH2)2 N N (CH2)2CO2Me
Sn
MeO2C(CH2)2 N (OH)2
N
Me N
Me
MeO2C(CH2)2
Me Me (CH2)2CO2Me
141
Spatially fixed porphyrin 145 was used to produce 80--86% and 48% enantiomeric ex-
cess in binding of esters of histidine and lysine by means of two-site interaction between
two nitrogen atoms of the guest molecule and two zinc ions of the host molecule to form a
tweezer structure [188]. Similarly, for chiral recognition of amino acid derivatives the work
[152] used porphyrin 146 rigidly bound via 1,1′-binaphthyl fragment. This host molecule
exhibits an extremely high affinity to lysine derivative due to the formation of tweezers.
Me
Et
Et Me
Et
N
Me Et
NH
Me N
HN Me
HN
R
Me NH
NH
N
Me HN
Me
Et N
Et
Me Et
Et
Me
142,143
142 : R = A, 143 : R = B
-(H2C)2OCHN -(H2C)2OCHN
A = B =
-(H2C)2OCHN -(H2C)2OCHN
Me
R
Et
Me
N
Me Et
NH
Et
HN N
Et HN Me
Me NH Et
N NH
Et
HN
Et Me
N
Me Et
R
Me
-(H2 C)2OCHN
144 : R =
-(H2 C)2OCHN
Optimization of the geometry of host--guest complexes showed that the selectivity mechan-
ims is based on spatial repulsion between two methoxy groups of porphyrin 146 and the
amide group of the amino acid derivative in the less preferable diastereomeric complex. Af-
ter inclusion of the chiral residue of leucine into the covalent bridge of achiral dimeric por-
phyrin 124 the obtained chiral compound 147 was used by the authors of [189] for optical
separation of a series of artificial bidentate ligands of different lengths, containing a leucine
group in the middle of the molecule (the source of optical activity) and two end pyridine
(providing for the two-site binding. A 80% value of enantiomeric excess with significant
R R R R
N N
N
N Zn N N Zn N
N
N N
R R R R
145 : R = Ph(t-Bu)-m,m
Me Me Me Me
Et Me
Me Et
N N N N
Zn A Zn
N N N N
Et Me Me Et
Me Me Me Me
146 : A MeO
MeO
N N
Zn
N N
R
A R A
N N
Zn
N N
147 : A = -CO(NHCMe2CO)3NHC(CHPh)CONHCH(i-Bu)CONH(CHPh)CO(NHCMe2CO)3NH-
enhancement of the CD signal in the Soret band region due to the formation of a preferable
tweezer complex. Although the detailed mechanism of chirality enhancement is not pre-
sented in the work, comparative studies showed the observed effect to be stipulated by the
stabilization of the twisted geometry of the host molecule due to the helicity of the guest
molecule. An opposite tendency is described in [190]. In particular, in the presence of cit-
Ph Ph
N Ph Ph N
N N
Co Co
N N
Ph N N Ph
O O
O O
S S
148
Et Et
Et Et
N N
Zn CH2 *
N N
Et Et
Et Et
149
rene (1-methyl-4-isopropenylcyclohexene-1) a negative single CD signal of porphyrin 148

was significantly decreased and hypsochromically shifted with a different degree of mod-
ulation of chirality for two enantiomers. A similar CD signal decrease was found in the in-
teraction of chiral bis-chlorine 149 with chiral ligands due to induced conformational
changes; herewith, the chiroptical response of antipodal ligands differs significantly [191].
This makes it possible to use chlorine 149 for purposes of chiral recognition, using the new
principle of enantioselectivity based on the model of two-site host--guest interaction in
combination with the model of exciton interactions of chromophore fragments of dimer, the
chiral arrangement of which is controlled by the stereochemistry of the guest molecule.
4 Conclusion
Material presented indicates that supramolecular systems based on porphyrins are of un-
doubted interest for establishing the absolute configuration of organic molecules of various
nature and developing molecular receptors for a certain type of substrate. The occurrence
of tetrapyrrole chromophores in such systems enables application of spectral methods tra-
ditionally used in porphyrin chemistry for studies of molecular recognition processes. Re-
cent advances in this field are a promising prerequisite for the development of new
molecular technologies, such as chiral nanotechnology, intelligent molecular devices and
chiroptical memory.
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5 Chemical Activation of
Porphyrins in Coordination
Core Reactions
D.B. Berezin1,2 and B.D. Berezin1

1Institute
of Solution Chemistry, Russian Academy
of Sciences, 153045 Ivanovo, Russia
2Ivanovo State University of Chemistry and Technology,
7 F. Engels Prospect, Ivanovo, 153000, Russia;
email: berezin@isuct.ru
Depending on the molecular structure and effects of the medium, porphyrin NH

bonds of the coordination core can be considered as localized or partially delocal-
ized. Delocalization can be caused by intramolecular effects such as molecule pola-
rization by means of push-pull substituents or special nonplanar macrocycle con-
formations and by a series of external factors like strong solvation interactions in a
solution containing the electron-donor component in a polymer state, sorption or
other types of solid state effects. Activation of NH bonds in porphyrin molecules
results in crucial changes of their coordination core reactivity. This chapter
analyzes the causes and ways of activation in relation to the possibility of classi-
fying porphyrins by their ability to activate or not to activate NH bonds. A number
of reliable quantitative criteria allowing to talk about the degree of the chemical
activity of NH bonds for porphyrins or their analogues of any structural group is
presented. Activation of NH bonds can not only affect the H2P core reactivity but
in some cases cause a reorganization of π-chromophore and reaction centres, which
are different tauthomeric processes. Products of such a reorganization are good
models for biological supramolecular systems. Chemical activation of porphyrin by
itself is considered as a powerful tool for the reactivity control of these molecules,
including biological aspects.
Keywords: porphyrin, coordination core, chemical activation, reactivity control

170 D.B. Berezin and B.D. Berezin
Introduction
Coordination cores in molecules of porphyrin ligands (H2P) and their complexes with p, d
and f metals (MP) are some of their main reaction centres [1–3]. These are complex mul-
tifunctional centres, as a rule, N4H2 and MN4, respectively. Their unique reactivity is de-
termined by a combination of a number of factors. They include not only the chemical
affinity of atoms and atomic groupings constituting these cores, with respect to electro-
philic-nucleophilic reagents, their capability of synchronous donor-acceptor interactions,
the possibility of redistributing electronic density from one atomic grouping to another for
reasons of their involvement in the united system of π-conjugation [1, 4]. One of the most
effective factors is a relatively rigid shielding of these reaction centres by an atomic envi-
ronment and π-electron cloud of the aromatic macrocycle (Fig. 1). The extent of shielding,
in turn, can change under the influence of external effects. This phenomenon was given the
name of the macrocyclic effect (MCE) of porphyrins and metalloporphyrins [4–8]. MCE
affects the physicochemical properties and reactivity of H2P and MP by means of π-elec-
tronic and structural (steric) components. Their effects on various physicochemical pro-
cesses occurring in the molecule are, as a rule, different and nonsymbate one to another [8].
(a) (b) (c)
Figure 1 Atomic and π-electronic shielding of the reaction centre MN4 of metalloporphyrins MP
(a), X(MP) (b), (X)2MP (c) [4–5].
Initially, research to achieve understanding of the mechanisms of complex enzymic

reactions involving porphyrins and their analogues was aimed at studies of the electronic
effects of substituents located mainly at the periphery, i.e., in β- and meso-positions of H2P
and MP molecules, and their influence on the structure and properties of molecules [1,
9–11]. Namely the functional substitution was considered to be the most efficient means
of controlling the properties of H2P and MP. Herewith, most porphyrins were traditionally
seen as planar molecules and some (often significant) deviations from planarity, revealed
using X-ray diffraction analysis (XDA) of crystals, were referred to crystal packing effects
[1, 12, 13]. The last quarter of the 20th century witnessed the appearance of a large number
of new porphyrin structures with unusual spectral and other physicochemical properties,
drastically different from the earlier known compounds [14]. A large part of them are por-
phyrins with a strongly nonplanar structure [15–17]. As the result, an idea was proposed
and took root in late 1980s that the key role in the exhibition of their unique properties by
porphyrins and their complexes in the bioenvironment, the properties of the coordination
core including, is played by the conformational mobility of the macrocycles [18, 19].
However, in recent years authors again come back increasingly often to the role of
polarization effects interrelated with the planar structure of H2P [20–24]. Indeed,
conformational conversions of H2P molecules can cause secondary changes due to the
asymmetric redistribution of π-electronic density in them. The emerging polarization of
molecules on the whole and separate chemical bonds in particular, e.g., the chemical acti-
vation of NH bonds, can drastically change the reactivity of compounds.
Of special significance is the role of the medium, in particular, the nature of solvent
(if processes are run in a liquid-phase system), which is capable of significantly intensifying
or inhibiting the polarization processes in H2P molecules.
Thus, another important factor, which along with atomic electronic shielding (MCE),
determines the reactivity of porphyrins in vivo, is polarization of the molecule and, as a rule,
chemical activation of NH bonds in its coordination core. This conclusion has been evident
earlier. It follows, e.g., from the reaction mechanism of complexation of porphyrins with
d metal salts [1, 5, 25], the limiting stage of which is namely polarization by the solvent and
dissociation of NH bonds of the H2P macrocyclic ligand.
An important problem, which needs to be solved, is to elucidate the relation between
the nonplanar structure of the porphyrin macrocycles and the chemical activity of NH bonds
in the coordination core of these molecules. As a particular case of a jumpwise change of
the properties under the influence of conformational regroupings, the phenomenon of NH
activation is very important for modelling and studying the processes involving H2P in vivo,
where in the course of acceptance of metal ions by bioporphyrins NH activation is per-
formed also under the influence of the donor-acceptor protein-lipid environment.
1 Porphyrin Ligands with Localized and Delocalized NH bonds

In accordance with a classification proposed with account for the significance of the chem-
ical activity of NH bonds, porphyrins can be divided into three groups [26]:
• with the planar aromatic π-system and chemically low-active NH bond;

• with the nonplanar π-system of aromatic chromophore and chemically active NH bond;
• with the planar aromatic π-system and chemically active NH bond.
The first group includes many porphyrin bioligands, e.g., chlorophylls, blood porphy-
rins, cytochromes [5, 27], as well as their synthetic β- and meso-substituted analogues, e.g.,
β-octaethylporphin {I, H2(β-Et)8P}, meso-tetraphenylporphin {II, H2TPP} and meso-
tetrapropylporphin {III, H2T(n-Pr)P}. These compounds are also called classical H2P [28],
R1 R R1 R X
N
R1 R1 X
NH N meso-position NH N NH N
R R R R N N
N HN β−position N HN N HN
R1 R1 X
N
R R1 R X
R1
I–III, V–VII IV, X VIII–IX

I. R = H, R1 = Et; II. R = Ph, R1 = H; III. R = n-Pr, R1 = H; IV. R = Ph; V. R = Ph,
R1 = Et; VI. R = R1 = Ph; VII. R = Ph, R1 = Br; VIII. X = H; IX. X = Br; X. R = H.
because the properties they exhibit totally correspond to those traditionally known in por-
phyrin chemistry. The second group are dodeca-substituted (peripherally highly substitut-
ed) H2P (compounds IV–VII) [28–33] and some other strongly nonplanar molecules. The
third group comprises rigid highly aromatic porphyrazines (compounds VIII–IX) [34–37],
as well as tetrabenzoporphyrins {e.g., H2TBP (X)}, having substituents in condensed rings
but not in meso-positions [38]. Meso-substituted tetrabenzoporphyrins (e.g., IV) belong to
the second group of compounds, dodeca-substituted porphyrins [31–32, 38–39]. H2P with
nonclassical properties constitute the second and third groups of compounds [26, 28].
1.1 Factors causing the delocalization of NH bonds in porphyrin molecules

The high chemical activity of NH bonds in H2P molecules, related to their partial or total
delocalization [1, 4, 5, 35], can be achieved if several conditions are satisfied [28, 39]:
• the structure of the macrocycle is changed to become rigid (aromatic) or, vice versa,
strongly nonplanar;
• electron-acceptor groups or groups of various electronic nature which lead to the
push-pull-type polarization of π-chromophore [40–43], are present in the molecule;
• the properties of the medium, contributing to the polarization of NH bonds of the H2P
molecule, are changed by means of solvation and chemical interactions, e.g., with
molecules of electron-donor solvents [4–5], or by adjusting the reaction NH centres of
porphyrins to the reaction centres of the medium in the solid phase during the sorption,
immobilization etc. [44–45].
At least two out of the three conditions, each of which is necessary but not sufficient,
are required to be satisfied to achieve the NH activity of H2P.
1.1.1 Spatial structure and polarization of the porphyrin molecule

Analysis of vast literature data [4–5, 19, 22, 29–30, 34–37, 46, 56–57] and own studies
[7–8, 24, 26, 28, 31–32] indicate that the porphyrin ligand molecule equally acquires a ca-
pability of delocalizing NH bonds both when it becomes more planar and highly aromatic
and, vice versa, under the influence of significant extraplanar distortions of the π-chromo-
phore. As a result, a situation, at first glance paradoxical, emerges when opposite structural
changes lead to the same type of changes in some properties of the molecules [58]. Thus,
for instance, with the advance of benzo- and aza-substitution of porphyrin molecules their
Saddle Ruffled Domed Wave
Figure 2 Main types of nonplanar conformations of H2P and their complexes ( and assume the
location of atoms over and under the initial plane of the macrocycle, respectively) [16, 19].
structural rigidity is enhanced in the sequence: porphyrins proper → benzoporphyrins →

azaporphyrins, which is supported by numerous quantum chemical [35, 46, 48, 59], X-ray
diffraction [1, 35] and fluorescence [48, 60] data. In the case when the periphery of the H2P
molecule, i.e., its β- and meso-positions, is overloaded with bulky functional groups, por-
phyrin becomes more apt to extraplanar deformations and by the geometry of the most sta-
ble conformation belongs to one of the known types [16, 19]. Among them, the most
frequent types are saddle, ruffled, domed and wave conformations (Fig. 2), as well as stepp-
ed conformation; their combinations are also widely known, which are especially charac-
teristic of porphyrins with asymmetric substitution. Thus, a combination of the ruffled and
domed conformations yields a gabled structure [57]. The largest distortion is achieved if
even at free β-positions all four meso-positions are substituted by bulky groups (iso-Pr,
cyclo-Hex, tert-Bu {compounds XI–XIII}). The distortion of the porphyrin macrocycle
also increases to a maximum with the increase in the number of peripheral substituents of
moderate voluminosity up to twelve (compounds IV–VII) at eight β-positions occupied.
Each of the typical conformations can be characterized in
one way or another by the degree of intensity of the nonplanar
R
structure. For instance, saddle conformations typical of dode-
ca-substituted H2P (compounds IV–VII), as well as of a sig-
nificant number of Zn porphyrins (ZnP) (Table 1) are charac- NH N
terized by the mean deviation of β-carbon atoms from the ini- R R
tial plane of the macrocycle (∆Cβ, Å) obtained from XDA da- N HN
ta. The value of ∆Cβ changes for the currently known strongly
nonplanar, saddle-distorted porphyrins and their complexes
from 0.7 up to more than 1.3 Å [19]. A deviation of the order R
of 1 Å is considered to be very significant. The ruffled struc- XI.XI. R R == iso-Pr;
iso-Pr;
ture of nonplanar porphyrins, which include meso-substituted XII. XII. RR= = cyclo-Hex;
cyclo-Hex;
molecules with bulky substituents (compounds XI–XIII), as XIII. R=
XIII. R = tret-Bu.
tert-Bu.
well as most nickel complexes (NiP) are characterized by the
deviation from the averaged plane of the meso-carbon atoms (∆Cmeso, Å) (see Table 1).
Table 1 Comparative X-ray analysis data for planar and nonplanar H2P as well as their complexes
with Zn(II) and Ni(II) [19, 65].
Porphyrin Configuration X-ray diffraction data(a)

of ligand
∆Cβ, Å ∆Cmeso, Å
H2(β-Et)8P (I) Pl 0.08[0.06] 0.04[0.03]

H2TPP (II) Pl 0.23[0.18] 0.14[0.45]
H2T(n-Pr)P (III) Pl 0.08 0.02
H2T(iso-Pr)P (XI) Ruf (0.27)[0.27] (0.29)[0.74]
H2T(cyclo-Hex)P (XII) Ruf (0.26)[0.29] (0.06)[0.77]
H2T(tert-Bu)P (XIII) Ruf (0.33) (0.90)
H2TPTBP (IV) Sad (0.77) (0.08)
H2(β-Et)8TPP (V) Sad 1.17(1.09)[1.24] 0.03(0.04)[0.03]
H2(β-Ph)8TPP (VI) Sad 1.28(0.36)[0.96] 0.07(0.01)[0.86]
H2(β-Br)8TPP (VII) Sad 1.26(0.92) 0.32(0.25)
(a) ( ) X-ray analysis data for Zn complexes; [ ] for Ni complexes; ∆C and ∆C
β meso, mean deviations
of C atoms in β- and meso-positions of the initial plane of the macrocycle.
Property
Ì HOMO–LUMO energy gap

Ê H2P redox properties
Ì stability of MP complexes
Coordinating ability of H2P

in electron-donor solvents
Ê Ì
in proton-donor solvents
Acid-base properties of H2P

NH acidity
Ê
N basicity

Ì Relative H2P solvation heats
by weakly solvating solvent
Ê by electron-donor solvent
group
Ƚɪɭɩɩɚ2 2 group 1
Ƚɪɭɩɩɚ 1 Ƚɪɭɩɩɚ
group 3 3 solubility of H2P
Rigidity of H2P macrocycle
Figure 3 Porphyrin structure–property dependences: group 1, planar classical H2P; group 2, non-
planar nonclassical H2P; group 3, planar nonclassical H2P.
The properties related to the reactivity of the coordination core of the H2P molecule
on the whole and the chemical activity of NH bonds in particular are especially dependent
on the peculiarities of the macrocycle geometric structure [58]. Herewith, both porphyrins
with the rigid π-macrocycle (compounds VIII–X) and strongly nonplanar compounds
IV–VII acquire more pronounced acidic properties [35, 61–62], and in a some cases be-
come capable of interaction with molecules of electron-donor solvents [30, 32, 55], which
is exhibited both in their solvation characteristics [32,63–64] and in the change of the re-
activity of these ligands in the complexation reaction (1) with salts of d metals in elec-
tron-donor media [36, 52, 55] as compared with predominantly planar porphyrins proper
(compounds I–III). As the result, there arises a nonlinear dependence of the change of some
properties of H2P on the change of structure of the molecules (Fig. 3).
–2Solv
H2P + MX2(Solv)n–2 [H2P … MX2(Solv)n–4]# MP + 2HX + (n–4)Solv. (1)
For porphyrins with nonplanar structure, an important role in activation of NH bonds

is played by the type of nonplanar conformation of the ligand (see Section 1.4).
The rise of rigidity of porphyrins with the advance of their benzo- (compound X) and
aza-substitution (compounds VIII–IX) [35] leads not only to the flattening of the ligand,
but also to its more pronounced π-electron deficiency. Therefore, the regular consequence
of this is an enhancement of the acidic properties of NH groupings of molecules. Similar
changes of reactivity are also observed in simpler five- or six-membered nitrogen-contain-
ing heterocycles with two or more heteroatoms or atomic groupings. Thus, in the sequence
of compounds pyrrole – imidazole – 1,2,3-triazole – tetrazole the NH acidity in an aqueous
solution increases 12.5-fold [66–67] with the rise of the intensity of π-electron deficiency
of the aromatic heterocycle (Table 2). In a DMSO medium, the acidity of the heterocycles
is 5–6 orders of magnitude lower, as in this solvent there is a problem with the solvation of
anion [68].
The causes of the increase of the acidic properties and, in particular, of the chemical
activity of NH bonds in nonplanar dodeca-substituted porphyrins remain a topical subject
of studies up to the present. As a rule, inclusion of acid-base centres into a unified system
of conjugation leads to the symbateness in the change of the chemical activity of these cen-
tres [69]. It is observed as in the case of five-membered aromatic heterocycles, when an in-
crease of acidity of NH groupings occurs against the background of a decrease of the main
properties of nitrogen tertiary atoms (see the sequence pyrazole – triazole – tetrazole in
Table 2). A similar situation is also observed in the case of complex macroheterocyclic
Table 2 NH acidity (pKa) and N basicity (pKBH+) of five-membered nitrogen-containing hetero-
cycles in DMSO and H2O [66–67].
N N N N
Heterocycle N N N
N N N N N
H H H H H
Pyrrole Pyrazole Imidazole 1,2,3-Triazole Tetrazole
pKa298(DMSO) 23.3 20.4 18.9 – –

pKa298(H2O) 17.5 14.2 14.2 9.3 4.9
pKBH+298 – 2.5 – 1.2 –2.68(a)
(a) measured in sulfuric acid
compounds, in particular, porphyrins with predominantly planar structure (I–III). Thus, in-
troduction of one electron-acceptor chlorine atom each into each of the meso-phenyl rings
of the H2TPP molecule (II), even into meta-positions, increases the strength of this NH acid
in the elimination of the first proton ((pKa1, equation (2)) almost 1.5-fold in a DMSO me-
dium. Simultaneously, the main properties (measured in acetonitrile) weaken in the total
process of elimination of two protons from porphyrin dication (pK3,4, H4P2+ ' 2H+ +
H2P) by two orders [61, 69]. Even the fact that the acidity and basicity of compounds was
assessed in different solvents does not disturb the general pattern of acid-base interaction
(equations (2)–(5)).
KK11
H2P(solv) H+(solv) + HP –(solv) , (2)
KK21
HP –(solv) H+(solv) + P2–(solv) , (3)
KK31
H4P 2+(solv) H+(solv) + H3P+(solv) , (4)
KK41
H3P +(solv) H+(solv) + H2P(solv) . (5)
Interference of spatial factors related to the change of the geometric structure of the
π-macrocycle very often results in the simultaneous increase of NH-acidic and N-basic
properties of nonplanar porphyrins. For instance, already in passing from unsubstituted por-
phin (H2P) to slightly more nonplanar H2TPP (II) both the rise of acidity and basicity of the
molecule are observed simultaneously (Table 3). This phenomenon acquires a much more
pronounced character in strongly nonplanar dodeca-substituted porphyrins IV–VII. Takeda
and Sato [29] were among the first to explain the simultaneous enhancement of both acidity
Table 3 Constants of acid-base interaction of porphyrins [61].

Cl
Cl
Porphyrin NH N NH N NH N
N HN N HN
N HN
Cl
Cl
H2P H2TPP H2T(m-Cl)PP

298
pKa1 (DMSO) 22.35 21.15 19.82
298
pK3,4 (AN) 15.35 19.80 17.80
and basicity of these molecules as the result of spatial deshielding and increased accessibil-
ity of atoms and chemical bonds of the coordination core of the conformationally mobile
macrocyclic ligand for chemical reagents. As it became clear later, this explanation is not
sufficient, as far from any disturbance of the planar structure of the H2P molecule chemi-
cally activates NH bonds in it (see Section 3). On the contrary, according to the classical
polarization views, a greater conformational mobility of the molecule should lead to a wors-
ening of the conditions of the distribution of π-electrons on the conjugated system and,
therefore, its localization on individual atoms and bonds, in particular, on coordination core
atoms. This conclusion is consistent with the increase of the basicity of tertiary atoms of
nitrogen in the molecule of strongly nonplanar H2P ligand; however, it does not explain but,
vice versa, contradicts the enhancement of NH acidity. What is more, 1H NMR studies of
a number of nonplanar porphyrins, whose conformations in the solid phase can be charac-
terized as ruffled [70], have shown that rather significant extraplanar distortions of these
molecules lead to a mere 5% decrease of the π-electron ring current. Herewith, the dou-
ble-dipole model was used, which proved itself to be good in the assessment of ring currents
of planar porphyrins, including chlorophyll ligand molecules [9, 10, 71–72]. Apparently,
an insignificant decrease of the ring current is a specific feature of nonplanar porphyrins.
Its presence makes it possible to assume that π-chromophores of such nonplanar molecules
shall be as readily polarized under the influence of the electronic effects of substituents,
light quanta and chemical reagents.
Table 4 Dipole moments of nonplanar porphyrins [22].
Porphyrin H2TPP cis-H2(β-Et)4TPP H2(β-Et)6TPP H2(β-Et)8TPP

(II) (XIV) (XV) (V)
∆Cβ, Å 0.23 0.76 0.95 1.17

µ (D) 0 1.36 1.89 1.18
Indeed, a recent work [22] has found that dodeca-substituted porphyrins with an in-
creased chemical activity of NH bonds possess, despite the highly symmetric structure,
rather significant dipole moments (µ), while in planar H2P pertaining to a similar structural
group this value is equal to zero (Table 4).
Thus, practically planar tetraphenylporphin (II) has the
zero dipole moment, whereas the symmetric octa-
β-alkyl substitution of this molecule, leading to the non-
R planar structure of the π-system in β-octaethyltetraphe-
R nylporphin {V, H2(β-Et)8TPP}, increases this
NH N characteristic of H2P up to 1.2 D. In the case of porphy-
rins (XIV–XV), which are more planar as compared
N HN with compound V but are additionally polarized due to
R R1 the asymmetric substitution, the dipole moments are
R R1 even higher.
The most nonplanar from asymmetrically substituted
H2P (compound XV) possesses the maximal dipole mo-
XIV. R = Et, R1 = H; ment (1.89 D). An unusual fact is that the dipole mo-
XV. R = R1 = Et; ment of saddle-distorted ligands is directed
orthogonally or almost orthogonally to the plane of H2P
(see Fig. 4).
Ghosh et al. [23] acknowledge that the entire body of changes in the properties of H2P
molecules in passing from the planar structure of the macrocycle to a strongly nonplanar
conformation can not be explained by only the difference in the geometry of the molecule.
Based on the above said, it can be concluded that the cause of changes in the physicochem-
ical properties and reactivity of nonplanar porphyrin ligands, in particular, those caused by
an increased chemical activity of coordination core’s NH bonds, is internal polarization of
nonplanar molecules. It also determines many of their photophysical properties [22, 49–50]
and reactivity [29–30, 52].
Polarization is a consequence of conformational conversions of H2P nonplanar mole-
cules. Such structural changes are associated with the asymmetric change of bond lengths
and valence angles, and lead to the redistribution of π-electronic density in it. Thus, they
are the cause of secondary electronic changes in the molecule, which affect the reactivity
of its reaction centres (N4H2, MN4 etc.).
Polarization of H2P molecules under the influence of the electronic effects of substit-
uents has an additional positive effect on the activation of NH bonds. In the case of highly
aromatic molecules, such as tetrabenzoporphyrins (X) and porphyrazines (VIII–IX), the
Figure 4 Structural causes of the formation of the orthogonal dipole moment in saddle-distorted
porphyrin ligands by the example of H2(β-Et)8TPP (V) [22, 33].
planar structure creates the most favourable conditions for their polarization by the light
wave, which is exhibited in the change of the photophysical characteristics of the com-
pounds. Besides, this is favourable for internal polarization of H2P by means of substituents
of various electronic nature and for the emergence of the push-pull effect at the time when
simultaneously donor and acceptor atomic molecular functions (compound XVI) occur
[40–43, 71].
The influence of the
push-pull effect proper on
the reactivity of porphyrin
coordination cores and on
NH bonds practically has NH N
not been studied. Only the (M e)2N C C C C NO
optical properties were in- N HN
vestigated, including the
nonlinear optics properties,
as well as such parameters
of “push-pull” porphyrins XVI
as polarizability β and di-
pole moment µ [40–43]. The highest polarizabilities of the order of 5000·10 –30 esu and
dipole moments of more than 10 D were found in “push-pull” porphyrins carrying elec-
tron-donor (e.g., NMe2) and electron-acceptor (e.g., NO2) groups in phenyl rings, which
are linked with meso-positions of the macroring via an ethynyl spacer [41]. A later work
[43] showed the most efficient among π-linking spacers (–CSC–, –N=N–, –CH=CH–,
–CH=N–) to be exactly the ethynyl spacer.
The consequences from introducing substituents of any electronic nature – donors or
acceptors – into the H2P molecule (as a rule, with a chemically low active NH bond) have
been studied well [1–3, 9–11]. A change of reactivity of the coordination core of such mol-
ecules is observed in complexation reactions of H2P with solvatosalts of d metals
[MX2(Solv)n–2] (1) and acid-base interaction ((2)–(5)), as well as of dissociation (6) and
metal exchange (7) in MP porphyrin complexes. Peripheral substituents of the same elec-
tronic nature in H2P molecules affect different processes differently. The cause of this is
oooo
MP + nH·Solv+ + nX – H4P2+ 2X – + MX2(Solv)n + n–4HX, (6)
MP + M′Xm(Solv)p–m M′P + MXm(Solv)p–m (7)
the difference of the mechanisms of the above listed reactions and the different degree of
involvement of NH bonds in these mechanisms. Thus, the complexation reaction (1) has a
strong dependence on the donor-acceptor properties of the solvent, in which it is run, and
for H2P with a chemically inactive NH bond (e.g., para-substituted derivatives
{H2(p-R)4TPP} of tetraphenylporphin II) is accelerated by electron-donor substituents in
phenyl rings, and is slowed down by electron-acceptor substituents [1–3, 73]. The rate of
reaction (1) in this case is determined by the strength of reacting solvatosalts of the com-
position [MX2(Solv)n–2] [74–78] and is, therefore, maximal in carboxylic acids (AcOH,
C2H5COOH) and other weakly coordinating solvents, such as esters, ketones, nitriles and
alcohols [5]. Reaction (1) of porphyrins with a chemically active NH bond is, in contrast,
accelerated by electron acceptors, which are simultaneously coordination-core activators,
and slowed down by electron donors [5, 31, 32, 35]. The highest catalytic effect for reaction
(1) is achieved in a medium of polar electron-donor solvents, also activating NH bonds in
the H2P molecule. A characteristic example are porphyrazines (VIII–IX, XVII–XVIII)
substituted in β-positions by electron-acceptor halogen atoms [35, 54]. From the data of Ta-
ble 5, it follows that the rate constants (kv) of reaction (1) of NH-active macrocycles with
ZnAc2 increase with the rise of electronegativity of β-substituents, as well as at an increase
Table 5 Kinetic parameters of the formation of zinc complexes with tetraazaporphin (VIII) and its
halogen derivatives (IX, XVII–XVIIII) in AcOH–Py mixtures, CH2P = 2·10 –6 mol/l, CZnAc2 =
3.3·10–4 mol/l [54].
Porphyrin Solvent kv298 , E, ∆S #,

s–1· mol–1 kJ · mol –1 J · mol–1 · K–1
H2TAP (VIII) AcOH 0.0087 50 –190

H2(β-Br)4TAP (IX) 0.0070 55 –185
H2TAP (VIII) AcOH – Py = 99–1 0.0054 52 –180
H2(β-Br)4TAP (IX) 0.0020 78 –110
H2(β-Cl)4TAP (XVII) 0.0011 77 –128
H2(β-F)4TAP (XVI) 0.00074 72 –146
H2TAP (VIII) AcOH– Py = 3– 2 0.191 62 –106
H2(β-Br)4TAP (IX) 0.197 71 –120
H2(β-Cl)4TAP (XVII) 0.225 54 –161
H2(β-F)4TAP (XVI) 0.270 58 –147
H2TAP (VIII) Py 5.79 44 –161
H2(β-Br)4TAP (IX) very fast
H2(β-Cl)4TAP (XVII) very fast
H2(β-F)4TAP (XVI) very fast
of concentration of the electron-donor component of the solution (Py). Thus, kv in reaction

(1) of H2TAP in passing from pure acetic acid to pure pyridine increases more than 2.5-fold,
and in the sequence of β-substituents H < Br < Cl < F from 1.5- to 7-fold [54]. The dou-
ble-action effect of electron acceptor and solvent is exhibited both in the reactivity of the
coordination core of rigid [34–37, 53–55] and strongly nonplanar [8, 19–20, 22–23,
29–32, 56–58,71] porphyrins with a chemically active NH bond.
Dissociative processes in porphyrin complexes, such as dis-
sociation of MP (6) or metal exchange (7), initiated by the at-
X
tack of electrophilic reagent (H·Solv+, H3O+, HX, M2+) are,
N
X as a rule, accelerated by electron-donor substituents, because
NH N the latter increase the chemical affinity of the reaction centres
N N (nitrogen atoms) in reactions of the type of (6) and (7). Thus,
for instance, when only the σ-effect of substituents affects the
N HN
MP dissociation reaction in a solvent–H2SO4 medium, as is
X the case in metallotetraphenylporphyrins {MT(R)PP}, it is
N
X accelerated, irrespective of the nature of metal (Fig. 5) {co-
valent σ-complexes of Zn(II) and Fe(III), σπfwd, Mn(III)
XVII. X = F: complex; σπinv, Cu(II), Ni(II), Pd(II) complexes; predomi-
XVII. X = F; XVIII. X = Cl
XVIII. X = Cl nantly ionic Cd(II) complex}, in the following sequence of
N N
-ɨɛɪ
σ πinv .
-ɨɛɪ
σ
πinv .
M -ɨɛɪ.
πinv
σ
-ɨɛɪ.
πinv
σ
N N
Figure 5 Electronic effects of coordination in molecules of metalloporphyrins [4–5].
the functional groups located in the phenyl rings of the macrocycles: –NO2, –NH3+, –Cl,
–Br, –COOH < H < –CH3, –OH, –OCH3 [79–80]. If a substituent enters into direct con-
tact with the π-macroring when being in β- or meso-positions of the MP molecule, reaction
(6) proceeds with account of the electronic effects of substituents ±I and ±C, as well as the
electronic effects of coordination – σ-donor–acceptor interaction (N → M), forward dative
π-bond (πfwd, N → M) and inverse dative π-bond (πinv, N ← M) (Fig. 5) [1, 4–5]. A number
of dissociation rates in such MP can change and even reverse depending on the combination
of the above listed effects [80].
Studies of the effect of basicity of substituted meso-tetraphenylporphin (II) (Table 6)
on the rates of metal exchange reaction (7) in the {CdP/Zn(ClO4)2/Py/0.5M H2O} system
within the temperature range of 288–310 K has shown that electron-donor groups in the
phenyl rings of H2TPP accelerate the metal substitution process [81–82].
Thus, the true reaction rate (kv298, l·mol–1 ·s–1) in passing from p-CN to p-OCH3 de-
rivative increases from 1.3 up to 10.2. The metal exchange rates in the considered cases are
600–6000 times higher than the rate of complexation of zinc perchlorate with respective
ligands (Table 6).
Table 6 Kinetic parameters for the metal exchange reaction of {CdP/Zn(ClO4)2 /Py/0.5M H2O} of
meso-substituted CdTPP [81].
Complex kexch /kcompl k298, ∆H #, ∆S #,

l·mol–1 ·s–1 kJ·mol –1 J · mol–1
CdT(p-CH3)PP 1640 8.3 70.2 8.7

CdT(o-CH3)PP 6700 8.5 61.0 –21.7
CdT(p-OCH3)PP 1000 10.2 58.1 –30.5
CdT(p-CN)PP 1400 1.2 54.7 –58.5
CdT(p-H)PP 1530 5.7 48.1 –68.5
CdT(p-Cl)PP 600 1.3 45.1 –90.3
The rates of acid-base interaction processes involving the coordination core of the H2P
molecule (equations (2)–(5)) react to the electronic nature of substituents in the porphyrin
macrocycle in a way traditional for this type of reactions, i.e., the basicity increases and the
acidity goes down at the drop of π-electronic density in the macrocycle [61].
Even the introduction of polarizing substituents into H2P molecules with inactive NH
bonds (compound II) strongly changes the physicochemical properties and reactivity of the
compounds. Thus, the substitution of one of β-positions in the H2TPP molecule (III) stabi-
lizes two trans-tautomers [33, 72], increases the dipole moment of the molecule up to
approximately 7 D [33], due to the charge-transfer (CT) interactions cardinally changes the
photophysical pattern of the process in its main S0 and excited (S1, T1) states [83].
1.1.2 NH activation in the course of porphyrin–solution component interaction and

porphyrin–solid phase interaction
The environment of the H2P macrocyclic ligand or its complexes in solid, liquid or gas
phase plays an important and sometimes determining role for the successful execution of
their useful functions [8, 68, 84]. Namely the rapid and significant response in the electronic
structure and geometry, in the properties of compounds to the change of the nature of the
medium is one of the conditions to recognize it as a biologically active or technically useful
substance, e.g., in molecular recognition processes. For this reason, studies of the effect of
the phase state of a substance and of the properties of the medium on the progress of various
physicochemical processes involving biologically active substances or their models are still
topical.
Porphyrins belong to just the type of compounds, whose properties strongly depend
on the change of environment of the molecules. This occurs, in particular, due to the com-
plexity and multifunctionality of the coordination core of the H2P molecule (MP). As the
greater part of chemical reactions proceed namely in the liquid phase, interactions of the
“H2P·Solv” and “H2P·solution component” type have been studied better than solid-phase
interactions [11, 64, 85].
Herewith, the most investigated interactions are those involving H2P with a chemically
inactive NH bond. Their behaviour in solution is readily predictable, and changes of pho-
tophysical and coordination properties, solvation and stability of compounds to the action
of light, temperature or chemical reagents obey the “classical” regularities, well known in
porphyrin chemistry already by early 1980s [1, 9–10]. These compounds in the absence of
active functional groups at the periphery of molecules are little soluble in organic solvents,
both in weakly solvating (hexane, benzene) or weakly proton-donor ones (chloroform, ni-
tromethane) and in electron-donor ones (e.g., pyridine, DMSO, acetonitrile). Their solubil-
ity under standard conditions changes within 1·10–5 –1·10 –3 mol/l [86]. The low solubility
of H2P (Table 7) is predominantly due to the universal character of solvation of their mol-
ecules as the result of strong shielding by means of the macrocyclic effect of coordina-
tion-core atoms – in this case the only centres of specific solvation [6–8, 64]. This type of
Table 7 Solubility of porphyrins in organic solvents [86].
Porphyrin S298 ·105, mol/l
Benzene Ethanol
H2TAP (IX) 3.16(a) 1.9

H2TATBP (XXII) 39(a) (49)(b) 0.3
H2TBP (X) 0.83 (560)(b) –
H(β-Et)8P (I) 290 5.0
H2TPP (II) 590 1.8
H(N-Me)TPP (XX) 1980 5.3
H(N-Ph)TPP (XXI) 780 2.8
Measured in chlorobenzene(a) and pyridine(b).

interaction assumes a weak sensitivity of compounds to the nature of the solvent, e.g., in-
significant changes in their photophysical properties [48, 60, 71], high chemical and ther-
mal stability of most H2P in solutions [1–5, 64]. At the same time, the nature of solvent
plays a significant role in the change of reactivity of N4H2 coordination core, in particular,
in reactions of acid-base interaction [61] and complexation [75].
Owing to the polarization of molecules of NH-active porphyrins [22, 71], their reac-
tivity in solutions of electron-donor solvents significantly increases; this applies not only
to nonplanar H2P possessing the intracyclic reaction centres more accessible for reagents
[30, 32, 52], but also to planar porphyrins with rigid π-chromophore [53–55]. As we have
already mentioned, the rise of aromaticity of the H2P molecule assumes not only the shield-
ing of intracyclic reaction centres at the cost of the MCE structural component, but also the
redistribution of the π-electronic density by means of the MCE electronic component,
which entails the pronounced chemical activity of particular bonds [7–8]. In a number of
chemical processes, the latter of the two factors prevails. At the same time, the solvation
characteristics of planar and nonplanar H2P with a chemically active NH bond differ no-
ticeably. Nonplanar H2P are well soluble in organic solvents irrespective of the type of
bond, as, e.g., N-substituted porphyrins XIX–XXI with a chemically inert NH bond [85],
whereas planar ones are still little soluble, especially in weakly proton-donor media (see
Table 7) [86].
Thus, the solubility of predominantly planar compounds I–II with a chemically inac-
tive NH bond in benzene decreases by 1–3 orders with the rise in the macrocycle rigidity
(compounds X, IX, XXII). At the same time, for planar NH-active compounds the value of
S298 is always higher in electron-donor solvents (e.g., in pyridine) as compared with uni-
versally solvating ones (Table 7) owing to the additional specific interaction of the Solv
molecule with NH bonds of the coordination core of the H2P molecule. In a medium of
weakly proton-donor solvents (EtOH), porphyrins, in contrast, are very weakly soluble, in-
cluding at the expense of depolarization of NH bonds in them. Indeed, in some cases con-
sidered the solvent can regulate the extent of chemical activity of these bonds. Partially
delocalized NH bonds of these compounds experience an additional activation (polariza-
tion) in a medium of electron-donor solvents [32] and, on the contrary, are completely lo-
calized already in a weakly proton-donor medium (carboxylic acids) [31]. These structural
changes are reflected in a characteristic jumpwise change of the complexing properties of
NH-active porphyrins in solvents of the types considered, and are also seen in the EAS [30,
55] (see Sections 1.2.1 and 1.3).
Consider schematically the mechanism of action of electron-donor (Py) and elec-
tron-acceptor (AcOH) solvents on the state of the coordination core (N4H2) of H2P mole-
cules of various structural groups: porphyrins proper (classical) and their rigid, or,
conversely, strongly nonplanar nonclassical derivatives (Fig. 6) [31].
In molecules of classical H2P (compounds I–II) [28] the macrocycle is almost planar
[13, 65], so the access of solvent molecules to –N= and –NH– solvation sites is hindered.
Besides, NH bonds in such molecules are polarized very weakly. Both of these factors lead
to a situation, when the interaction of NH groupings with molecules of electron-donor Py
is almost absent in the main state. Thus, a change of enthalpy in the transfer of H2TPP (II)
from inert solvent benzene to electron-donor Py, which characterizes the change of relative
solvation, is very small (∆trH0 = –1.9 kJ/mol) [7, 64]. In this case the NH bond remains
totally localized (is not activated), and the reaction proceeds with difficulty.
Due to the low basicity, as well as owing to structural causes, the molecule of AcOH
enters into only a very weak acid-base interaction with tertiary –N= atoms of the classical
meso
meso meso
meso
Figure 6 State of the coordination centre of porphyrin molecules with a chemically inactive group
(a) and a chemically active NH bond (b) in proton-donor (AcOH) and electron-donor (Py) media; b1,
nonplanar H2P; b2, planar H2P; , transfer of π-electronic plane in the macrocycle [31, 39].
H2P molecule. The H associate formed (Fig. 6, a) is, apparently, characterized by a low de-
gree of proton transfer, because it is not registered in the EAS. Nevertheless, protons in this
associate act as electron acceptors, decreasing the electronic density in the macrocycle and,
thus, promoting the rather weak polarization of the NH bond, e.g., in reaction (1).
This mechanism works only for predominantly planar porphyrins with a moderate
rigidity of the macroring. In the case when the macrocycle acquires a nonplanar conforma-
tion under the influence of multiple substitution at the periphery of the molecule or in any
other way [8, 16–17, 19, 87] (see also Section 1.4), the basicity of its tertiary nitrogen atoms
sharply increases, and the action of AcOH leads to partial or total protonation of these N
atoms (equations (4) and (5)) and their blocking as coordination centres in reaction (1)
(Fig. 6, b1).
Recently, it has been found that
R1 R R1 strongly distorted dodeca-substi-
N
tuted H2P, e.g., compounds
R1 R1 V–VII, in their titration in toluene
N-X N NH N solutions by weak organic bases
N
R R N form H complexes (or proton-
N HN
N HN transfer complexes, PTC (equa-
R1 R1 N tions (8)–(9)), which are charac-
terized by a rather profound
R1 R R1
transfer of NH proton to the mol-
XIX. R = H; R1 = Et; X = Me XXI. ecule of electron-donor solvent
XX. R = Ph; R1 = H; X = Me and, for this reason, are clearly
XXI. R = X = Ph; R1 = H seen in the EAS [30, 55, 85]. This
fact makes it possible to assert
that the NH bond in them is noticeably polarized. The distortion of the macrocycle increases
the accessibility of this bond for the attack by the Py molecule, and the formation of the H
complex with proton transfer to pyridine additionally polarizes the bond, bringing it closer
to the state characteristic of the transition state of reaction (1). Indeed, reaction (1) with non-
planar H2P proceeds very slowly in AcOH and easily in Py (see Sections 1.3 and 1.4). The
difference in complexation reaction rates in these solvents is up to more than seven orders
of magnitude [8, 51, 57, 85].
Another group of nonclassical H2P – most often these are the aza derivatives of por-
phin [26, 28, 34–37], such as compounds VIII–IX, XVII, XVIII, XXII – is characterized
(Fig. 6, b1) by a high aromaticity of the macrocycles and an enhancement of the macrocyclic
effect (MCE) [6–8]. The MCE structural component [8] contributes to the shielding of –N=
and –NH= centres from the penetration of reagents. This is one of the causes of a decrease
of the main properties of tertiary nitrogen atoms of tetraazaporphin (H2TAP, VIII) and tet-
ra-β-bromotetraazaporphin {H2(β-Br)4TAP, IX} [61]. For this reason, there is almost no
interaction of them with AcOH. The bridge (meso-) aza atoms are involved in only a weak
associative interaction with AcOH molecules, because their unshared electron pairs are in-
volved to a significant degree in the conjugation with the π-system of the macroring [1, 35].
As for the state of the NH bonds, they are completely localized in a solution of AcOH,
which does not contribute to the exchange of NH protons of H2P with the medium.
Simultaneously, intramolecular effects, such as the enhancement of the MCE π-elec-
tronic component [8], evoke a significant polarization of NH bonds in molecules of rigid
H2P. This polarization follows from an increase of their acidity [61], which for azaporphy-
rins in a DMSO medium is 10–12 orders higher as compared with classical H2P. Protona-
tion of NH bonds facilitates the formation of the PTC of rigid macrocycles with molecules
of electron-donor solvents, where these bonds are already partially delocalized [55] (Fig. 6,
b2). Formation and stability of PTC are considered in more detail in Section 1.2.
Apparently, the considerable potential of activating NH bonds and modifying the prop-
erties is in the effect of the solid-phase environment on porphyrins. Unfortunately, these
issues are far from being understood at present, and there are only separate works on the
topic in the literature [44–45, 88–89]. All these publications note the catalytic effect of the
solid phase in H2P or MP formation reactions. Tsukahara and Suzuki [44] studied the com-
plexation ability of H2TPP (II) sorbed on Kieselgel 60HR. They showed that its reaction
with Zn2+ ions sorbed on SiO2 proceeds much faster in a sequence of weakly polar or non-
polar solvents (CCl4 > toluene > 1,2-dichloroethane > acetone) as compared with the reac-
tion in solution. At room temperature, reaction (1) is complete in only 5 min in all solvents
except weakly coordinating acetone at an H2TPP–salt concentration ratio of 1–18/260,
whereas in solution it runs slowly and at increased temperatures [75]. This indicates the de-
localization of NH bonds under the influence of sorbent’s active centres, as well as the spe-
cific adjustment of activated reaction centres of the solvatosalt to the reaction centres of
porphyrin. Another example of solid-phase activation of NH bonds was observed when the
chlorophyll ligand and its derivatives immobilized on water-soluble polymer material
(polyvinyl alcohol) reacted in a glacial acetic acid and its aqueous solutions with Cu(II),
Zn(II) and Co(II) acetates at rates not lower and even higher than in the absence of polymer
carrier [45]. The above mentioned facts may come as a surprise because at first glance the
solid-phase and, moreover, polymer environment strongly hinders sterically the attack of a
reagent, e.g., the solvatosalt in reaction (1) on the coordination core of the H2P molecule.
Letts and Mackay [88] studied the complexation reaction of H2TPP with Cu(II), Mg(II),
Mn(II), Zn(II) and Co(II) ions in a water-emulsion system in the presence of cationic or an-
ionic surfactants, noted both the facts of its inhibition and acceleration by various deter-
gents. In the literature, there are facts of heterogeneous catalysis of not only MP formation
reactions (1) but also of condensation processes to form H2P ligands. Thus, for instance,
Onaka et al. [89] observed a more than twofold increase of the yield of tetra-meso-alkylpor-
phyrins (35–45%) in a system containing porous montmorillonite K10.
1.2 Interaction of organic solvents and porphyrins with delocalized-type bonds

Porphyrins with chemically active NH bonds enter into quite a number of specific interac-
tions involving these bonds. They can be the already mentioned weak acid-base interactions
or tautomeric conversions, as well as their combinations. Depending on a particular process
and type of ligand, such kinds of interactions can lead to absolutely different consequences
– from the destruction of H2P chromophore to the formation of supramolecular structures
on its basis.
1.2.1 Acid-base interactions

Traditionally porphyrins are assigned to compounds with weakly pronounced NH-acidic
properties [1, 5, 9–10, 61]. The cause of the decrease of acidity of these aromatic tetrapy-
rrole compounds even as compared with individual pyrrole is, apparently, the spatial shield-
ing of NH sites by atomic-electronic π-macrocyclic environment (the macrocyclic effect)
[8]. However, due to the activation of NH bonds by means of the above considered struc-
tural polarization factors (the MCE electronic component [8]) they are not capable any more
of not only donating a proton to a strong base, e.g., tetraalkyl ammonium hydroxide
R4NOH, but of interacting by the acid-base type of interaction with molecules of elec-
tron-donor solvents (B) to form specific complexes with proton transfer (PTC) (equations
(8)–(9); Fig. 7, a).
Kk1
H 2P + B k –1
H2P·B, (8)
k21
K
H2P·B + B k –2
H2P·2B. (9)
R R Participation in such kind of processes is an indi-

vidual feature of porphyrins with the delocalized
type of chemical bond.
R N R It should be noted that planar (porphyrazines,
compounds VIII–IX, XVII–VXVIII, XXII–
NH N XXV) and nonplanar (dodeca-substituted, com-
N N pounds IV–VII) porphyrins behave differently in
N HN acid-base interaction reactions with electron-
R N R
donor solvents. First, nonplanar H2P enter into re-
actions of the type of (8)–(9) in an equilibrium
manner [30], whereas for planar porphyrines PTC
formation processes are realized in the kinetic re-
R R
gime [55, 90]. Second, although proton-transfer
XXIII. R = H; complexes are in both cases unstable in media
XXIV. R = Br;
XXV. R = NO2 with low permittivity (ε), the results of this insta-
bility are different. PTC with porphyrazines in
low-polar media are, as a rule, subjected to destruction to break down the macrocycle,
whereas in the case of dodeca-substituted H2P the decrease of permittivity of the medium
only sharply shifts the equilibrium (8) of the formation of the H associate towards the initial
compounds. For azaporphyrins, cases are known when the stability of PTC is high and, for
structurally thermodynamic reasons, their formation inhibits the coordination reaction (1)
λ, nm λ, nm
Figure 7 Change of electronic absorption spectra in the course of formation in the C6H5Cl–DMSO
system (a) and partial destruction in the C6H6 –tert-BuNH2 system (b) of the PTC of β-tetrabromo-
tetraazaporphin (IX) [90].
as compared with media, where the formation of PTC with porphyrins is not registered
spectrally, i.e., the extent of interaction of protons of NH groups with molecules of the sol-
vent is lower. No such cases were observed for strongly nonplanar H2P.
Interactions of planar derivatives of porphin, in particular, porphyrazines, have been
studied in the literature in greatest detail [55, 90–99]. Proton-transfer complexes were first
spectrally (EAS) observed in 1961 by Whalley in pyridine solutions of H2(β-Ph)8TAP
(XXIII) and called pyridine salts [91], which is not quite correct as the PTC is not a salt but
represents a product of incomplete acid-base interaction [92]. The scheme of complete
transfer of proton from acid HA to base B has the form:
K1 K1 K1 K1
HA + B AH…B Aδ– …H…Bδ+ A –…HB+
K1
A – ·HB+ A – + HB+, (10)
where AH…B is a molecular complex with the H bond, Aδ– …H…Bδ+ is an intermediate-
type complex with the delocalized H bond, A –…HB+ is an H-bound ion pair, A – ·HB+ is a
contact ion pair, A – + HB+ are complete proton-transfer products. The H2P with partial pro-
ton-transfer to the base molecule (B or Solv) represents, depending on the strength of acid
and base, permittivity and solvating power of the medium either an H associate (AH…B)
or an ion pair (A –…HB+) or else an intermediate complex with the delocalized position of
proton (Aδ– …H…Bδ+) (Fig. 8) [90]. Formation of such kinds of products is difficult even
in solution and is possible only in the case of weak acids and bases with spatially shielded
reaction centres [93].
The kinetics and mechanism of PTC formation with porphyrazines (VIII–IX,
XXII–XXV) have been studied in detail in a series of works [55, 90, 94–99].
Based on the spectral and kinetic data, a probable structure of the PTC of the type of
Figure 8 Assumed structures of the porphyrin PTC with a weak base molecule B: (a) ionized, (b)
H-bound [35, 55, 90].
H2P·2B (Fig. 8) has been proposed. It has also been shown that the formation of kinetically
stable proton-transfer complexes is observed only if NH-acidic properties of H2P are suf-
ficiently strongly pronounced, the electron-donor ability of the base is moderate and the per-
mittivity of the medium is high, as, e.g., in DMSO or inert solvent–DMSO mixtures as well
as in a DMSO–Py medium [90] (Table 8). Otherwise, e.g., in most media with low permit-
tivity based on aliphatic amines {Et2NH, n-BuNH2, tert-BuNH2PhCH2NH2, Et3N,
n-Bu3N, Pip}, PTC are subjected to gradual destruction to colourless products (Table 9, Fig.
7, b). Both the PTC formation and destruction rates are controlled by steric factors. The
more bulky the amine molecule and the more shielded its reaction centre, the more complex
the proceeding formation and destruction processes of PTC are (Fig. 9, Table 9). The ob-
served phenomena are well described within the framework of the PTC formation and dis-
sociation mechanisms [55, 90].
Table 8 Kinetic parameters of acid-base interaction of H2TAP derivatives with DMSO in C6H5Cl
(CH2TAP = 0.5 · 105 mol/l; C 0DMSO = 3.88 mol/l).
Porphyrin k298 ·105, E, ∆S #,

l·mol–1 ·s–1 kJ · mol –1 J · mol–1 · K–1
H2(β-Cl)4TAP (XVIII) 3.47 24 –250

H2(β-Br)4TAP (IX) 5.30 26 –237
Table 9 Kinetic parameters of acid-base interaction of H2TAP derivatives with N bases in benzene
{CH2TAP = 0.4–0.5·10–5 mol/l}.
Porphyrin N base k298 ·102, E, ∆S #,

2 ·mol2 ·s–1
l kJ · mol –1 J · mol–1 · K–1
H2(β-Br)4TAP BzNH2 0.78±0.04 29±2 –226±5

(IX) n-BuNH2 6.6±0.3 11±2 –270±6
tert-BuNH2 0.076 18 –283
Et2NH 4.0±0.2 15±2 –261±5
H2(β-Cl)4TAP BzNH2 0.65±0.03 31±3 –223±10
(XVIII) n-BuNH2 3.8±0.1 23±2 –235±7
tert-BuNH2 0.09 28 –251
Et2NH 1.10±0.05 30±3 –218±10
Figure 9 Dependence of log k of the destruction of H2(β-Br)4TAP (IX, filled dots) and
H2(β-Cl)4TAP (XVII, empty dots) in benzene in the presence of basic nitrogens at 298 K.
eff
DMSO
Figure 10 Dependence of log keff on CDMSO for reaction (1) of formation of Mg(β-,4- NO2Ph)8TAP
in benzene at CMgAc2 = 1.8·10 –4 mol/l; T = 308 K (1), 318 K (2), 328 K (3).
PTC formation processes are facilitated in the following sequence of compounds:

H2TAP (VIII) < H2(β-Ph)8TAP (XXIII) < H2(β-,4-BrPh)8TAP (XXIV) < H2(β-,4-
NO2Ph)8TAP (XXV) < H2(β-Br)4TAP (IX) < H2(β-Cl)4TAP (XVIII), which is determined
by the enhancement of the proton-donor properties of the ligands [35–36, 55, 90]. Thus,
while compound IX enters into acid-base interaction with molecules of bases already under
normal conditions, tetraazaporphin (VIII) and β-octaphenyltetraazaporphin (XXII) form
PTC only in prolonged boiling in a solution of base B.
Formation of PTC is a process of chemical activation of NH bonds in the H2P molecule
by organic solvents.
λ, nm
Figure 11 Change of electronic absorption spectra for H2(β-Br)8TPP (VII) in toluene in the
presence of piperidine, CH2P = 4.23·10 –4 mol/l; Cpip, mol/l: 0 (1), 0.008 (2), 0.016 (3), 0.025 (4),
0.033 (5), 0.041 (6), 0.058 (7), 0.083 (8), 0.083 (9), 0.166 (10).
lg CHP- / CH2P
Figure 12 Graphic determination of the number of base B molecules participating in the equilib-
rium with H2(β-Br)8TPP (VII) in toluene: 1, Pip; 2, DMF; 3, Py; 4, DMSO; n = tan α ≈ 1.
Indeed, in a number of cases H2P molecules activated in this way enter more readily
into complexation reactions (1).
At the same time, it should be noted that the complexing ability of PTC in reaction (1)
is determined by its kinetic stability and can be both strongly increased and decreased with
the change of the concentration of the base in the same reacting system (Fig. 10).
The ability of strongly nonplanar dodeca-substituted porphyrins to interact with elec-
tron-donor solvents was found comparatively recently [29–30]. It was revealed that the
substitution of a weakly solvating (inert) solvent (benzene, toluene) by an electron-donor
solvent completely changes the structure and number of bands in the electronic spectrum
of the molecule (Fig. 11) [30]. The fact that the number of bands decreases, and the EAS
starts to resemble more the spectrum of a metal complex, indicates the formation of struc-
tures with a higher symmetry (D2h → D4h). Similar spectral changes were also observed in
the interaction of porphyrazines with bases [55, 90]. In contrast with porphyrazines, PTC
in dodeca-substituted porphyrins are formed in an equilibrium manner. Karmanova et al.
[30] calculated the stability constants of forming delocalized acid-base forms from the re-
sults of titration of solutions of H2(β-Br)8TAP (VII). The stability of PTC proved to in-
crease with the rise of polarity of the medium, even if the electron-donor properties of the
base weaken: Py < DMF < DMSO < Pip. In contrast with solutions of porphyrazines, the
breakdown of PTC proceeded in DMSO without the destruction of ligands at the addition
of N bases. Besides, it was found that in the course of titration of compound VII by the base
only a PTC of the type of H2P·B is formed (equation (8)). In particular, this follows from
the value of the slope of the dependence log CSolv = f (log CHP– /CH2P) (see Fig. 12).
Table 10 Stability constants (Kst) of proton-transfer complexes (PTC) and thermodynamic
parameters of acid-base interaction of H2 (β-Br)8TPP) (VII) with organic solvents [30].
Base solvent Characteristic of solvent Kst298 ∆H, ∆S #,

kJ · mol –1 J · mol–1 · K–1
DN ε
Py 33.1 12.3 0.025±0.001 –18±1.1 –90±4

DMF 26.6 36.7 0.87±0.05 –11±0.5 –36±2
DMSO 29.8 46.7 2.87±0.17 –17±1.6 – 49±4
Pip – 4.28 6.3±0.4 –35±0.7 –102±3
1.2.2 Tautomeric processes

For porphyrins proper, not containing additional active functional fragments, e.g., meso-
tetraphenylporphin (H2TPP, II), tautomeric conversions are little characteristic. For them,
as for any other H2P, the coordination core of the molecules of which contains pyrrole and
pyrrolenine atoms of nitrogen, only the process of N–NH tautomerization is known (Fig.
13), whose conditions and mechanisms are considered in [35, 48, 60, 72,100–109]. This
type of tautomerization weakly depends on the nature of solvent, and the energy of its ac-
tivation is about 12 kcal/mol and little depends on the phase state of H2P. Quick-time
N–NH exchange by protons is reduced to the conversion of two equivalent or practically
equivalent trans-tautomers in each other. The synchronous (I) and asynchronous (II) mech-
anisms of ND–NH tautomerization is possible [100] (Fig. 13). A number of quantum chem-
ical [48, 108–109, 101, 104], 1H NMR spectral [60, 72, 102–103] and photophysical [48,
100, 105–106] data are in favour of the asynchronous mechanism,. The two-step process
II with thermally activated quantum mechanical tunnelling through the activation barrier is
realized at T = 200–300 K [101] and in the case of predominantly planar, symmetrically
substituted H2P is characterized by the rate constant ≈5·104 s –1 in CDCl3 [102]. In the
course of process II the activation energy of the system is required only to achieve the
cis-tautomer, which then is tunnelled by route II [107]. At temperatures ≈77 K the synchro-
nous mechanism of tautomerization predominates, however, the rate of the process is much
lower (≈10 –6 s –1 at 4.2 K) [101, 107], and the calculated activation barrier of the process
is ≈18.5 kcal/mol [104].
The type II process (Fig. 13) can also run with participation of excited states of the
H2P molecule as photoisomerization [105, 108]. A number of indirect proofs of its running
with participation of the triplet T1 state have been found. Thus, Shushkevich et al. [106],
by the example of tetrabenzoporphin (compound X), assessed the rate constant of NH re-
arrangement of the H2P molecule in triplet state to be ≈ 10 –1 s –1 (at 77 K).
Introduction of an efficient electron-acceptor [107] or “heavy” [102] substituent into
a pyrrole ring or meso-position of the macrocycle stabilizes NH tautomers. For instance,
meso-substitution of the β-octaethylporphin (I) molecule by a bulky 2-(4-nitrobenzyl)-1-
NH HN
N N
II N HN
NH N
I NH N
N HN
Figure 13 Synchronous (I) and asynchronous (II) routes of N–NH tautomerization of porphyrins.
naphthyl fragment decreases the rate of the process to 0.004·104 s –1, i.e., by about three
orders of magnitude. NH tautomerization does not change the properties of the H2P mole-
cule significantly, because the rearrangement of the π-conjugation contour in the conver-
sion of tautomers leads neither to the change of symmetry nor to the activation of the already
available reaction centres or the emergence of new reaction centres. .
Sophistication of the H2P structure can enrich the reactivity to a significant degree and
often provides a possibility for the existence of the molecule in various molecular forms
depending on the nature of the medium, on the presence of some or other reagents in solu-
tion. These manifestations determine the whole range of practically useful properties of the
molecules [110]. One of the examples of such compounds are H2P carrying active (one or
several) hydroxy or amine groups in meso-positions. In solutions, these compounds enter
into tautomeric conversions of, respectively, keto-enol or amine-imine type. As a rule, be-
ing in a nonionized medium, these compounds exist in keto or imine form, which is reliably
confirmed by spectral methods and follows from the data on their reactivity. Keto-enol
equilibria of oxoflorins have been studied the most [111–112]. Existing in nonionized me-
dia in a keto form, oxoflorins (XXVI) are subjected to tautomerization during the action of
strong acids (XXVIb), as well as in the course of coordination of these nonaromatic ligands
by double-charged metal ions (XXVIc), and form meso-hydroxyporphyrins (enols) (Fig.
14) possessing all the features of aromatic molecules. For this reason, oxoflorins are stron-
ger acids and bases as compared with H2P, they also enter more readily into complexation
reactions [111]. The conversion mechanisms of oxoflorons (Fig. 14) have not been studied;
however, it is evident that the process in this case involves chemically activated NH bonds.
Chlorophyll and its structural analogues are also subject to keto-enol tautomerization
involving the cyclo-pentane ring in accordance with scheme (11) [1]. As in the case of sim-
pler molecules, the tautomerization in chlorophyll a (H2Cl a, XXVII) can be shifted to-
wards ketone or enol, which is controlled by the nature of the peripheral substituents in the
moloooecule, the oo
OH
OH O O
NH H N NH HN
N N NH H N HX
MX2 HX
M -X N H HN
-2HX N HN -X N H HN
N N
XXVc XXV XXVa XXVb
Figure 14 Tautomeric forms of oxoflorins.

H 2C CH C H3
H 3C C2H 5
NH N
H N HN
C H3
H 3C
H 2C H
H O
H 2C C O O C H3
C 20 H 39 O O C
XXVII
molecule, the nature of the medium, by other factors. Thus, enol (compound XXVII) is
formed not only in the medium of alcoholic alkaline [1, 113–114], but also in an
ethanol–glycerol (1:1) system and also in solutions containing a large amount of the second
component. The enol form of chlorophyll a (XXVI) is reactive. Thus, in an alkaline medium
in the presence of air oxygen it is unstable and reacts to break down the cycle (11).
NaOH, ROH O2
C C
(11)
- + - +
O O Na H3COOC O O O Na
H3COOC H H3COOC H
Inversion of one of the pyrrole rings in the formation of the tetrapyrrole macrocycle
of H2P leads to the formation of a new class of compounds – single-inverted analogues
of porphyrins [115–116] (compounds XXVIII–XXIX). They, in particular, compound
XXVIII, are characterized by an unusual type of tautomerization (12) associated with the
intramolecular transfer of one of the intramolecular NH protons (the form XXVIIIa) to the
external tertiary atom of the molecule (the form XXVIIIb:
K1
XXVIIIa + Solv XXVIIIb…Solv. (12)
The existence (in the porphyrin ana- Ph Ph

logue II) of individual tautomeric forms 3 5 7
RN
a (2-aza-21-carbatetraphenylporphin, N 2 21
CH N
22
8
CH N
2 21 H
H2[ N, CH]TPP) and b (2-imino- Ph 20 10 Ph Ph Ph
21-carbatetraphenylporphin, H[2NH, NH N N HN
21 24 23 12
CH]TPP) in a medium of nonpolar 18
17 15 13
and polar solvents, respectively, was Ph Ph

first found in [117–118]. As shown by
further investigations, tautomeric pro-
XXVIII a. XXVIII b. R = H
cesses of this type take place in inverted XXVIII. R = CH3
H2P of any structural groups, including
the analogue of unsubstituted porphin [119], β- and meso-substituted compounds [120], and
play a role in the formation (by these ligands) of numerous structural types of complexes
possessing the catalytic, receptor and other useful properties [121–123].
Berezin and coworkers [124], attracting a wide range of media with differing polarities
A
3.0
ɚ
2.5
b
c A/6
2.0
1.5 a b
1.0
c
0.5
0.0
400 450 550 600 650 700 750 λ, nm
Figure 15 Electronic absorption spectra of compound XXVIII (c = 1.5·10 –5 mol/l) in organic sol-
vents: a, C6H6; b, DMSO; c, Py.
and coordinating abilities, used electron absorption and fluorescence spectroscopies, as

well as 1H NMR spectroscopy, to study the state of compounds XXVIII and XXIX in so-
lution and to obtain stability constants of the tautomeric form XXVIIIb in benzene in the
presence of electron-donor components of the solution – N,N-dimethylformamide (DMF),
dimethylsulfoxide (DMSO), hexamethylphosphotriamide (HMPTA) and N,N-dimethyl-
propyleneurea (DMPU).
Tautomeric conversion (12) is accompanied by a significant rearrangement of the main
contour of π-conjugation in the molecule of compound XXVIII, which, in particular, fol-
lows from the EAS data (Fig. 15, a and b). The band structures in the EAS of H2TPP (II)
proper and of the tautomers of its inverted analogue XXVIII are also strongly different. At
the same time, Belair et al. [118] have shown that the EAS of compound XXVIII can be
also described by the Platt–Gouterman four-orbital model used to calculate the absorption
spectra of porphyrins proper [9–10, 33, 48, 59–60, 107–108]. Inversion of one of the pyr-
role rings in macrocycle II leads to a strong batochrome shift of all EAS bands. For instance,
the intensive Soret band (B band) in the region of 420–450 nm and the batochrome band I
(Qx, 650–730 nm) are shifted by 19 and 81 nm in the case of tautomer a of compound
XXVII and by 24 and 51 nm in tautomer b relative to the bands of compound II. The quan-
tum chemical calculations also demonstrate a long wavelength shift of the EAS band cor-
responding to the electronic transition S0→S1 and an increase of the Qx band oscillator
strength, occurring in the sequence of compounds II < XXVIIIb < XXVIIIa, which is ex-
plained by a decrease of the singlet excited state S1, especially for tautomer XXVIIIa [124].
Significant differences in the spectra of the tautomeric forms of compound XXVIII
(Fig. 15) make it possible not only to reliably identify them by the spectral data, but also
imply the individual character of the forms in solution. Thus, according to the EAS data,
the individual form a exists in a medium of nonpolar solvents with weakly pronounced elec-
tron-donor properties, such as hexane (C6H14), benzene (C6H6), toluene, dichloromethane
(CH2Cl2), chloroform (CHCl3). The type of the spectrum of tautomer a is given in
Fig. 15, a. As a rule, the permittivity (ε) of such solvents is not higher than 5, and the donor
numbers (DN) do not exceed 15 (Table 11) [68, 75].
Table 11 Properties of organic solvents and forms of existence of compound XXVIII in solutions.
Solvent(a) ε (µ) DN AN Spectrally registered

forms(b)
C6H14 1.88 (0.08) 0 0 a

C6H6 2.28 (0) 0.1 8.2 a
CHCl3 4.72 (1.15) 4.0 23.1 a
AcOH 6.15 (1.70) 20.0 52.9 H4P2+
TFA 8.26 (2.28) – 105.3 H4P2+
Diox 2.20 (0.45) 14.8 10.8 a
(CH3)2CO 20.70 (2.10) 17.0 12.5 a+b
NM 37.78 (3.56) 2.7 20.5 a + H4P+
PrOH-1 20.10 (1.65) – 37.5 a + H4P+ + H5P2+
CH3CN 36.02 (3.44) 14.1 18.9 a+b
THF 7.58 (1.75) 20.0 8.0 a+b
Py 12.30 (2.37) 33.1 14.2 a+b
DEA 3.60 50.0 – a+b
Pip 4.28 (3.87) – – a+b
DMF 36.70 (3.80) 26.6 16.0 b
DMSO 46.68 (3.96) 29.8 19.3 b
HMPTA 30.00 (5.54) 38.8 10.6 b
DMPU 36.10 (4.23) 34.0 – b
(a)Characteristics of the solvents were taken from [13, 14]; (b) H4P+ and H5P2+ are designations for
single- and double-protonated forms of compound XXVIII.
Tautomer b is realized in a medium of polar electron-donor solvents, e.g., N,N-dime-

thylacetamide (DNAc) [117–118], DMF, DMSO (Fig. 15, b), HMPTA, DMPU. Tautomer-
ic equilibrium (12) is shifted towards the form b only if a number of conditions are
observed: the permittivity (ε), dipole moment (µ) and donor number (DN) of the solvent
are sufficiently high, and the acceptor number (AN) does not exceed the donor number: ε ≥
30, µ > 3.5 D; DN > 25: DN ≥ AN. Thus, in acetonitrile (CH3CN), a solvent with ε = 36 but
low DN = 14.1 at AN = 18.9, the EAS of compound XXVII are already observed to register
a mixture of tautomers a+b (Table 11). A similar situation is realized in pyridine (Py), de-
spite its high value of DN (Fig. 15, c; Table 11). A mixture of forms a and b is also registered
in other electron-donor solvents with low values of ε and µ, such as diethylamine (DEA, ε
= 3.6; DN = 50) or piperidine (Pip, ε = 4.28) [68, 75].
Tautomeric conversion (12) can be registered by various spectral methods. Thus, the
dependence of the position of short-wavelength maxima in the fluorescence spectra (754
nm in toluene and 721 nm in DMF), as well as a change of the structure of the emission
bands depending on the nature of the solvent, is well consistent with the views of the exist-
ence of two stable tautomeric forms in compound XXVIII (Fig. 15).
The fact that in both these solvents the fluorescence spectrum does not change in the
variation of excitation and registration wavelength (Fig. 16, a and b) indicates the presence
of only one of the two spectral forms of porphyrin XXVIII in each of the given media. Ear-
lier, using a similar method, the individual character of forms XXVIIIa and XXVIIIb in
CHCl3 and DMAc was confirmed [118]. In contrast, the fluorescence spectrum of com-
pound XXVIII in Py strongly depends on the excitation wavelength (Fig. 16, c), thus dem-
λ, nm
Figure 16 Steady state fluorescence spectra of compound XXVIII depending on the excitation
wavelength (λexc = 360–460 nm) in toluene (a), DMF (b) and Py (c). T = 298 K.
onstrating the presence of a mixture of compounds – tautomers a and b in a pyridine

solution.
The state of inverted porphyrins in solutions is confirmed by the data of 1H NMR spec-
tra (Table 12) obtained for 2-aza-21-carba-tetraphenylporphin (XXVIII) and its methylated
analogue 2-(N-methylase)-21-carba-tetraphenylporphin (H[2NCH3,21CH]TPP, XXIX) in
CDCl3 and DMSO d6. Transition of compound XXVIII from the tautomeric form a to form
b is accompanied by the transfer of intracyclic proton NH, shielded by the ring current of
the aromatic π-system of the molecule, to deshielded peripheral atom of nitrogen (2N). In-
deed, the tautomerization in the medium of DMSO d6 leads to the emergence of a unit sig-
Table 12 1H NMR spectral characteristics of inverted H2P analogues in CDCl2 and DMSO d6
[124].
Compound Solvent Chemical shift, δ (ppm)
δ 21CH δ NH δ CH δ NH δ NH3
(internal) (β + Ph) (external)
XXVIII CDCl3 –5.07 (1H) –2.34 (2H) 7.61–9.81 – –

DMSO d6 0.68 (1H) 2.22 (1H) 7.52–8.13 12.81 (1H) –
XXIX CDCl3 0.98 (1H) 3.62 (1H) 7.40–7.95 – 3.32
DMSO d6 0.87 (1H) not determined 7.44–8.01 – 3.52
nal (δ = +12.81 ppm) in compound XXVIII in a weak field, corresponding to NH proton in

position 2 (Table 12), whereas in tautomer a in CDCl3 this spectral region is clean. Con-
version (12) is accompanied by a very significant weak-field shift of signals of intracyclic
CH and NH protons of molecules of XXVIIIa and XXVIIIb (by 5.75. and 2.56 ppm, re-
spectively), and, the other way round, a shift of peripheral Cβ and C-phenyl protons into a
strong field (by 1.78 ppm). This is indicative of a significant decrease of π-electronic ring
current in the case of compound XXVIIIb as compared with XXVIIIa, i.e., of the decrease
of aromaticity of tautomer b. The data of 1H NMR spectra for tautomers XXVIIIa and XX-
VIIIb are consistent with the results of [125].
As seen from the data of Table 12, inverted porphyrin XXIX, in which the extracyclic
atom of nitrogen 2N is methylated, was not subjected to tautomerization (12), so the 1H
NMR spectra of this compound in CDCl3 and DMSO d6 are close. Methylation of nitrogen
Figure 17 Correlation of the value of Kt of compound XXVIII in reaction (12) with characteristics
of organic solvents: a) Kt = f(µ); b) Kt = f(DN); 1, DMF; 2, DMSO; 3, DMPU; 4, HMPTA.
atom 2N fixes in this molecule the conjugation contour, which corresponds to that of the
tautomeric form b of nonmethylated analogue of XXVIII. As the result, by the extent of
aromaticity and position of proton signals the macrocycle of XXIX is close to the tautomer-
ic form XXVIIIb (Table 12).
The increase of the acceptor number AN of the solvent relative to the donor number
DN indicates an increase of its proton-donor properties, so equilibria (13) and (14) can be
realized in a medium of solvents with DN < AN. As the result, a mixture of tautomer
XXVIIIa with single- (H2[2NH,21CH]TPP+) and double- ({H3[2NH,21CH]TPP}2+) proto-
nated forms of compound XXVIII is formed in solution (because the permittivities of these
media are, as a rule, not high), if the acceptor number of the solvent AN < 50 as in weakly
proton-donor propanol-1 (C3H7OH-1) and nitromethane (CH3NO2); or only the double-
protonated form is formed, when AN > 50, e.g., in a medium of acetic acid (AcOH) or tri-
fluoroacetic acid (TFA) (Table 11). According to the data of [115], the first proton in the
molecule of XXVII is added to the external, more sterically accessible nitrogen atom 2N,
and the second to the intracyclic atom 23N.
K1
H2[2N,21CH]TPP + HX {H2[2NH,21CH]TPP}+X – , (13)
K1
{H2[2NH,21CH]TPP}+X – + HX {H3[2NH,21CH]TPP}2+2X – (14)
The tautomers of compound XXVIII (a and b) are converted into each other in an equi-
librium manner during the change of the composition of the inert solvent–electron-donor
solvent system (12). Berezin et al. [124] obtained tautomerization equilibrium constants (Kt
= [b]/([a]·[Solv]2, where [a], [b] and [Solv] are the concentrations of tautomer a, tautomer
b and titrant, respectively, for process (12) in C6H6 –DMF, C6H6 –DMSO, C6H6 –HMPTA
and C6H6 –DMPU systems.
As seen from the data of Table 13, the stability of the tautomeric form b increases with
the increase of solvent’s polarity, which is characterized by the values of ε and µ, and its
donor strength expressed by the donor number DN. The values of Kt were found to correlate
the best with the values of µ and DN (Fig. 17, a and b). Apparently, the DN value of the
solvent determines the strength of the bond of the donor site of compound XXVIIIb with
the extracyclic NH proton, and the high dipole moment µ favours the shift of equilibrium
(12) to the right owing to the better stabilization of ionic particles, formed in the conversion,
by the solvent. At the same time, this correlation is observed only for solvents, whose donor
strength and polarity parameters are within a certain interval favourable for the formation
of tautomer b.
It should be noted that for bulky HMPTA the value of Kt is much lower (Fig. 17) than
it could have been expected based on the values of its µ and DN, which are maximal in the
sequence of solvents considered (Table 13). Evidently, this is explainable by the worsening
of the steric conditions for the solvation of the 2NH grouping of inverted porphyrin by the
bulky molecule of the solvent. Thus, the molar volumes (VM) for DMF, DMSO and DMPU
are within the limits of 100 cm3/mol, whereas VM for HMPTA is almost twice as high
(Table 13).
Table 13 Tautomerization constants of porphyrin XXVIII in a C6H6 –donor solvent medium and
characteristics of the solvents [124].
Solvent Kt ε(a) µ, D(a) DN(a) VM(a), cm3/mol
DMF 0.23±0.01 36.7 3.8 26.6 77.4

DMSO 0.39 ±0.01 46.7 3.96 29.8 71.3
HMPTA 0.47±0.02 30.0 5.54 38.8 175.7
DMPU(b) 0.67±0.01 36.1 4.23 34.0(c) 98.5
(a) Characteristics of the solvents were taken from [68, 75]; (b) from [126]; (c) donor strength Ds.
The value of Kt in the temperature range of 298–328 K within the limits of experimen-
tal error does not depend on temperature [124]. From this, it follows that tautomerization
process (12) proceeds with the activation energy E close to zero, which is consistent with
the data of quantum chemical calculations of tautomers a and b [109, 124, 127]. Thus, the
equilibrium state of (12) is largely controlled by entropy factors and, at an active participa-
tion of a solvent in the process, by solvation factors. Indeed, earlier studies showed tautomer
b to be stabilized by the molecule of an electron-donor solvent even in a crystal [117].
Several ways are possible to realize the tautomerization mechanism of inverted ana-
logues of porphyrins: within the involvement of a solvent by tunnelling or intermolecular
transfer of NH proton, as well as owing to its intramolecular transfer with participation of
a solvent [117, 128]. In the opinion of Furuta et al. [117], intramolecular proton transfer in
the course of the tautomeric conversion of compound XXVIII is performed owing to the
emergence of a strongly nonplanar conformation, in which the inverted pyrrole ring is in-
cluded towards the coordination core, and the inner and outer nitrogen atoms are brought
closer to each other. This treatment of the mechanism appears to be open to question, as the
reaction centres of a tetrapyrrole molecule involved in proton transfer are too far away one
from the other to enter into direct interaction, and the approximation of these centres to at
least a distance of an effective H bond (≤3 Å) should strongly destabilize the molecule.
According to the data of quantum chemical calculations and XDA data, both tautomers of
compound XXVIII are almost planar and, besides, close energy-wise molecules. The
difference in energy of the two tautomers is 3.4–5.7 kcal/mol according to the data of
[109, 127].
The strong dependence of the value of Kt on the nature of solvent, the evident impor-
tant role of the entropic contribution to the Gibbs energy of the tautomerization process, as
well as the fact of the involvement of two solvent molecules in the proton transfer found by
us (it follows from the values of tan α of the indicator dependence [log(Cb/Ca) = log (CSolv)
Figure 18 Dependence of the indicator relation (logCb/Ca) of the concentration of the tautomeric
forms of compound (II) in benzene on the logarithm of the concentration of the donor solvent
(log Csolv): 1, DMF; 2, DMSO; 3, DMPU; 4, HMPTA. tan α = 2.
N N
CH HN CH HN
Solv
- H Solv+ - iso
K ɢɡɨ
NH N N N
K1
a a1
-N Solv ...HN
CH N CH N
H Solv+
N HN K2 N HN
b1 b
Figure 19 Schematic tautomerization mechanism of inverted porphyrin analogues [124].
(Fig. 18)) enable us to propose an alternative mechanism of tautomerization involving a sol-

vent (Fig. 19).
According to the mechanism, conversion of tautomer a to tautomer b is realized via a
number of consecutive equilibrium stages. At the first stage (a ↔ a1), there is a polarization
under the influence of the molecule of electron-donor solvent (Solv), and then complete dis-
sociation of one of the intracyclic NH bonds in the molecule of inverted porphyrin (Fig. 19).
The proton can with equal probability be abstracted both from 22N and 24N – in both cases
the second stage (a1 ↔ b1), of the isomerization proper of the conjugation contour, is ac-
companied by intracyclic cis-tautomerization (22NH→23N or 24NH→23N). Presumably,
abstraction of the proton from the 22N grouping should be more probable due to its increased
acidity. In this case, the proton is located slightly closer to the electronegative 2-aza atom.
The last, third stage is accompanied by the protonation of the peripheral nitrogen atom (b1
↔ b). Intermediate particles a1 and b1 are highly reactive and are not revealed in the EAS
in any way. For this reason, the total tautomerization constant Kt (15) was calculated pro-
ceeding from the expression for the constants of the elementary stages of the process,
(16)–(18). Within the framework of this approach, the participation of two molecules of
electron-donor solvent in the course of tautomeric process (12) is well explained.
Kt = K1 ·Kiso ·K2 , (15)
K1 = [a1]/([a]·[Solv]), (16)
Kiso = [b1]/[a1], (17)
K2 = [b]/([b1]·[Solv]. (18)
Thus, inverted analogues of porphyrins, such are compound XXVIIIa, also pertain to
tetrapyrrole macrocycles with chemically active NH bonds (see the mechanism). Tautomer
XXVIIIb is less aromatic and, therefore, possesses no NH activity.
1.3 Quantitative assessment of the state of NH bonds in porphyrin molecules

The most evident quantitative assessment of the chemical activity of NH bonds in H2P mol-
ecules is the direct determination of their acidity by equations (2)–(3) (Section 1.1.1). How-
ever, the reliable and comparable data by the acidic dissociation constants of porphyrins are
far from being readily accessible [61]. In this connection, in [26, 58] it was proposed to use
a complex of criteria of state and reactivity of NH bonds in porphyrin molecules. They in-
clude the 1H NMR, kinetic and quantum chemical criteria.
1.3.1 Spectral criterion

1
The H NMR spectral criterion is based on the value of the chemical shift (19) of H2P NH
proton signals in the spectrum during the substitution of inert, weakly solvating solvents (SI
= C6H6, CHCl3, CH2Cl2), in which NH bonds of the porphyrin molecules are always local-
ized, by strongly electron-donor solvents (SB = DMSO, Py), in the medium of which pro-
tons of these bonds can be delocalized. The weak-field shift of the value
∆δNH = δSI – δSB, (19)
of δNH in porphyrins can be caused by two main causes – activation of the NH bond with
abstraction of protons by a donor solvent from the region of their shielding by the ring cur-
rent of the π-system and disturbance of the planar structure of the molecule [72]. The type
and depth of nonplanar conformation varies in various H2P [19], so it would not be correct
to compare the values of δNH of these compounds to make a judgement on the delocaliza-
tion of the NH bond. When substituting the solvent, the conformational composition of H2P
molecules does not in fact change [14], and the NH bond can only be activated, so to assess
the latter, one should use namely the value of the relative chemical shift of the signals of
NH protons, ∆δNH (19). It is positive, if porphyrin has chemically active NH bonds, and
negative in the cases when the bonds, as in classical H2P, can not be activated by elec-
tron-donor solvent (Table 14). According to the 1H NMR spectral criterion, the sequence
of chemical NH activity of H2P considered is as follows: IV << VIII < VI < VII. NH bonds
are minimally localized in weakly nonplanar porphyrin IV. Polarization of NH bonds in-
creases in transition to rigid nonclassical H2P (compound IX) and is maximal in strongly
nonplanar dodeca-substituted porphyrins (VI–VII).
Table 14 Effect of a solvent on the position of NH proton signals (δ NH, ppm) in 1H NMR spectra
of porphyrins.
Porphyrin Solvent δ NH ∆δ NH, ppm Ref.
H2TAP (VIII) CD2Cl2 –2.07 35

Py –0.97 +1.10 35
H2TPP (II) CDCl3 –2.76 –
DMSO d6 –2.91 –0.15 –
H2TPTBP (IV) CDCl3 –1.16 –
DMSO d6 –1.10 +0.06 –
H2(β-Et)8TPP (V) CDCl3 –2.00 72
DMSOd6 not determ. – 72
H2(β-Ph)8TPP (VI) CDCl3 –0.90 72
DMSO d6 +1.0 +1.90 72
Py d5 not determ. – 72
H2T(β-Br)8TPP (VII) CDCl3 –1.65 – –
DMSO d6 +0.52 +2.10 –
1.3.2 Kinetic criterion

Similar changes are observed in the kinetic assessment of the chemical activity of NH
bonds, which has been used in porphyrin chemistry over a period of years [5, 28, 34–35,
52–54, 129], but formulation of the kinetic criterion has not been done earlier. According
to the mechanism of complexation of H2P with metal salts MX2 (1), one of the most energy-
intensive stages of the process is elimination of NH protons from the coordination cavity
of N4H2 [1, 25]. For this reason, the rate of reaction (1) strongly depends on the state of NH
bonds and the nature of solvent (Solv), and in nonclassical porphyrins it strongly increases
in a medium of electron-donor solvents activating NH bonds [26].
In accordance with the kinetic criterion, porphyrin is chemically NH active, if the rate
of its coordination by d metal salts in a medium of electron-donor solvents (SB = DMSO,
DMF, Py) under comparable conditions is higher than the rate of reaction (1) in a proton-
donor solvent (e.g., SA = AcOH or ROH), i.e., for the true constants kv of the rate of reaction
(1) expression (20) is observed (Table 15). Herewith, d metal and anion of the salt can vary,
and the most suitable will be Cu(II), Zn(II), Co(II) acetates.
kv(SB) > kv(SA). (20)
The values of the constants given in Table 15 make it possible to determine compounds
I–II as classical H2P, and compounds IV–VII and VIII–X as nonclassical ones, and indi-
cate an increase of the chemical activity of NH bonds in the following sequence of porphy-
rins pertaining to groups (2) and (3) (Fig. 3): X < VIII < IX and IV < V < VI < VII. From
the data of Tables 14 and 15, it follows that polarization of the H2P molecule by peripheral
substituents, in particular, those of opposite electronic nature (the push-pull effect) enhanc-
es the chemical activity of NH bonds. This is well seen by the example of H2(β-Br)8TPP
(VII), carrying eight electron-acceptor functional groups in β-positions of the molecule, as
well as compounds VIII–IX (Table 15).
Table 15 Dependence of the rate of reaction (1) of porphyrin with Zn(II) and Cu(II) acetates (kv,
l ·mol–1 ·s–1) on the donor-acceptor nature of a solvent.
Porphyrin Concentration of salt, Rate constant (kv, l ·mol–1 ·s–1) Ref.

CMAc ·10–3, mol/l and direction of its change
2
AcOH Py
H2(β-Br)4TAP (IX) M=Zn, (0.3) 0.007±0.0003 930±140 → 37,53–54

H2TAP (VIII) M=Zn, (0.3) 0.0087±0.0003 5.79±0.04 → 37,53–54
H2TBP (X) M=Zn, (2.6) slow 0.0069±0.0018 → 31–32,38
H2(β-Et)8P (I) M=Zn, (5.0) 3.67±0.05 slow ← 1,75
H2TPP (II) M=Zn, (0.48) 38.6±0.90 slow ← 1,75
H2TPTBP (IV) M=Zn, (2.6) slow 0.100±0.001 → 31–32
H2(β-Et)8TPP (V) M=Cu(II), (0.05) 0.00065±0.00004 1230±40(a) → 51–52
H2(β-Ph)8TPP (VI) M=Cu(II), (0.005) slow 754±20 → 51–52
H2(β-Br)8TPP (VII) M=Zn(II), (0.22) slow fast 130
(a)
Csalt = 0.005 mol/l.
Chemical activation of NH bonds by molecules of an electron-donor solvents is exhib-

ited not only in the change of the rate constants, but also of other kinetic parameters of in-
dicator reaction (1), in particular, its entropic characteristics.
A recent work [85] found significant differences in the kinetic parameters of reaction
(1) of the complexation of N- (XIX–XX) and dodeca- (V–VI) substituted molecules of por-
phyrins taking in solution nonplanar conformations, which are predominantly saddled, but
differ in symmetry and degree of deformation [15, 19]. It is not hard to notice (Table 16)
that the change of entropy ∆S # in the course of activation of reaction (1) reagents for dodeca-
substituted porphyrins is always more positive than for N-substituted porphyrins, but only
an electron-donor solvent (DMF).
The change of entropy of the reaction proceeding in solution without the change in the
number of moles of substance is related to the change of solvation of particles in the for-
mation of the transition state [1–8, 11, 21, 25, 31–32, 34–39, 52–54, 57, 74–77, 84–85].
From this perspective, the rise of the value of ∆S # in reaction (1) can be explained by either
the desolvation of the transition state, which is little probable, or by an additional solvation
of the initial reagents, achieved under certain conditions. In our opinion, the rise of ∆S # of
the complexation reaction of NH-active dodeca-substituted porphyrins in an electron-donor
solvent is directly related to the ability to form PTCs [30]. In the case of a nonpolar medium
or NH-inactive compounds, there is no increase of ∆S # (Table 16).
Formation of PTC leads to the increase of solvation of the initial H2P in reaction (1)
and its acceleration owing to NH activation as compared with NH-inactive porphyrins. It
is known [1, 5, 25] that one of the most energy-intensive contributions to the total energetics
of reaction (1) is the dissociation of NH bonds in the H2P ligand. Formation of PTC is char-
acteristic of dodeca-substituted H2P and not characteristic of N-substituted porphyrins (see
also Section 1,4), which is reflected on the activation parameters of the reaction (Table 16).
Earlier studies of the kinetics of coordination of tetrabenzoporphyrins with chemically
active NH bonds by zinc acetate in a pyridine–diethylamine medium found [32] that, with
the rise in the concentration of the base DEA, an increase of the rate of entropy is observed
along with an increase of the reaction rate in the course of intermediate state formation.
ooooo
Table 16 Effect of the nature of porphyrin on the entropic characteristics of complexation reaction
(1) with Zn(II) and Cu(II) salts in C6H6 and DMF.
Porphyrin Salt Solvent kv298 , Ea , ∆S #,

l· mol –1 ·s–1 kJ · mol –1 J · mol–1 · K–1
H(N-Me)(β-Et)8P CoAc2 DMF 30.94±1.39 53.6±1.3 –63±2

(XIX) Co(Acac)2 DMF 18.13±0.69 59.7±1.8 –29±1
Co(Acac)2 C6H6 very slow
H(N-Me)TPP (XX) ZnAc2 DMF 18.8±0.73 32.8±2.0 –119±9
Zn(Acac)2 DMF 12.51±0.50 40.2±1.2 –97±3
Zn(Acac)2 C6H6 very slow
H2(β-Et)8TPP (V) ZnAc2 DMF 9.66±0.11 62.6±0.7 –24±1
Zn(Acac)2 DMF 4.01±0.21 88.9±4.0 56±2
CoAc2 DMF 17.55±0.47 60.0±2.2 –26±1
H2P(β-Ph)8TPP (VI) ZnAc2 DMF 1.81±0.05 93.2±2.3 64±1
Zn(Acac)2 DMF 5.29±0.21 68.5±2.1 –9±1
Zn(Acac)2 C6H6 83.79±2.18 32.5±1.3 –108±4
CoAc2 DMF 5.39±0.30 77.8±1.8 22±1
Co(Acac)2 DMF 2.58±0.09 77.1±3.6 13±1
Co(Acac)2 DMFC6H6 10.48±0.56 79.3±3.1 –32±1
Thus, for instance, during the change of the concentration of DEA in the
H2TPTBP(IV)–ZnAc2 –Py–DEA system from 0 to 1 mol/l, the true reaction rate constant
increases from 0.1 up to 3.93 l⋅mol –1 ⋅s –1, and the entropy change value increases from
–143 up to –48 J·mol –1 ⋅s –1. Herewith, the value of kv was found to directly depend on
the molar concentration of DEA (Fig. 20). Such changes can not be explained by a decrease
of additional solvation in the formation of the transition state of reaction (1), but are caused
by a decrease of the entropy of the initial state, as the result of the interaction of DEA mol-
ecules with chemically active NH bonds of H2P before entering into reaction (1).
kv, mol/(l·s)
CDEA, mol/l
Figure 20 Dependence of the rate constant of reaction (1) on the concentration of DEA for H2TBP
(X) in the Py–DEA system (a) and H2TPTBP (IV) in Py–DEA (b) and DMSO–DEA (c) systems.
1.3.3 Quantum chemical criterion

In the recent years, a very popular method has been the quantum chemical assessment of
NH acidity of porphyrins [109]. Thus, Stuzhin [47] has shown that for NH-active H2P (e.g.,
IV–VII) the value of a relative stability of dianions P2– (δ∆Hf(0-2)) calculated by the AM1
method is more positive compared with classical H2P (compounds I–II) (Table 17). Por-
phyrins whose chemical activity of NH bonds is high are characterized by the value of
δ∆Hf(0-2) > 0 (compounds V–VII, IX, XVII). This fact is consistent with the results of as-
sessing the state of these bonds in accordance with the spectral and kinetic criteria and is
supported by EAS data (Fig. 21).
Table 17 Quantum chemical (AM1) and spectral (EAS) assessment of the chemical activity of NH
bonds in H2P molecules (I–II, IV–X, XVII–XVIII).
δ ∆Hf (0-2), Parameters of Qx band in EAS [λQx, nm; (ε)]

Porphyrin kcal/mol(a)
CHCl3(b) DMSO(b)
H2(β-F)4TAP (XVII) +3.11 – –

H2(β-Cl)4TAP (XVIII) –0.30 – –
H2(β-Br)4TAP (IX) +4.04 639 (4.71) 620
H2TAP (VIII) –23.18 617 (4.75), C6H5Cl 613 (4.71), Py
H2TBP (X) –16.55 663 (3.45) 663 (3.53)
H2(β-Et)8P (I) –39.01 619 (3.74) 619 (3.74), DMF
H2TPP (II) –25.64 646 (3.55) 646 (3.68), DMF
H2TPTBP (IV) –2.34 694 (3.85) 696 (3.57)
H2(β-Et)8TPP (V) +9.66 696 (4.06) 715 (3.97)
H2(β-Ph)8TPP (VI) +13.28 709 (4.36) 749 (4.64)
H2(β-Br)8TPP (VII) +22.82 738 (3.84) 792 (4.16)
(a)
By the data of [47]; (b) using the data from [8, 35, 39, 55].
1.3.4 Insufficiency of absorption spectrum analysis of porphyrins

Nonclassical porphyrins with highly chemically active NH bonds are readily distinguish-
able from inert classical porphyrins by comparing their EAS in weakly solvating and elec-
tron-donor solvents (Fig. 21). With electron-donor solvents, these NH-active H2P form H
associates, which are also called proton-transfer complexes (PTC) (21) [30, 32, 55, 93].
oooooooooooooooo
K1 K1
Solv
H2P + Solv HPδ–‘ …Hδ+…Solvδ– Solvδ–…Hδ+…Pδ+‘…Hδ+…Solvδ– (21)
However, electron spectroscopy data can not be a reliable criterion, as they make it possible
to characterize the chemical NH activity of H2P only in the cases when this property is pro-
nounced in compounds [32]. A large number of porphyrins, pertaining in accordance with
the above three criteria to H2P with a chemically active NH bond, e.g., compounds IV, VIII
and X, do not practically change their EAS in electron-donor solvents (Table 17) because
their PTC formation equilibria (21) are strongly shifted to the left. An example of an EAS
for a weakly NH-active porphyrin IV [32, 39] is given in Fig. 21, a. In contrast, compound
V, which possesses a high chemical activity of NH bonds, forms in DMSO a PTC, which
(a)
(b)
λ, nm λ, nm
Figure 21 Electronic absorption spectra of (a) H2TPTBP (IV) and (b) H2(β-Et)8TPP (V) in CHCl3
(1) and DMSO (2).
is accompanied by a decrease in the number and by a shift of EAS bands (Fig. 21, b). Here-
with, the shift of the most batochromic Qx band (Table 17) is the greater, the easier the PTC
is formed by porphyrin.
Some porphyrins can not be tested for the chemical activity of NH bonds by all three
criteria. It has been found [26, 58, 85] that H2P, for which at least two of the three assess-
ment criteria of the state of NH bonds are observed, can be considered to be NH-active. A
larger number of criteria of NH activity can be suggested using other methods.
1.4 Nonplanar structure of the macrocycle and chemical NH activity in its

coordination core
Porphyrins can acquire a nonplanar structure as the result of a number of structural changes
[19, 85, 87], such as:
• Modification of the periphery of the H2P molecule:

(a) multiple (nona-, deca-, undeca- or dodeca-) substitution of peripheral hydrogen
atoms simultaneously in β- and meso-positions [19, 22, 85, 130–135]
(b) substitution of meso-positions by bulky functional groups (iso-propyl, tert-butyl etc.)
[19, 136–140]
• Modification of the H2P coordination centre:
(a) substitution of N atoms of intracyclic NH groups (N-substitution) [15, 85, 87,
141–144]
(b) protonation of tertiary atoms of nitrogen {mono- (H3P+) or di- (H4P2+) cationic
forms of H2P, equations (4)–(5)} [61, 145–147]
(c) complexation with metals whose ionic radius is much smaller (Ni(II)) or, vice versa,
exceeds (Zn(II)) the size of the coordination cavity (some metalloporphyrins)
[16, 19, 65]
• Changes in the H2P molecule leading to a decrease of aromaticity:
(a) formation of radical forms of porphyrins [16, 65]
(b) reduction of Cβ –Cβ bonds in pyrrole rings of H2P macrocycles (bacteriochlorins,
corrins etc.) [9–10, 87]
• Combined action of several factors (dodeca-substitution + N-substitution; dodeca-
substitution + protonation etc.) [87, 143–145].
As the result of extensive X-ray diffraction studies [16, 19], all known H2P and MP
with nonplanar structure were classified by the type of molecule distortion as saddled, ruf-
fled, domed, waved, stepped, twisted, gabled etc. [19].
The porphyrin nonplanarity criteria can be not only XDA data [12–13,19, 65], which
describe the structure of macrocycles in the solid phase, but also fluorescence data [22, 33,
49–50, 107, 135, 140, 142, 147], RR (resonance Raman) spectroscopy [134, 148–149], and
in some cases NMR spectral results [72, 132, 150–151].
A crucially important issue is that of the relation between the type of nonplanar struc-
ture of the porphyrin macrocycle and the chemical activity of NH bonds in its coordination
core. If such a relation does exist and, thus, a simple change of a nonplanar conformation
of the H2P molecule may cardinally change its properties, the mechanism of the processes
involving these molecules in vivo can be assumed to be similar [8, 17, 27].
The most pronounced distortion types of the planar structure of the H2P molecule in-
clude, besides dodeca substitution, the substitution of meso-positions by bulky, most often
alkyl, groups, which leads to the ruffled conformation, as well as the substitution of one or
a larger number of nitrogen atoms in the coordination core to form N-substituted porphyrin
analogues [8, 19, 87]. In the latter case, the saddled conformation of an asymmetric type
was found [142].
Section 1.3, which discussed the criteria of the chemical activity of NH bonds in H2P
molecules, did not consider other types of strongly nonplanar macrocycles, except dodeca-
substituted.
Figure 22 Absorption spectra (solid line) and fluorescence spectra (dashed line) of H(N-CH3)TPP
(XX) in toluene, T = 298 K [142].
At the same time, both meso-substituted H2P [19, 49–50, 70, 72, 136–140] and N-sub-
stituted porphyrin analogues [15, 24, 61, 71, 85–87, 141–144] by their spectral character-
istics, physicochemical properties and reactivity behave as compounds with a nonplanar
structure. Thus, all bands in the visible range of their EAS are shifted batochromically rel-
ative to the predominantly planar H2P [15, 139], which, however, is not a reliable criterion
of nonplanarity [23]. The photophysical characteristics describing the properties of excited
states are more reliable [107]. Significant Stokes shifts of adjacent bands in the fluorescence
spectra relative to absorption bands (22), low quantum yields of fluorescence and triplet
states’ yields are considered to be features of nonplanar porphyrins [33, 49]. Besides,
oooooooo
∆ν1 = ν1fl – ν1abs (22)

characteristic modes appear in vibrational spectra of nonplanar porphyrins in the region of

1300–1700 cm–1, the so called marker lines ν2–ν4, sensitive to the change of H2P spatial
structure [149]. The 1H NMR spectra are observed to have weak-field shifts of NH proton
signals owing to the deshielding of the coordination core in the distortion of the molecule
planar structure [72]. As the result of activation of extraplanar vibrations, porphyrins of the
structural groups considered acquire an increased solubility in organic solvents (Table 7)
[86, 152], redox properties [153] unusual for predominantly planar H2P. The stability of the
ligands and their complexes to the action of elevated temperatures [64, 154] and chemical
reagents [136, 154] decreases. For instance, strongly nonplanar H2T(tert-Bu)P (XIII),
which exists predominantly in ruffled conformation (∆Cmeso = 0.9 Å), proves so reactive
that in the presence of acids or d metal salts nucleophilically attaches solvent molecules
(CH3OH), thus forming porphodimethene structures [136]. N-methylation of the H2TPP
(II) molecule decreases the stability of its zinc complex in a medium of glacial AcOH
100-fold [154]. Apparently, as the result of partial localization of π-electronic density the
basicity of tertiary nitrogen atoms in pyrrole rings of nonplanar macrocycles (XI–XIII,
XIX–XXI) increases, as well as the ability of the nitrogen atoms to be protonated even in
the presence of mean-strength acids [61,85, 154]. Their complexation ability in a medium
of solvents with moderate donor-acceptor characteristics, e.g., in acetonitrile, increases too
[155].
The issue of the appurtenance of these nonplanar macrocycles (XI–XIII, XIX–XXI)
to porphyrins with chemically active NH bonds has not been discussed earlier. To the best
of our knowledge, there are no data at all at present on the acidity of NH bonds in porphyrin
molecules with bulky meso-substituents. Sufficiently contradictory information is available
on the effect of intracyclic N-substitution into H2P molecule on their acidic properties [15,
61].
In such a case, use can be made of the above criteria of the chemical activity of NH
bonds in porphyrin molecules [26]. Unfortunately, there is no information in the literature
on the dependence of these 1H NMR spectra of meso-substituted H2P (XI–XIII) and their
N-substituted analogues (XIX–XXI) on the nature of solvent. Besides, in most cases the
proton signal of an NH group is not exhibited at all in 1H NMR spectra of N-substituted
porphyrins [15]. For this reason, we shall use two other NH activity criteria – quantum
chemical and kinetic [26].
Table 18 compares the results of a quantum chemical calculation of enthalpies of form-
ing H2P ligands, their mono- (HP – , ∆Hf(–1), kcal/mol) and dianions (P 2– , ∆Hf(–2),
kcal/mol), as well as heats of deprotonating the ligands to mono- and dianions in the gas
phase for predominantly planar NH-inactive porphyrins (I–III), as well as nonplanar meso-
substituted porphyrins (XI–XIII) and N-substituted H2P analogues (XIX–XXI). The val-
ues of ∆Hf(–2) taken as estimates show that, in contrast with dodeca-substitution of H2P
(compounds IV–VII), meso- or N-substitution in the macrocycle does not lead to chemical
activation of NH bonds.
Thus, the values of ∆Hf(–2) show only very weak tendencies to the shift towards more
positive values (a mere 5–8 units), whereas even in compounds with weakly pronounced
NH activity, such as H2TBP (X) and H2TPTBP (IV), these shifts reach 26.5 and 23.3 units,
respectively (Table 18). As N-substituted porphyrins are monobasic acids, analysis of quan-
tum chemical data for them can be done using the value of ∆Hf(–1). As it follows from the
data of Table 18, this value does not practically increase relative to unsubstituted porphin,
either, as compared with NH-active compounds (IV and X).
Thus, quantum chemical data indicate the absence of a noticeable chemical activity of
ooooo
Table 18 Enthalpic characteristics of deprotonation of {∆H f(–1) and ∆H f(–2) (kcal/mol)} and
formation of mono- and dianionic forms {∆H (–1) and ∆H (–2) (kcal/mol)} of porphyrins with planar
and nonplanar structure (calculated by the AM1 method).
Macrocycle ∆Hf(0) ∆Hf(–1) ∆Hf(–2) δ ∆Hf(0–1) δ ∆Hf(0–2) ∆H(–1) ∆H(–2)
H2P 221.00 200.81 264.09 20.19 –43.09 347.01 777.49

H2(β-Et)8P (I)(a) –39.01
H2TPP (II)(a) –25.64
H2T(n-Pr)P (III) 151.91 132.54 192.37 19.37 –40.46 347.83 774.86
H2T(iso-Pr)P (XI) 178.54 160.12 217.27 18.42 –38.73 348.78 773.13
H2T(cyclo-Hex)P (XII) 131.62 111.92 167.04 19.70 –35.42 347.50 769.82
H2T(tert-Bu)P (XIII) 195.96 176.05 231.19 19.91 –35.23 347.29 769.63
H(N-Me)(β-Et)8P (XIX) 129.24 113.26 – 15.98 – 351.22 –
H(N-Me)TPP (XX) 352.27 329.70 – 22.57 – 344.63 –
H(N-Ph)TPP (XXI) 392.90 372.72 – 20.18 – 347.02 –
H2TBP (X) 265.25 237.53 281.80 27.72 –16.55 339.48 750.95
H2TPTBP (IV) 401.29 371.91 403.63 29.38 –2.34 337.82 736.74
(a)
Data of [47].
NH bonds in nonplanar meso-substituted H2P and N-substituted porphyrin analogues. Let

us turn to the data on the kinetics of complexation reaction (1) of these ligands, as they make
the basis of the kinetic criterion of H2P NH activity. It follows from the obtained experi-
mental data (Table 19) that the complexation rates of N-substituted compounds decrease
with the increase of the electron-donor value of the solvent in the sequence: PrOH-1 > DMF
> DMSO > Py [31, 85]. This corresponds to the behaviour of porphyrins with a localized
NH bond. Moreover, in a medium of pyridine, which activates NH bonds, in the reaction
of nonplanar H(N-Me)TPP (XX) with zinc acetate the rate of the process not only fails to
increase but, upon reaching a certain concentration of the product, an equilibrium is settled
[31]. An additional argument in favour of the absence of NH activity in N-substituted por-
phyrins are the negative values of activation entropy change in reactions (1) of their coor-
Table 19 Kinetic parameters of reaction (1) of N-substituted porphyrins with ZnAc2 in organic
solvents [31, 39, 85].
Porphyrin Csalt ·103, Solv kv298 , Ea , ∆S #,

mol/l l· mol –1 ·s–1 kJ · mol –1 J · mol–1 · K–1
2.6 Py 0.014 77.0±1.4 –30±2

H(N-Me)(β-Et)P 2.0 DMSO 5.20 52.8±2.8 –56±3
(XIX) 2.0 DMF very fast – –
0.2 DMF 131.4 7.1±0.1 –189±15
0.2 PrOH-1 very fast – –
2.6 Py very slow – –

H(N-Me)TPP
(XX) 2.0 DMSO 6.14 34.1±2.8 –124±7
2.0 DMF very fast – –
0.2 DMF 18.8 32.8±2.0 –119±9
0.2 PrOH-1 very fast – –
dination by d metal salts (Table 16). In contrast, the positive values of ∆S # = S # – S init in
NH-active H2P are associated with a significant solvation of the initial state on the coordi-
nate of reaction (1).
There are practically no data on the complexation kinetics of nonplanar porphyrins in
ruffled conformation. Our studies, using H2T(iso-Pr)P (XI) and H2T(cyclo-Hex)P (XII)
(∆Cmeso for their complexes with Ni(II) are equal to 0.74 and 0.77 Å [19], respectively),
have shown that the rates of this reaction with copper, zinc or cadmium acetates in electron-
donor solvents (DMF, DMSO), assuming NH activation, are very low, and respective com-
plexes within the temperature range of 298–338 K are not formed in practice. This fact is
a good proof of a localized NH bond in the molecules of meso-substituted porphyrins.
It can also be noted that H2P with bulky meso-substituents, as a rule, do not in practice
enter into complexation reactions with d metal salts in acetic acid and other solvents with
the pronounced proton-donor function. The disturbance of the planar structure of the mac-
rocycle leads to partial localization of the π-electronic density in pyrrole rings and an in-
crease of basicity of the molecule. As protonated forms of porphyrins produced in this case
in proton-donor media in reaction (1) are inactive [1, 7], this rule is also valid for other non-
planar H2P, such as N- or dodeca-substituted molecules. Some dodeca-substituted com-
pounds also enter into reaction (1) in a medium of AcOH, but at very low rates [51–52].
The best medium for MP synthesis in the case of nonplanar meso-substituted porphyrins is
a polar solvent with moderate donor-acceptor properties, e.g., CH3CN, in which reaction
(1) with more nonplanar H2T(cyclo-Hex)P (XII) proceeds at 298 K very rapidly, and with
H2T(iso-Pr)P (XI) with kv ~ ⋅10 l⋅mol –1 ⋅s –1.
Thus, the criteria of the chemical activity of NH bands in the coordination core are a
universal tool for the assessment of the reactivity of porphyrins and are applicable for both
H2P belonging to various structural groups and their analogues. The second important con-
clusion is that a disturbance of the planar structure of the H2P molecule does not automat-
ically assume the presence of chemically active NH bonds. The chemical activity of these
bonds depends in a specific way on the type of conformation, in which a nonplanar macro-
cycle is. Thus, only the occurrence of the symmetrically distorted pronounced saddled con-
formation, possessing a high orthogonal dipole moment [22], leads to the emergence of an
NH activity in nonplanar H2P. Nonplanar porphyrins in ruffled conformation are low-polar
and exhibit no chemical activity of NH bonds.
2 Reactivity of the Coordination Core in Molecules of Porphyrin Ligands

As we have already mentioned, the chemical activity of NH bonds is vividly exhibited in
the character of the progress of those chemical processes, which affect these bonds in one
way or another. For H2P ligands and their analogues, these are primarily complexation re-
actions (1), acid-base interaction reactions ((2)–(5)), as well as coordination-core reactions
of nucleophilic substitution.
2.1 Complexation reactions

The features of reaction (1), which follow from its mechanism (Fig. 23), have already been
considered in Section 1. Here we would only list them once again, bearing in mind the sig-
nificance of the state of NH bonds in the H2P molecule on the rate of this reaction:
1. The reactivity of porphyrins as aromatic macrocyclic ligands in complexation

N... H . solv X. solv solv . H

solv
N X. solv
solv solv
N Ɇ solv 1 N
+ Ɇ
N
solv X. solv N solv
solv . H ... N solv . H N X. solv
Salt
ɋɨɥɶ
Porphyrin
ɉɨɪɮɢɪɢɧ 2 Intermediate
ɂɧɬɟɪɦɟɞɢɚɬ
×
solv .H......N
solv
N X. solv
Ɇ Metalloporphyrin
Ɇɟɬɚɥɥɨɩɨɪɮ
N
X. solv
solv . H ......N
solv
Transition state
Figure 23 A schematic mechanism of the complexation reaction of porphyrins with d metal salts
[1, 5, 11, 25].
reaction (1) is controlled by the intensity of the macrocyclic effect, i.e., is determined
by the rate of the electronic and structural MCE components affecting the process
[6–8].
1.1. The structural component always hinders reaction (1) to a greater or smaller extent
due to the spatial shielding of the coordination core by the surrounding atoms and
π-electronic cloud.
1.2. The π-electronic MCE component is determined by the extent of aromaticity of the
H2P macrocycle, and in the case of high delocalization of π-electrons in the macroring
induces the polarization of the molecule and some chemical bonds in it.
Thus, the character of the macrocyclic effect depends on the type of H2P structure –
planar or nonplanar [8].
2. The progress of reaction (1) (Fig. 23) for porphyrins with a different degree of rigidity
strongly depends on the nature of a solvent.
2.1. Reaction (1) of H2P with a chemically inactive NH bond proceeds the most readily
in a medium of solvents – moderate-strength ligands (carboxylic acids, alcohols),
because it is determined by the strength of the solvatosalt [MX2Solv(n-2)] [75]. The
strength of the solvatosalt depends on the coordinating ability of the metal ion and the
nature of ligands – molecules of a solvent (Solv) and anions of a salt (X – ). If the bonds
M–X and M–Solv in the solvatosalt are sufficiently strong, an intermediate is formed,
a complex with H2P of amine type (Fig. 20), which is easily broken by electron-donor
additives [1, 25].
2.2. Complexation reaction (1) for H2P with a chemically active NH bond is accelerated
in solutions of electron-donor solvents. In this case, the state of the NH bond is the
major factor determining the rate of reaction (1) [1, 32, 129].
Chemical activation of NH bonds not only leads to accelerate reaction (1), e.g., owing
to the interaction of these bonds with molecules of an electron-donor solvent, but also to a
possibility of controlling more complex and fine processes, e.g., in the course of tautomeric
conversions. Thus, it has been found that, depending on the state of tautomeric equilibrium
F F
F F F F
F F F F
F FN F F F F F F
HN
C N p-TolSO2NHNH2 C N
II
F Cu III F F Cu F
N N DDQ N N
F F F F F F F F
F F F F
F F F F
F F
Figure 24 Redox processes involving the fluorinated analogue of compound CuXXVIII [121].
of compound XXVIII, the oxidation of metal occurring in the ligand can be controlled; here-
with, the metal is in a bidentate state (XXVIIIb) or a tridentate state (XXVIIIa) [117,
122–124]. Thus, Maeda and Furuta [122] propose a simple method of oxidation of Cu(II)P
DDQ to Cu(II)P, which proceeds without the change of the geometry of the coordination
node (plane-square complexes) (Fig. 24). The inverse process is performed in the presence
of p-toluene sulfonyl hydrazide. Both processes proceed quantitatively and almost instan-
taneously. They are realized as easily electrochemically {Cu3+ \Cu2+ = 0.15V} [121].
2.2 Proton ionization of NH bonds in H2P molecules

The rise of chemical activity of NH bonds leads to the formation of proton-transfer com-
plexes with weak organic bases [30, 55, 90, 93]. In a PTC, the uncompleted acid-base in-
teraction is realized (equations (8)–(10)). Herewith, an obligate event is enhancement of
NH-acidic properties of H2P with a chemically active bond in reactions accompanied by
the total proton transfer (equations (2)–(3)), i.e., when the interaction of H2P with a strong
base is realized [61].
What is more, the assessment of the values of proton dissociation constants of NH-ac-
tive porphyrins (pK1 and pK2, equations (2)–(3)) is a good proof for the validity of chem-
ical activity criteria of NH bonds. These criteria can be used on other objects in the absence
of such constants [26]. Table 20 presents the values of dissociation constants for a number
of porphyrins with chemically inactive (I–III, XX) and active (IV–X, XXII–XXIII) bonds.
As it follows from the table data, the NH-acidic properties are indeed enhanced in sequenc-
es of compounds, in which an increase of NH activity is observed (Section 1.3). But H2P
nonplanar macrocycles, which possess no NH activity, drop out of this sequence and behave
similar to planar compounds I–III. Thus, for nonplanar [142] H(N-Me)TPP (XX), the other
way round, the NH activity is observed to be decreased almost 10-fold as compared with
unsubstituted tetraphenylporphin [61] (Table 20). Thus, the rise of chemical activity of NH
bonds in H2P molecules increases the NH acidity of compounds in a cardinal way. For in-
stance, the acidity of nonplanar NH-active compounds can be enhanced by up to 13 orders,
and of planar ones, up to 15 orders as compared with classical unsubstituted porphin ac-
cording to the values of pK1 (equation (2), Table 20).
Table 20 Constants of acid dissociation of porphyrins in DMSO–KOH [222] and

DMSO–NR4OH systems.
Porphyrin pK1 pK2 Reference
H2(β-Br)4TAP (IX) 7.26 ±0.02 7.96±0.02 35, 53

H2Pc (XXII) 10.73± 0.03 – 35, 61, 155
H2(β–Ph)8TAP (XXIII) 13.73 ±0.03(a) – 156
H2TAP (VIII) 12.36 ±0.18 – 61
9.68 ±0.02(b) – 156
H2TBP (X) 18.53±0.05 – 61, 155, 157
H2T(n-Pr)P (III) 23.91±0.02 – 61, 155
H2P 22.35±0.02 – 61, 155, 157
H2TPP (II) 21.15±0.03 – 61, 155
H(N-Me)TPP (XX) 22.07±0.04 – 61, 155
H2TPTBP (IV) 18.42±0.05 – 155
H2(β-Et)8TPP (V) 17.93 17.49 62
H2(β-Ph)8TPP (VI) 12.52±0.02 15.39±0.06 62
H2(β-Br)8TPP (VII) 9.14±0.06 12.90±0.03 62
(a)
Sulfo derivative of H2Pc(SO3– ); (b) solvent DMF.
2.3 Nucleophilic substitution reactions in the coordination core

Porphyrins can be subjected to numerous nucleophilic substitution reactions in the coordi-
nation core to form a number of simple or bridge N-substituted porphyrin analogues [15].
A scheme of one of the simplest substitution processes – at the saturated carbon atom,
namely, alkylation, benzylation etc. – is given in Fig. 25.
regrouping
Figure 25 Schematic reaction of the nucleophilic substitution by H2P molecule coordination core
atoms in saturated carbon atom [15, 85].
Although the substitution process is nucleophilic, by one of the tertiary nitrogen atoms
of the coordination core, progress of the reaction at the regrouping stage depends to a great
extent on the chemical activity of the NH bond.
3 Biosignificance of the Phenomenon of NH Activation

Studies of the phenomenon and conditions of the chemical activation of NH bonds in por-
phyrin molecules are of great importance for understanding the processes occurring in por-
phyrin-containing biosystems. Although the major part of bioporphyrins are attributed to
the group of planar classical porphyrins with a chemically low-active NH bond [28, 58],
selective activation of bonds in these molecules in vivo, nevertheless, can occur. It can pro-
ceed under the influence of the protein-lipid environment, leading to conformational chang-
es of a certain character [17, 19] and, probably, to the polarization of the macrocycle [22,
26]. As we showed above, depending on the type of H2P conformation emerging in solu-
tion, the polarization of the macrocycle and individual bonds in it can occur or not occur at
all [26, 85]. It can be expected that processes in biosystems proceed in accordance with sim-
ilar mechanisms.
Incorporation of metals (Fe, Mg, Co, Ni and others) into molecules of heme precur-
sors, chlorophylls or cytochromes is performed in vivo on the enzymic level [10–11, 14,
27, 258]. Nevertheless, the effect of conformationally initiated activation of NH bonds can
also accelerate metal inclusion processes by several orders, as it occurs in solution [26, 35,
52, 85].
Unfortunately, until now it has not been established in detail in conformations of which
types precursors of bioporphyrins exist and how they structurally change in vivo, as well as
which factors regulate the process of metal inclusion into H2P ligand [14]. It can only be
assumed that their main conformation in this process would be saddled [16, 19], as the result
of the formation of which the synthesis of MP proceeds more efficiently owing to the ac-
cessibility of the reaction centre on the whole and the activation of NH sites in particular
[26].
Thus, studies of the peculiar features of conformational conversions of the porphyrin
macrocycles and related changes in the physics, chemistry and reactivity of compounds has
wide prospects. Thus, for instance, knowledge of the requirements imposed on porphyrins
– molecular-recognition receptors or photosensitizers in photodynamic therapy [110] –
from the point of view of the geometry of conformations, from the positions of the best cor-
respondence of reaction centres of the participants in the process will undoubtedly make it
possible to intensify it significantly.
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6 Synthesis, Structure
Peculiarities and
Biological Properties
of Macroheterocyclic
Compounds
M.K. Islyaikin1, E.A. Danilova1, Yu.V. Romanenko1,

O.G. Khelevina1 and T.N. Lomova2
1Ivanovo
State University of Chemistry and Technology,
7 F. Engels Prospect, Ivanovo, 153000, Russia; email:
islyaikin@isuct.ru
2Institute
of Sciences, Ivanovo, 153045, Russia; email: tnl@isc-ras.ru
Recent advances in the chemistry of macroheterocyclic compounds – structural

analogues of porphyrin and hexaphyrins – are reviewed. Aromaticity of various
macrocyclic molecules and their fragments was studied using the geometric (EN,
GEO and HOMA) and magnetic (NICS) criteria based on experimental data and
results of quantum chemistry calculations at the DFT level. The coordinating and
biological properties of macroheterocyclic compounds as well as of their metal
complexes are under consideration.
Introduction
A conspicuous group of natural macrocyclic pigments among the great diversity of macro-
heterocyclic compounds are hemoglobins, chlorophylls, cytochromes and other species.
They play an exceptional role in vital processes of photosynthesis and respiration, are in-
volved in regulation of fine metabolic processes in living organisms. These substances are
tetrapyrrole macroheterocyclic compounds (porphyrins), the basis of which is porphin 1. It
CRC but not checked

220 M.K. Islyaikin, E.A. Danilova, Yu.V. Romanenko, O.G. Khelevina and T.N. Lomova
has a planar structure, and their inner macroring consisting of 16 carbon and nitrogen atoms
contains 18 π-electrons and is aromatic.
Owing to their unique structure, porphyrins and their metal complexes exhibit quite a
number of interesting properties and find the most broad application in science, technology
and medicine. In particular, they are used as photosensitizers for photodynamic therapy of
cancer [1], as catalysts [2] and electrochemical sensors [3]; are promising materials for non-
linear optics, optoelectronics, liquid-crystal systems [4] etc.
X N
N HN N HN
X N N
X
NH N NH N
X N
1 X = CH; 1a X = N 2
Numerous synthetic derivatives of porphyrins are known, e.g., tetraazaporphin (por-

phyrazine) 1a and tetraazatetrabenzoporphin (phthalocyanine) 2, which are of great practi-
cal significance. In particular, phthalocyanines are deep-blue colour pigments and dyes
unsurpassed for their purity and brightness [5, 6].
The endeavour to expand the colour gamut of phthlanocyanine dyes led to the discov-
ery of a new generation of macroheterocyclic compounds, structural analogues of phthalo-
cyanine, in whose molecule one (3) or two (4) isoindole fragments are substituted by
residues of aromatic diamines,
N
N
R
R
N NH
N N N
N
NH N NH
R
N
N
3 4
where R are aromatic or heteroaromatic cycles.

Perfection of the methods of synthesis, use of various diamines made it possible to ob-
tained macroheterocyclic compounds (Mc), which differ in the number and composition of
small cycles combined into a macrocyclic system. As the result, Mc were synthesized,
which differ in the size of the coordination cavity, composition and nature of atoms in it,
which is of undoubted interest for coordination chemistry. The broadest potentialities of a
CRC but not checked

Synthesis, Structure Peculiarities and Biological Properties of Macroheterocyclic Compounds 221
structural modification of the macrocycle skeleton, which do not rule out the incorporation
of pharmacophore groupings, introduction of substituents by the periphery, as well as metal
atoms into the inner coordination cavity, make this class of compounds rather promising
and attractive for the search of substances with practically valuable properties, in particular,
with potential biological activity.
1 Synthesis, Structure and Properties of the Initial Compounds

The common method of synthesis of symmetric-structure macroheterocyclic compounds 4
is interaction of diamines (A) with phthalodinitrile or its functional derivatives 1,3-diimino-
isoindoline (B) or 1,1-dialkoxy-3-iminoisoindolines (Scheme 1).
A
NH
2H N A
NH2
+ 2 B B B
2
NH
A
Scheme 1
The reaction of three-link BAB products with the functional derivatives of phthalod-
initrile – 1,3-diiminoisoindoline or 1,1-dialkoxy-3-iminoisoindolines – is used to produce
nonsymmetric Mc 3 (Scheme 2).
A A
NH
B B + B B B
NH HN NH
B
Scheme 2
As initial diamines, use is usually made of aromatic five- or six-membered carbo- and
heterocyclic compounds containing amino groups in 1,3-positions. The use of 1,4-, bi- and
polynuclear diamines leads to the formation of Mc with an increased internal cavity [7, 8].
Being included into the macrosystem, cyclic fragments of diamines can support or in-
terrupt macroring conjugation. In this connection, of special interest are five-membered
heterocyclic diamines, e.g., based on 1,2,4-triazole 5–8 and 1,3,4-thiadiazole 9, as they are
heteroanalogues of pyrrole and their presence in the macrosystem can lead to the formation
of an intracyclic conjugation system similar to the porphyrazine system.
Substituted diaminotriazoles – 1(H)-3,5-diamino-1,2,4-triazole 5 (guanazole), 3,5-di-
amino-1-phenyl-1,2,4-triazole 6, as well as 3,5-diamino-1-(α-naphthyl)-1,2,4-triazole 7 –
are obtained by condensation of hydrazine hydrate, phenyl hydrazine chloride or α-naph-
thyl hydrazine, respectively, with dicyandiamide [9, 10].
R 5 R = H; N N
N N 6 R = ɋ6ɇ5;
H2N NH2
H2N NH2 7 R= ɋ10ɇ7 S
N 8 R = C12H25
5-8 9
1-Dodecyl-3,5-diamino-1,2,4-triazole 8 was produced by direct alkylation of initial di-

amine 5 by dodecyl bromide in MeOH in the presence of MeONa [11]. 2,5-Diamino-
1,3,4-thiadiazole 9 is formed in the interaction of dithiourea with H2O2 [12].
Key substances for the synthesis of substituted macroheterocyclic compounds are sub-
stituted o-dinitriles [13].
The main methods of producing substituted phthalodinitriles are dehydration of dia-
mides of substituted phthalic acids, substitution of halogen atoms in o-dihalogen derivatives
for nitrile groups by the Rosemund–von Braun reaction, as well as substitution of mobile
atoms and groups by the reaction of nucleophilic substitution and conversion of substituents
of dinitriles already available in molecules. Methods for producing substituted phthalodini-
triles are reviewed in rather many works. Some of them are presented in [13–16].
Maleodinitrile and its derivatives are also of interest for the synthesis of Mc. However,
in the literature there is almost no information on the synthesis of Mc with pyrrole frag-
ments. At the same time, already the first attempts of using substituted maleodinitriles and
their functional derivatives led to the discovery of new macroheterocyclic compounds [17,
18], which were obtained based on 3,4-di(4-tert-butylphenyl)-2,5-diiminopyrroline [19] 10
and 1,2-dialkylthiomaleodinitriles [20] 11–13.
NH Alk
AlkS CN
NH C4H9 11,
AlkS CN C8H17 12,
NH
10 C12H25 13
Phthalodinitrile and its substituted are rarely used for Mc synthesis directly. Usually,
they are transferred into more reactive alkoxy- or diiminoisoindole derivatives.
The reaction of phthalodinitrile with alcoholates of alkaline metals in an alcoholic me-
dium (Alk = Me, Et, Pr) smoothly proceeds at room temperature [21] in accordance with
Scheme 3.
OAlk OAlk AlkO OAlk

CN
AlkONa AlkOH
N N NH
-AlkONa
CN
NNa NH NH
14 15 16 16a
Scheme 3
Introduction of a substituent into a phthalodinitrile molecule is accompanied by a de-

crease in symmetry, which predetermines the possibility of forming regioisomer alkoxy
compounds: 4(7)- in the case of 3-substituted and 5(6)- for 4-substituted phthalodinitriles.
OAlk Substituents have a significant effect on the reactivity of phthalo-

7
6 1 dinitrile. Thus, substituted phthalodinitriles, containing electron-
R N2 acceptor or weak electron-donor substituents in positions 3 or 4,
5 3 pass rather easily into respective alkoxy compounds already at
4 NH room temperature (e.g., 17–20). In contrast, 3-amino-, 3,6-di-
hydroxy-, 3,6-dimethoxyphthalodinitriles form no alkoxy com-
R pounds under these conditions.
3(6)NO2- 17 There are several points of view on the mechanism of this reaction
3(6)Cl- 18 in the literature. The dominating one is the hypothesis [21] formu-
3(6)Br- 19 lated well back in 1956, in accordance with which phthalodinitrile
4(5)tBu- 20 14 in a strongly polarizing alcoholic medium passes into bipolar
ion 14a (Scheme 4). The latter, by attaching a molecule of alkaline
metal alcoholate, transforms into an isoindolenine derivative 15:
OAlk
CN OAlk
N N
CN Na
N NNa
14 14a 15
Scheme 4
It should be emphasized that the formation of bipolar ion 14a is postulated, and the
role of the “polarizing effect of the medium” is not disclosed. Herewith, no proofs support-
ing this mechanism are given in the literature.
Borodkin [22] studied the interaction of phthalodinitrile with sodium methanolate in
benzene and methanol. He notes that, in the former case, a product of sodium methanolate
addition to one of the nitrile groups is formed, whereas in the reaction in methanol the re-
action product is an isoindolenine derivative. Therefore, the mechanism given in Scheme 4
is, apparently, not realized. It is also not ruled out that the isoindolenine derivative is formed
as the result of the intramolecular cyclization of the monoaddition product. The results of
kinetic studies of the reaction of phthalodinitrile with phenol given in a review [23] are in
favour of such a mechanism.
As phthalodinitrile can be recrystallized from alcohol and other organic solvents,
while addition of alcoholate to its alcoholic solution already at room temperature leads to
the formation of alkoxy compounds, the activating role of alkaline metal becomes evident.
Interaction of phthalodinitrile and its substituted derivatives with nucleophilic re-
agents in the presence of alkaline metal cations was studied using the MNDO quantum
chemical method [24]. The action of cation was found to cause a polarization of the nitrile
group, as the result of which the carbon atom of this grouping acquires a significant positive
charge and becomes a strong electrophilic site. As the result of the nucleophilic attack, a
product of sodium methanolate molecule addition for this group is formed (Fig. 1); after an
intramolecular regrouping, this product turns into an isoindole derivative.
Thus, based on the generalization of the literature data and performed theoretical stud-
ies, the following mechanism of isoindolenine cycle formation appears to be the most prob-
able: formation of the product of alkaline metal alcoholate addition for one of the nitrile
O15 H16
C7
C1
N8
126.4
6A
2.7
C2 Li
C9
N10
Figure 1 A model of the product of addition of LiOH to the phthalodinitrile molecule.
groups of phthalodinitrile followed by the intramolecular cyclization of the addition

product (Scheme 5):
OAlk OAlk
CN
AlkONa NNa N
CN CN
NNa
14 21 15
Scheme 5
The effect of substituents on the direction and rate of this conversion was explained
within the framework of this two-step mechanism [25].
1,3-Diiminoisoindoline and its substituted species are obtained by the interaction of
respective phthalodinitriles in methanol in the presence of sodium methanolate with ammo-
nia [21]. Apparently, this compound is formed via 1-methoxy-3-iminoisoindolenine, which
in the interaction with ammonia is converted into the end product. This reaction mechanism
was considered in [26] using the semiempirical AM1 method. Conformational and tauto-
meric conversions of 1,3-diiminoisoindoline, as well as the influence of substituents and
solvents on the energetics of isomeric transitions was studied in [27]. Analysis of the critical
points of potential energy surface showed the cis-form 22 to be the most stable (Scheme 6).
H H
H H
N N N
NH NH N
N N N
H H H
22 22a 23
∆Hf = 81.24 kcal mol-1 ∆Hf = 83.19 kcal mol-1 ∆Hf = 88.14 lcal mol-1
Scheme 6
Figure 2 Structure of 2-(1-aminoisoindolenine-3-ylidenamino)-5-thioxo-1,3,4-thiadiazole-4-in 24.
Figure 3 Packing of the solvate of 2-(1-aminoisoindolenine-3-ylidenamino)-5-thioxo-1,3,4-thiadi-

azole-4-in 24 in a crystal.
Planar inversion (53 → 53a) proceeds with overcoming a relatively low energy barrier
(~20 kcal·mol –1). For realizing tautomeric conversions (53a–54) as the result of intramo-
lecular transfer, this value is ~70 kcal·mol –1. Calculations performed in “supramolecular”
approximation showed that the prototropic rearrangement with participation of hydroxyl-
containing solvents (MeOH, H2O) was made with lower energy expenses by about 17
kcal·mol –1. The data obtained are qualitatively consistent with ab initio calculations
(6-31G basis) for intermolecular proton transfer in formamidine hydrate [28], where the ac-
tivation energy is also observed to decrease by 20.8 kcal·mol –1 as compared with the in-
tramolecular transfer in formamidine.
Of special interest is the structure of products of condensation of 1,3-diiminoisoindo-
line with amines at one of the imino groups, as they are intermediate substances in Mc syn-
thesis.
The structure of 2-(1-aminoisoindolenine-3-ylideneamino)-5-thioxo-1,3,4-thiadiaz-
ole-4-in 24 was studied using X-ray diffraction analysis [29]. It has been shown that com-
pound 24 is crystallized as the solvate with one molecule of DMFA and water. In the crystal,
the molecule is planar, despite the much shorter intramolecular contact N(1)...S(1) 2.74 Å
(the sum of van der Waals radii 3.34 Å [30]) (Figs. 2, 3).
166 H
N N
B
S S N C
N
∆Hf, kcal/mol
164 NH2
162
0 50 100 150 200

ϕ, deg
Figure 4 Internal-rotation energy profiles for tautomer 24a. The structural formula is given for ϕ =
180°.
The energy profiles of inner rotation of the thiadiazoline (thiadiazole) fragment around
the ordinary bond N(3)-C(9) in molecule 24a were studied using the semiempirical quan-
tum chemical AM1 method. The results of the analysis are given in Fig. 4.
In the configuration space studied, the internal rotation proceeds with overcoming the
low energy barrier ∆∆G = 4.02 kcal·mol –1, which makes it possible to assign this molecule
to structurally nonrigid molecules.
The low activation barrier of internal rotation suggests that not internal but external
factors shall be of determining significance in the stabilization of this or that configuration,
which is what is observed in the crystal (Figs. 2, 3), where form C is stabilized owing to
intermolecular interactions.
bis(1-Imino-3-isoindolinylideneamino)arylenes and azoles (TZP) are important inter-
mediate products in the synthesis of both symmetric- and nonsymmetric-structure Mc
[31–33]. bis(1-Imino-3-isoindolinilydeneamino)arylenes 25, 26 are the most studied of
them.
N X N
NH HN
NH HN
25 X = CH; 26 X = N
Despite the similarity in structure, they exhibit different reactivities. Thus, in the in-
teraction with diiminoisoindoline, compound 25 forms a triisoindolebenzene macrocycle,
but from compound 26 a similar product can not be obtained under the same conditions.
A structural feature of three-link products 25 and 26 is the occurrence of two suffi-
ciently “heavy” isoindole fragments connected with the diamine residue by aza bridges,
which appears to determine the aptitude of the molecules to the formation of nonplanar
Figure 5 Model of complex 25 with DMFA. Figure 6 Model of complex 26 with AlOH.
structures as the result of internal rotation relative to single N–C bonds.

Planar inversion of hydrogen atoms of terminal imino groups, the tautomerism and in-
ternal rotation in molecules of bis(1-imino-3-isoindolinilydeneamino)arylenes (aryl =
1,3-phenylene, 2,6-pyridindiyl) were studied using quantum chemical methods [34]. Anal-
ysis of the critical potential energy surface points showed the internal rotation to proceed
with low energy barriers and contribute to the structural nonrigidity of molecules. Solvation
by aprotonic solvents, such as DMFA (Fig. 5) and, to an even greater degree, complexation
with metals (Fig. 6) were found to stabilize the molecule, without rendering any significant
effect on the electronic structure of terminal imino groups. Besides, the distance N(5)–N(6)
decreases as compared with the initial three-link product. These factors together make it
possible to consider such metal complexes as convenient templates for Mc synthesis.
The most widespread method of producing TZP is by heating aromatic diamine with
1,3-diiminoisoindoline or 1,1-dialkoxy-3-iminoisoindoline in methanol at a temperature of
about 40°C for several hours [31].
Compounds 26, 27–29 are produced in accordance with Scheme 7:
NH N R N
H2N-R-NH2
NH NH HN 26, 27-29
NH NH HN
H3CO OCH3
NH MXn
NH L(X)
N R N
Cl Cl 26a,b; 27a-e;
28a,b,d; 29b
NH . HCl N M N
NH NH HN
H Ph
N N
N N N N
R=
N S
N N
26 27 28 29
M = Zn (a), Al (b); Cu (c); Co (d); Ni (e) Scheme 7

Information on products of TZP interaction with metal salts is rather scarce and of con-
tradictory character. Thus, the work [35] reports the synthesis of complexes of
2,6-bis(1-imino-3-isoindolinilydeneamino)-pyridine with Cu, Ni, Co and Au of 1:1 com-
position. A later paper [36] presents data on the synthesis of copper complexes of TZP
based on 1,3-phenylene, 1,2,4-triazolidene-3,5 and 4-chloro-1,3,5-triazinylidene-2,6,
which represent 2:1 complexes.
The work [37] reports the synthesis of metal complexes 28a,b of 1:1 composition
(Scheme 7), which were further used for template synthesis of ABBB-type Mc.
2 ABBB-type Macroheterocyclic Compounds

Recent years have witnessed an increased interest in noncentrosymmetric analogues of por-
phyrin [38] and phthalocyanine [39, 40]. These compounds exhibit practically valuable
properties: nonlinear optical properties, ability to form ordered monomolecular
Langmuir–Blodgett layers, etc.
A rather elegant approach is when one of the pyrrole or isoindole fragments is substi-
tuted by a residue of aromatic diamine, as the result of which an ABBB-type structure is
formed.
The first representative of this class of Mc
was the triisoindolebenzene macrocycle 30 obtain-
ed by Elvidge and Golden [31] in 1957 by the in-
teraction of 1,3-bis(1-imino-3-isoindolinilydene- N X N
amino)phenylene with 1,3-diiminoisoindoline in
boiling ethanol. N N
H
Similar to phthalocyanine, Mc are capable of
forming complexes with metals. Thus, respective N
N N
metal complexes were first obtained by the inter-
action of a solution of 30 with copper(II) or nick-
el(II) acetates in boiling pyridine or with cobalt(II)
in boiling benzyl alcohol [31]. However, these
compounds were characterized only by the ele- 30 X=CH; 31 X=N
mental analysis data.
The same complexes were studied in [41] us-
ing electron and IR spectroscopy. In particular, it has been shown that complexation is ac-
companied by a batochrome shift of the long-wavelength absorption band, lying at 510 nm
in the spectrum of a solution of metal-free compound 30 in α-chloronaphthalene. It has also
been found that the nature of the metal complex former renders a significant effect on the
value of this shift. Thus, the batochrome shift increases in the sequence Ni < Co < Cu (λmax
= 520, 530, 550 nm, respectively). The work also notes that metal complexes are cations;
however, the nature of counterion (L) is left without discussion.
Later publications [42, 43] take acetate anion in the cases when metal acetates were
used for complexation.
The formation and destruction kinetics of nonsymmetric-structure Mc metal complex-
es is considered in [44, 45]. It has been shown that metal complexes 30a–c are rather stable
compounds and possess a high thermostabilizing activity [46] with respect to polycapra-
mide, and complex 30a (L =AcO) found application in practice as a polymer thermal and
light stabilizer [47] and is manufactured under the name of Stabilin-9.
Finding practically valuable properties inspired investigators to study in greater detail
oooo
L
L N N
N N
N Cu N
N M N NH
N N
N
N N
30a, M=Cu; 30b, M=Ni; 32

30c, M=Co, L=OAc, Cl, OH
the structure of nonsymmetric Mc. Thus, for a complex nonsymmetric-structure Mc with

copper, the work [48] presents structural formula 32, which assumes the absence of coun-
terion. However, in performing the complexation with anhydrous copper(II) chloride under
conditions excluding the presence of moisture, complex 30a was synthesized, which con-
tains chloride anion as a counterion [49]. X-ray electron data [50] are also in favour of
structure 30.
Attempts to use this approach for the synthesis of triisoindolepyridine macrocycle 31
failed to lead to the desired result [31].
Nevertheless, using respective complexes of 2,6-bis(1-imino-3-isoindolinylidene-
amino)pyridine as templates for cyclization with 1,3-diiminoisoindoline, Bamfield and
Mac [35] succeeded in producing complexes of compound 31 with copper(II), nickel(II)
and gold(III) (structures 31a–c).
N N N N N N
NH M N N Au N
N N
N N N N
31a, M = Cu; 31b, M = Ni 31c
These compounds proved to be unstable to the light and to alkaline media, so further
studies in this direction were stopped.
3 Azolophthalocyanines
Of special interest among macroheterocyclic compounds of ABBB type are Mc with azole
fragments because, as compared with tetrapyrrole precursors, they have no centre of sym-
metry during the preservation of the internal macroring of the structure close to that of the
porphyrazine one.
Analysis of the literature [31] as well our own experimental studies have shown that
attempts to obtain Mc with three isoindole fragments and heteroaromatic diamine residue
by the method similar to the above method for the benzenetriisoindole macrocycle fail. Pos-
sible causes are the discrepancy of geometric sizes between the reaction centres of
bis(1-imino-3-isoindolinilydeneamino)heteroarylenes and 1,3-diiminoisoindoline, as well
as the low stability of TZP under reaction conditions [32]. As shown by quantum chemical
calculations presented above, TZP-based metal complexes can be of interest as templates
for the synthesis of nonsymmetric-structure Mc.
Synthesis of metal complexes of azole-containing macroheterocyclic compounds was
done by template condensation of respective TZP metal complexes with phthalodinitrile in
a phenol medium [37] (Scheme 8).
CN
N R N Ln N R N Ln
CN
N M N N M N
PhOH
NH HN N N N
M = Zn (a), Al (b) 33a,b; 34a,b; 35b; 36a,b

N N Ph
N N
N N
N S
N N
133a,b 34a,b 35b 36a,b

Scheme 8
The choice of the condensation medium is explained by that initial TZP metal com-
plexes are well-soluble in phenol. Besides, phthalodinitrile in the interaction with phenol
passes into reactive phenoxy compounds [51]. Phenol, being a weak acid, slows down side
reactions, in particular, formation of phthalocyanines, which, in the case of weakly-soluble
reaction products, are extremely difficult to get rid of.
The EAS of Mc metal complexes 33a,b–36a,b are characterized by sufficiently inten-
sive absorption bands in the region of 500–600 nm; the intensity and position of these bands
are largely dependent on the nature of solvent used for spectral measurements. For instance,
the zinc and aluminium complexes of Mc with a thiazole fragment 34a,b feature three ab-
sorption bands in the region of 500–620 nm, which indicates the pronounced aromatic char-
acter of the macrocycles.
Triazole-containing Mc of ABBB type, called triazolephthalocyanines, are the most
extensively studied to date [52]. The methods of synthesis and the properties of triazoleph-
thalocyanine metal complexes are given in the literature [53].
The major methods of their production are statistical condensation and interaction of
TZP with respective nitrile or its functional derivatives in the presence of nickel acetate in
organic solvents. In the former case, a mixture of substituted 1,3-diiminoisoindoline, 3,5-di-
amino-1,2,4-triazole and nickel acetate in a molar ratio 3:1:1 is heated in organic solvents
(usually in 2-ethoxyethanol, butylonitrile, butanol) to form Mc containing the same substit-
uents in all three isoindole fragments. In the latter case, one succeeds in obtaining
ABB′B-type Mc with different substituents.
Nickel proved to be the most convenient
in the synthesis of this type of Mc; the
N N
successful use of copper salts is also re-
N N ported. Other metals do not lead to the
N
formation of required compounds.
X N Ni N X Recently, Torres et al. succeeded in pro-
ducing metal-free triazolephthalocya-
N nine [54], however the purification of
N N this substance proved very difficult.
There are reports in the literature [53] on
the production of nickel complexes with
substituted triazolenaphthalocyanines
Y of ABBB and ABB′B types.
Triazolephthalocyanines have a con-
37a, b stant dipole moment, are capable of
37a X = OC8H17, Y = NO2, forming ordered Langmuir–Blodgett
37b X = OC8H17, Y = tBu layers [55, 56] possessing semiconduc-
tor properties [57], exhibit interesting
nonlinear optical properties [58, 59], liquid-crystalline properties [60] and possess a high
thermal stability [61].
The use of substituted pyrrolines instead of substituted isoindoles led to the discovery
of a new class of noncentrosymmetric Mc of ABBB type – derivatives of porphyrazine,
called by the authors [18] triazoleporphyrazines. Thus, hexa(4-tert-butylphenyl)triazole-
porphyrazine 38 and its N-butyl-substituted analogue 39 were produced with good yields
by the interaction of 3,4-di(4-tert-butylphenyl)-2,5-diiminopyrroline with 3,5-diamino-
1,2,4-triazole or 1-dodecyl-3,5-diamino-1,2,4-triazole, taken in a molar ratio 3:1, in dried
butyl alcohol (Scheme 9).
R
N N
N N N
NH N N
R H
N N N
3 NH + N N
H 2N N2H
N
NH
38 R = H, 39 R = C12H25.
Scheme 9
Respective complexes 38a–c are synthesized by the interaction of triazoleporphyra-

zine 38 with nickel, copper and cobalt acetates in DMFA at a temperature of 100°C
(Scheme 10).
R R
R N N R N N
N H M(OAc)2 nH2O N
N N N N
N H N N M N
N N N N
R N R R N R
R R R R
1 38a-c
R = 4-tBuPh M = Ni (a), M = Cu (b), M = Co (c)
Scheme 10
The structure of synthesized compounds was established based on the results of ele-
mental analysis, NMR, IR and electron spectroscopy, as well as mass spectrometry.
Comparative analysis of the spectral characteristics of Mc 38, its alkyl-containing an-
alogue 39, as well as metal complexes 38a–c has shown that triazoleporphyrazine 38 exists
in a configuration where the hydrogen atom is localized in position 1 of the triazole cycle.
Similar to porphyrazine 1a, triazoleporphyrazine 40 is the molecule with a complex
multicontour conjugation system. The structure of 40 can be presented by means of three
tautomeric species 40a–c (Scheme 11).
H
N N N N N N
N N N N N N N N N N
N N
H H
N N N N N N N HH N
H H H
N N N N N N N N
N N N N
1a 40a 40b 40c

Scheme 11
The features of the geometric and electronic structure of molecules of porphyrazine

1a, tautomeric species 40a–c, as well as the Ni complex of triazoleporphyrazine 41 were
considered in [62] using the quantum chemical method (DFT B3LYP/6-31G d,p). The re-
sults of the calculations are given in Table 1 and Fig. 7.
Table 1 Total (E(RB+HF-LYP)) and relative (Erel) energies, ionization potential (IP) and dipole
moment (µ) of structures 1a, 40a–c and 41 optimized at the level of DFT B3L YP/6-31G**.
Structure E(RB +HF-LYP), Erel,(a) IP, µ,

a.u. kcal·mol –1 eV D
1a –1053.72663826 – 5.82 0.00

40a –1085.75771329 12.54 6.45 2.66
40b –1085.77012851 4.75 6.48 7.21
40c –1085.77770003 0.00 6.36 6.13
41 –2592.8681607 – 6.41 6.67
(a)For
structures 40a–c.
H 1.309
11 N N
1.328 1.0127.7
3
5
8.
1.38
N N 10
1.381 13
1.4
10
2
1.36
3
1.3 1 107.8
2.1
.3 107.4 128.3 N
11
1 N 58
1.369 113.8 108.1 .362 N 1.
1
5
N1 104.6 N 12
1.010
.2
1.32
20 1 .35 N 2
1.307
127 32
1.8
1.302
.6 5 . 9 .2 1 127.4
1 11 1.47 H
1.3
.7 126 110.7 2
127 .8 1.48
1.485
.
77
6 06
1.350
N N105.4 11
1.3
2
10 109.7 110.1 1
32
6. 6. 06.
1.347
95
1.347
3 0 1
1.
105.8N N 105.1 110 111.6
0
1.47
35
5.6 5 .7 2 H 127.4
10112.6
1 .3
1.010
1.
14
H 112.2
1.026
1.4 12
1.348
1.3
84 12 83 48 8 2.
5.0
25
.3 125.3 1. 4 N 3
1.366
7 N
11
9
1 N .3 111.0 128.1
1. 1
12
87N .38 1.35 1.31

130.4
12
.5
1.3109.9 1
0.
7 106.4
N
8
130
1.45
1.307
10
106.7 106.9 1.313
8.
1.467
1
6
10
8 .2
67
1.360
8.2
1 .4
10
1.349
40a 40b
1.279 1.294
N N N N
3
5
0 5
7. 7.
1.39
1.42
10 10
1 .33 5
1.
111.6 1.326 1 111.0
N 34 102.9 129.0 N N .3 103.0 128.0 N
9 9 61 2
N 0. N 9.
5
9
12 11
1.31
1.31
1.860
9 127.3 4 127.5
1.44 12 1.45
12
1.3
1.3
106.4 0
4. 7. 110.0 9
3
. .
89
77
1.012 08 4 06
1.353
1.365
1.878
N H H N110.9 110 N Ni N105.6 110
8.0 6.9
106.7 110.7
9
7
1.869
1.45 1.45
36
36
0 125.9 4 127.4
1.
1.
12 12
1.328
1.326
2. 0.
64 N 4
76 N 4
N 1 .3 105.3 127.9 N N 1 .3 105.8 127.5 N
1.33 111.2 1.31 110.3
7 9
1.457
1.473
10
10
6.
6.
8
1
1.349 1.352
40c 41
Figure 7 Lengths of bonds (Å) and valence angles (°) in molecules of tautomeric species 40a–c and
Ni complex 41 optimized on the level of DFT B3LYP/6-31G**.
The difference in the values of energy in the sequence 40a > 40b > 40c (Table 1) is
relatively small. Tautomer 40c, the structure of whose internal ring is the most close to that
of the porphyrazine ring, is the most advantageous from the energy point of view.
In the sequence 40a, 40b, 40c the length of the N–N bond of the triazole cycle de-
creases and, in the case of structure 40c, approaches the length of the double bond N=N
(Fig. 7). At the same time, the N–C bonds linking the N–N grouping with the internal mac-
rocycle become longer to make 1.425 Å for 40c. This indicates that the N–N bond in this
structure is significantly isolated from the conjugation system of the macroring. In the case
of complex 41, the length of this bond is 1.294 Å and it occupies an intermediate position
between respective bonds in structures 40b and 40c.
Based on the geometric characteristics (Fig. 7) we [62] calculated the HOMA [63] and
NICS [64, 65] criteria and analyzed the aromaticity of 1a, 40a-c and 41 (Table 2, Fig. 8).
Table 2 EN, GEO and HOMA indices of tautomers 40a –c and Ni complex 41.
N NH N N N N N N
a a a a
N N N N N N N N N
N N N N
H H
N N d N N b d N N b d N H H N b d N Ni N b
H H H
N N N N N
N N N N N N N N
c c c c
1a 40a 40b 40c 41
Structure EN GEO HOMA
Porphyrazine 1a 0.115 0.322 0.562

fragment a, c 0.211 0.230 0.559
fragment b, d 0.265 0.514 0.221
internal cycle 0.022 0.036 0.942
40a 0.122 0.464 0.414
triazole cycle a 0.044 0.034 0.922
fragment b 0.332 0.812 –0.145
fragment c 0.332 0.484 0.184
fragment d 0.316 0.801 –0.117
40b 0.103 0.322 0.567
fragment b, d 0.280 0.154 0.566
fragment c 0.252 0.325 0.422
40c 0.098 0.287 0.615
fragment b, d 0.220 0.253 0.527
fragment c 0.289 0.568 0.143
41 0.080 0.277 0.643
fragment b, d 0.237 0.356 0.408
fragment c 0.241 0.383 0.376
As it follows from the data presented in Table 2, the total aromaticity increases in the
sequence of triazoleporphyrazines 40a < 40b < 40c < 41, which corresponds to the tendency
of bond length equalization in these molecules.
Transition from 40a to tautomers 40b and 40c is observed to be accompanied by an
increase of the total aromaticity against the background of a decrease of the local aromatic-
ity of the triazole cycle. The latter is even due to the fact that part of the triazole cycle in-
cluding atom N-4 and adjacent carbon atoms are involved in the conjugation system of the
H N N N N
-11.6 N N
N N -12.6 -5.60 -5.87
-13.2
N N N N N N N N
N N N N N N
H -3.84 N H
-15.5 -6.90 -14.1 -14.8
N N N N NH HN N Ni N
N N
H H
H
N N N N N N N N N N N N
N N N
1a 40a 40b 40c 41
Figure 8 Calculated NICS values (ppm) for structures 1a, 40a–c, 41.
internal macroring. This leads to the seclusion of the N–N bond, which in the case of struc-
ture 40c acquires a pronounced double character. As the result, the alternation of single and
double bonds in the triazole cycle increases, which is manifested in an increase of index
GEO from 0.034 up to 0.225 in passing from structure 40a to 40c. It should be noted that
the difference in the HOMA values of total and local aromaticity of the triazole nucleus de-
creases in the sequence 40a > 40b > 40c (0.508, 0.320 and 0.042, respectively). Aromaticity
of the internal cycle in this case increases (40c, HOMA = 0.948).
Formation of nickel complex 41 is accompanied with a greater equalization of bonds
and an increase of the HOMA value for the entire molecule up to 0.643. The bonds at the
bridge nitrogen atoms become shorter and equalize with the simultaneous elongation of
C–N bonds, nitrogen atoms of which directly interact with nickel. As the result of these
changes, the aromaticity of the internal macroring (HOMA = 0.913) decreases as compared
with similar characteristics of metal-free structures 40b and 40c.
Analysis of the aromaticity criteria for [18]heteroannulenes designated on respective
structural formulas 40b, 40c and 41 by bold lines (Table 2) shows that the largest value of
HOMA = 0.803 is in structure 40b, in which the conjugation contour passes through the
azo group of the triazole cycle. The respective contour of structure 40c is characterized by
a smaller value, HOMA = 0.753. In the case of metal complex 41 the contour including the
N–N group of triazole appears to be more aromatic (HOMA = 0.756) than the alternative
contour (HOMA = 0.634).
The magnetic criterion NICS (Fig. 8) confirms the tendencies of aromaticity change
in the sequence 1a, 40a–c, 41, revealed using the HOMA criterion.
Thus, 40a is the least aromatic, NICS = –6.90 ppm, whereas 40c appears to be the most
aromatic of the tautomers (NICS = –14.8 ppm). A rather interesting fact was also that the
triazole nucleus successively loses local aromaticity, which is also consistent with HOMA
criteria for this sequence of tautomers. Analysis of aromaticity using the NICS criterion
supports the conclusion that for the efficient participation in the formation of the aromatic
macrocycle, its constituent heterocycles should lose part of their aromaticity, which should
be compensated for by the gain in energy owing to the established conjugation in the macro-
system.
The spectrum of compound 38 given in Fig. 9 is characterized by four intensive ab-
sorption bands. Their intensity decreases with the advance to the long-wavelength region.
The band at 244 nm, which appears to be the result of electronic transitions in the phenyl
nuclei of substituents, is the most intensive. The band at 324 nm, by analogy with tetraaza-
porphin, can be interpreted as a Soret band. The bands in the visible spectrum are the result
of electronic transitions involving highest occupied (HUMO) and lowest unoccupied
(LUMO) molecular orbitals. An intensive band at 417 nm, exhibited in the spectrum of
D
A 1.2 391
1.5 374
1.0 325338
328
421 0.8
1.0 324 535 376
417 0.6 328 472 518 629
527 1 1
0.4 497 606 2
0.5 2
0.2 3
0.0 0.0
300 400 500 600 700 300 400 500 600 700
λ, nm λ, nm
Fig. 9 Fig. 10
Figure 9 Electronic absorption spectra: 1, 38; CHCl3, c = 2.16·10 –5; 2, 39, CHCl3, c = 1.71·10 –5
(mol·l –1).
Figure 10 Electronic absorption spectra: 1, M=Ni, 38a, CH2Cl2, c = 1.71·10 –5; 2, M=Cu, 38b,
CHCl3, c = 2.05·10 –5; 3, M=Co, 38c, CHCl3, c = 1.03·10 –5 (mol·l –1).
substituted porphyrazine [66] as a weak band at 450 nm, apparently, is due by its origin to
the disturbance of symmetry of the molecule owing to the substitution of the pyrrole frag-
ment by the triazole fragment.
The long-wavelength band at 527 nm in substituted triazoleporphyrazine 2 is shifted
hypsochromicly by 139 nm as compared with the band of respective porphyrazine [66].
The shape of absorption spectral bands of 38 and of dodecyl-substituted compound 39
is practically the same (Fig. 9), which indicates the identity of their chromophore systems.
At the same time, in compound 39 the 1H-triazole form is fixed by the presence of the alkyl
substituent in position 1. This fact supports the presence of a hydrogen atom in position 1
of the triazole cycle in the molecule of triazoleporphyrazine 38. The absorption bands in
the case of 39 are slightly shifted towards the long-wavelength side, which is a consequence
of the exhibition of a weak +I effect by the substituent.
Three groups of bands can be singled out in the EAS of complexes 38a–c (Fig. 10):
the long-wavelength absorption band at 602–629 nm and a broadened band at 570 nm,
which is exhibited in the case of compounds 38a and 38b as an inflection; a broad band in
the mid part of the spectrum at 472–518 nm; and two bands in the ultraviolet part of the
spectrum at 325–391 nm. The arrangement and sufficiently high intensity of the
long-wavelength absorption band indicate the aromatic character of metal complexes. The
nature of metal has an influence on the position of the absorption bands; the batochromic
shift of the long-wavelength absorption band increases in the sequence Ni < Co < Cu.
4 State of Triazoleporphyrazines in Proton-donor Media

Study of the state and stability of the macrocycles and their complexes in proton-donor me-
dia is an important issue in the chemistry of porphyrazine, the solution of which makes it
possible to determine the limits of their possible applications, to develop efficient methods
of synthesis and isolation of pure ligands and complexes. The stability of porphyrazines and
their metal complexes in a proton-donor medium is determined by the form of existence of
compounds in this medium (neutral or acidic).
Porphyrazines (H2Pz) are weak multicentre conjugated bases. The number of por-
phyrazine donor sites involved in acid-base interaction, the character of the interaction, as
well as the values of stability constants of acidic species produced depend on the structure
of porphyrazine and the properties of the proton-donor medium. Porphyrazine ligands can
produce acidic species both by the intracyclic and extracyclic (meso-) atoms of nitrogen. In
metal complexes, ϕn orbitals of the intracyclic nitrogen atoms participate in the formation
of bonds with complex-forming metal, therefore, only meso-atoms of nitrogen are involved
in acid-based interaction [67].
Triazoleporphyrazines, which differ from porphyrazines by the substitution of one of
the pyrrole cycles by a triazole residue, have two additional nucleophilic centres in the tri-
azole cycle, which can be considered as possible protonation sites. Besides, the proximity
of these atoms to the internal macroring makes it possible to assume a significant effect of
protonation on the properties of triazoleporphyrazines.
The process of acid-base interaction with participation of porphyrazines is of a com-
plex character [68]. Proton transfer from acid HA to base B proceeds via the stages of form-
ing acidic associate, H associate, ion-ionic associate and totally ionized protonated species:
1
HA + B B…HA B…H…A BH+…A – BH+ + A– (1)
acidic iassot
acidic associate H associate H associate ion-ionic associate protonated
protonatedassociate associate species
The forming acidic species differ one from another by the degree of proton transfer
from the molecule of acid to the donor site and should be spectrally distinguishable. The
final result of acid-base interaction depends on the electronic and geometric structure of acid
or base, as well as on the solvation features of anion A– and cation BH+, which is determined
by the nature of solvent. In media with low ionizing ability, the electron-donor centres of
porphyrazine, by participating in weak acid-base interaction, form H associates and ion-
ionic associates. The total proton transfer is possible only in a strongly ionizing medium.
Interaction of porphyrazines with acids is accompanied with characteristic changes in
the visible range of the electronic absorption spectrum (EAS), which correspond to the for-
mation of various acidic species. A quantitative measure of basicity is the stability constant
of the acidic species (Ks). Porphyrazines are Hammett indicators [67], and the value of Ks
can be determined using the Hammett equation:
pKs = nH0 + log Ii , (2)
where H0 is the Hammett acidity function, Ii = Ci /Ci–1 is the ratio of an ith and (i–1)th acid-
base species being in equilibrium (indicator ratio), n is the number of donor sites involved
in the acid-base interaction at a given stage. The values of acidity functions are known from
the literature [69]. In mixtures of carboxylic acids with organic solvents, for which the H0
functions are not known, one can determine the concentration constants of stability of acidic
species, using a modified Hammett equation:
pKs = –x logCHA + log Ii , (3)
where x is the number of acid molecules involved in acid-base interaction at a given stage.
The values of n and x can be determined as the slope of lines log Ii = f(H0) or log Ii =
f(logCHA).
We investigated the stage and stability of 1H-hexakis(4-tert-butylphenyl)triazolepor-
phyrazine 38 (H2TrPz) in a medium of benzene and 100% acetic acid.
.
H
N N
N N N
N N
H
N N N
38
Introduction of a small amount of acetic acid (0.4 mol·l –1, H0 = 7.5) into a benzene
solution of triazoleporphyrazine leads to a hypsochromic shift of the Q band in the EAS
(Fig. 11). Formation of the acidic species is completed in an acetic acid–H2SO4 –antipyrin
medium (H0 = 3.95). Herewith, the absorption maximum of the Q band is shifted hypso-
chromicly to 473 cm –1.
The stability constant of acidic species 38, as determined by the Hammett equation, is
equal to 4.48±0.18. The number of donor sites involved in acid-base interaction is 0.97,
i.e., ~ 1 (Fig. 12).
Taking into account that acid-base interactions of triazoleporphyrazines occur in a me-
dium with low ionizing ability, it can be concluded that an ion-ionic associate is formed in
this case.
A
1.6
531
1.4 1
2
1.2 3
4
1.0
0.8
0.6
0.4
0.2
0.0
450 500 550 600 650 λ, nm
Figure 11 Change of electronic absorption spectra of triazoleporphyrin 38 in the process of

acid-base interaction in time (benzene, 1; benzene–acetic acid system: 2, H0 = 6.08; 3, H0 = 5.33; 4,
H0 = 4.5).
log I
0.5
0.0
4.0 4.5 5.0 5.5 6.5 6.0 H0
-0.5
-1.0
-1.5
-2.0
Figure 12 Dependence of log I on H0 for the acid-base interaction of triazoleporphyrin 38 in a

benzene–acetic acid medium.
Α
531
1.6
1.4
1.2
1.0
0.8
0.6
0.4
0.2
0.0
450 500 550 600 λ, nm
Figure 13 Change of acid-base interaction of triazoleporphyrin 38 in the process of destruction in

a benzene–acetic acid medium at a temperature of 50°C and H0 = 5.95.
Comparison of the values of pKs for triazoleporphyrazine, tetraazaporphin (1.0) and

octaphenyltetraazaporphin (–1.33) has shown that substitution of the pyrrole cycle by a tri-
azole fragment increases the basicity of the macrocycle. An earlier work [68] has shown
that the first stage of the acid-base interaction of porphyrazines proceeds via one of four
meso-atoms of nitrogen and leads to a batochromic shift of the Q band in the EAS. The high
basicity of the macrocycle of triazoleporphyrazine, the hypsochromic shift of the Q band
of its EAS suggest that the protonation centre is the nitrogen atom of the triazole cycle in
position 4 (or N-3, in accordance with the numeration of Table 2). Quantum chemical stud-
ies we carried out at the DFT level [70] confirm this assumption.
Triazoleporphyrazine proved to be less stable than porphyrazines. In the course of time
at an elevated temperature, it breaks down already in solutions of acetic acid in benzene.
Figure 13 shows a change of the EAS of triazoleporphyrazine in the process of destruction.
313 K
0.07
0.06
log C0H2TrPz / CH2TrPz

0.05
323 K
0.04
0.03 333 K
0.02
0.01
0.00
0 10 20 30 40 50 τ, min
Figure 14 Dependence of log (C 0H2TrPz / CH2TrPz) on time τ for the destruction reaction of triazole-
porphyrin in a benzene–acetic acid medium at temperatures 313 K, 323 K, 333 K and H0 = 4.87.
log ke 333
-4.0
-4.1 323
-4.2
-4.3
313
-4.4
-4.5
-4.6
1.10 1.12 1.14 1.16 1.18 1.20 1.22 log CCH3COOH
Figure 15 Dependence of log keff on log C CH3COOH for the destruction reaction of triazoleporphy-
rin 38.
The stability of the macrocycle in solutions is assessed by kinetic stability, which is

characterized by the rate constant of its destruction. We studied the kinetic stability of tri-
azoleporphyrazine in protonated form in solutions of acetic acid in benzene with acidity
functions H0 equal to 5.32, 5.17, 4.87, 4.80 and 4.60, within the temperature range of
313–333 K. Kinetic measurements were carried out at a large excess of acetic acid, i.e., un-
der conditions of the pseudofirst-order reaction. The first order for triazoleporphyrazine is
confirmed by linear dependences of log(C 0H2TrPz /CH2TrPz) on reaction time (Fig. 14).
The effective reaction rate constants were calculated by equation (4):
k = 1/τ ln(C 0H2TrPz /CH2TrPz). (4)
Table 3 presents the kinetic parameters of the triazoleporphyrazine destruction

reaction. The experimental data show that the rate of destruction increases with the increase
of the concentration of acetic acid. The dependences logkeff = f(logCAcOH) are of linear
character (Fig. 15) with the slope close to 2, i.e., the reaction is of second order with respect
to the concentration of acetic acid.
Table 3 Kinetic parameters of destruction of triazoleporphyrin 38 (CH02TrPz = l·10–6 mol/l).
CCH3COOH, H0 T, K keff ·105, s–1 E, ∆S #,

mol·l–1 kJ·mol–1 J · mol–1 · K–1
13.10 5.32 313 2.00±0.10 37±2 –224±2

323 3.15±0.15
333 4.90±0.18
13.98 5.17 313 2.44±0.10 37±2 –223±3
323 3.85±0.11
333 5.95±0.14
14.85 4.88 313 3.21±0.12 39±3 –215±2
323 5.20±0.20
333 8.20±0.26
15.72 4.80 313 3.50±0.10 39±2 –215±4
323 5.71±0.45
333 9.00±0.14
16.60 4.60 313 4.25±0.20 39±4 –213±2
323 6.80±0.20
333 10.60±0.40
Destruction of porphyrazines can be protolytic and acidolytic under the action of acid
molecules, or a more complex solvoprotolytic process under the action of solvated proton.
Destruction of porphyrazines in aqueous solutions of H2SO4 is hydrolytic [71], i.e., pro-
ceeds under the action of hydroxonium ions H3O+. To split the macrocycle, the system must
in all cases be destabilized by two H3O+ ions. Destruction of triazoleporphyrazine in a ben-
zene-acetic acid medium is acidolytic, i.e., proceeds under the action of AcOH molecules.
The following scheme of destruction of the triazoleporphyrazine macrocycle can be
proposed. First there occurs the protonation of nitrogen atom, which was localized in posi-
tion 4 of the triazole cycle (centre N-3 in Table 4); herewith, the aromaticity of the macro-
cycle decreases. In the further successive interaction with two acetic acid molecules, there
occurs the protonation of nitrogen meso-atom. Further, acetate ion attacks the α-carbon
atom of the pyrrole fragment, and the bond C–N in the macrocycle breaks down.
Quantum chemical studies were carried out by the AM1 method to elucidate the most
reactive nucleophilic centres (nitrogen atoms) in the molecule of the monoprotonated spe-
cies of triazoleporphyrazine. Table 4 presents the main thermodynamic characteristics of
diprotonated species, which show that the most probable sites of attack by acetic acid mol-
ecules are nitrogen atoms N-8 and N-10 of the macrocycle (Table 4).
We considered the effect of metal on the basicity of the triazoleporphyrazine macro-
cycle by the example of acid-base interaction of Cu(II) and Ni(II) complexes with 1H-hexa-
kis(4-tert-butyl)triazoleporphyrazine 1 and 2, of a nickel(II) complex with 3,4-di(4-tert-bu-
tylphenyl)dibenzotriazoleporphyrazine 3 in benzene–AcOH and dichloromethane–AcOH
media, as well as of a Ni(II) complex with [4-(n-triphenylmethylphenoxy)] 7,8:12,13:17,
18-tribenzotriazoleporphyrazine 4 in a dichloromethane–AcOH medim.
Table 4 Major thermodynamic characteristics of diprotonated species (AM1 method, 298 K).
2 1 H
N N
10 a 4
N N
N
3
. 2H
d N 9 5 N b
H
N N7 N
8 c 6
Position of ∆Hf, ∆S, ∆G, ∆∆Hf,

protons kcal·mol –1 kcal·mol –1 ·deg – 1 kcal·mol – 1 kcal·mol –1
3,2 730.64 126.1 693.06 –72.99

3,4 720.08 125.5 682.68 –83.55
3,6 712.93 124.4 675.86 –90.70
3,8 711.58 124.5 674.48 –92.05
3,10 712.27 125.1 674.99 –91.36
3,5 715.61 123.6 678.78 –88.02
3,7 733.39 123.7 696.53 –70.24
3,9 712.68 123.9 675.76 –90.95
∆G – calculated by the formula ∆G = ∆Hf – ∆ST

∆∆Hf – thermal effect calculated for diprotonation: ∆∆Hf = ∆Hf (H2TrPzH22 +) – [Hf (H2TrPzH+) +
∆Hf(H+)], where ∆Hf(H+) = 314.9 kcal·mol –1.
N N N N
N N N N
N N
N Ni N N Cu N
N N N N N N
38a 38b
N N N N
N N N N
N N
N Ni N N Ni N
N N N N
N N
O CPh3
42 43
0.5
628
1
2
0.4 3
4
0.3 5
6
7
0.2
0.1
0.0
400 450 500 550 600 650 700 750 λ, nm
Figure 16 Change of electronic absorption spectra of compound 42 in the process of acid-base

interaction in a dichloromethane–acetic acid medium: 1, in CH2Cl2, 2–7, in CH2Cl2 –AcOH (C AcOH
= 0.176–17.64 mol·l –1).
In the acidification of solutions of complexes in benzene and dichloromethane by a

small amount of 100% acetic acid, as in the case of the triazoleporphyrazine ligand, there
occurs a hypsochromic shift of the Q bands in the EAS of the complexes. Formation of acid-
ic species in a benzene–AcOH medium for compound 38b occurs within the interval of H0
= 6.55–4.61 (CAcOH = 0.40–16.48 mol·l –1; for compound 38a, H0 = 6.60–4.60 (CAcOH =
0.87–16.59 mol·l –1); compound 42, H0 = 6.55–4.60 (CAcOH = 1.74–16.59 mol·l –1)
(Fig. 16).
The thermodynamic constants of stability (pKs) of acidic species of compounds 38a,b,
42 in a benzene–AcOH medium and the concentration stability constants of acidic species
of compounds 42 and 43 in a dichloromethane–AcOH medium are presented in Table 5.
Table 5 Stability constants (pKs) of acidic species of triazoleporphyrins.
Compound Medium pKs
H2TrPz 38 benzene-ACOH 4.48±0.18

38a benzene-ACOH 6.22±0.03
38b benzene-ACOH 6.50±0.04
42 benzene-ACOH 6.45±0.05
42 dichloromethane-ACOH 0.43±0.04
43 dichloromethane-ACOH 0.27±0.07
Based on the dependences logIi = f(H0) and logIi = f(logCAcOH), it has been found
that one proton is involved in acid-base interaction (the slope of these lines in a
benzene–AcOH medium for compounds 38a is 0.89; 38b, 1.14; 42, 1.07; and in a
dichloromethane–AcOH medium for compound 42, 0.80; for 43, 0.82) (Figs. 17, 18).
The high basicity of the macrocycle of triazoleporphyrazine complexes, a hypsochro-
mic shift of the Q band in the EAS suggest that the protonation centre is a nitrogen atom in
the pyrrole cycle in position 1. Quantum chemical studies of protonated species of triazole-
porphyrazine we recently carried out [70] support this suggestion.
log I
0.9
0.6
0.3
0.0
5.4 5.7 6.0 6.3 6.6 6.9 H0
-0.3
Figure 17 Dependence of log I on H0 for the acid-base interaction of compounds 38a,b and 42 in a
benzene–acetic acid medium: °, compound 38a; •, compound 38a; S , compound 42.
log I
0.9
0.6
0.3
0.0
-1.0 -0.5 ,0.0 0.5 1.0 log CCH3COOH
-0.3
-0.6
Figure 18 Dependence of log I on log C AcOH for the acid-base interaction of compounds 42 and 43
in a dichloromethane–acetic acid medium: S , compound 42; •, compound 43.
Experimental data show that nickel and copper complexes are more basic than the
ligand; and the copper complex is more basic than the nickel complex. The presence of the
triphenylmethylphenoxy substituent in the molecule of triazoleporphyrazine decreases the
basicity of the macrocycle.
5 ABAB-type Macroheterocyclic Compounds

The first representative of this vast class of Mc (2+2 or ABAB type) 4 was the symmetric
pyridine macrocycle [72, 73] 44, which Campbel [73] called hemiporphyrazine. Subse-
quently, this term was extended to the entire class of Mc 4. Torres [33] proposed to add the
name of residue R as a prefix to the word hemiporphyrazine. For instance, compound 120
can be called pyridinohemiporphyrazine (HpH2).
Methods of synthesis and properties of ABAB-type Mc and their metal complexes are
considered in detail in a review [33].
N
N NH
N
N
NH
N
N
44
The structure of compound 44, as well as of its complexes with Ni 44a, Cu, Co, Mn,
Zn and Ge, was studied using the X-ray diffraction analysis [74–79].
Figure 19 Structure of 44a.
It has been found that the molecule of symmetric macroheterocyclic compound 44,
constructed from isoindole and pyridine nuclei, is in a solid state capable of acquiring both
the planar and nonplanar configuration depending on the nature of complex-forming metal,
as well as on the presence or absence of solvated molecules of solvent.
Hecht and Luger [75] present information on the studies of symmetric benzene mac-
roheterocyclic compound 45 by the XDA method. It has been shown that in the crystalline
state the macrocycle exists as a solvate, which includes molecules of ethanol and water and
has a nonplanar structure.
N
NH
N N
NH
N
45
Spectral studies of a symmetric-structure tert-butyl-substituted benzene Mc we have

carried out in 1981 using 1H NMR spectroscopy [80] show that this substance in solution
can form stable complexes with a solvent, in particular, with DMFA. However, their struc-
ture remained unknown.
N(1s)
O(1s)
N(3)
N(5)
O(2s)
N(2s)
45a 45b
Figure 20 Structure of complexes 45a,b according to the X-ray diffraction analysis data. (Hydro-
gen atoms at carbon atoms are omitted for clarity.)
In this connection, we [81] carried out an X-ray study of complexes of dibenzenehemi-

porphyrazine with DMFA. Monocrystals suitable for X-ray studies were obtained by slow
diffusion of water vapours into a solution of 45 in DMFA.
It was shown that 45 formed complexes with DMFA of composition 1:1 45a and 1:2
45b, in which the macrocycle has a saddled shape (Fig. 20).
In the case of 1:1 complexes, exocyclic atoms of nitrogen N(2), N(3), N(5), N(6) lie
in one plane to an accuracy of 0.04 Å.
Benzene rings and isoindole fragments deviate from the midplane of the molecule to
the opposite sides. This conformation of the macrocycle is, probably, due to abridged in-
tramolecular contacts H(1N)…C(28) 2.72 Å (the sum of van der Waals radii [71] 2.87 Å),
H(1N)…C(10) 2.75 Å, H(1N)…C(9) 2.77 Å, H(1N)…C(27) 2.77 Å, H(4N)…C(10) 2.68
Å, H(4N)…C(11) 2.74 Å, H(4N)…C(23) 2.79 Å, H(4N)…C(28) 2.69 Å.
The macrocycle forms an oval cavity, whose size is determined by the distances
H(10)…H(28) 3.33 Å and H(1N)…H(4N) 2.64 Å. Owing to this, the DMFA molecule is
above the macrocycle, thus forming three-centre hydrogen bonds N(1)-H(1N)…O(1S) 2.14
Å (angle N-H…O 160°) and N(4)-H(4N)…O(1S) 2.17 Å (angle N-H…O 152°).
Addition of one more DMFA molecule (complex 45b of composition 1:2) leads to a
significant change of conformation of the macrocycle (Fig. 20). Exocyclic atoms do not lie
in one plane. The angle between the lines N(2)...N(6) and N(3)...N(5) is 5.8°. In compound
45b, one molecule of DMFA occupies the position similar to that found for a complex of
composition 1:1, thus forming three-centre hydrogen bonds N(1)-H(1N)…O(1S) 1.98 Å
(angle N-H…O 172°) and N(4)-H(4N)…O(1S) 2.12 Å (angle N-H…O 166°).
The second molecule of the solvent is on the opposite side of the macrocycle and is
linked with it by the weak intermolecular hydrogen bond C(10)-H(10)...O(2S) 2.24 Å (an-
gle C-H…O 158°).
In crystalline state, intermolecular interactions have a strong influence on the structure
of macromolecules, thus distorting the manifestation of internal factors. An idea of the
structure of compounds in isolated state is given by quantum chemical calculations.
In this connection, we carried out a theoretical study of the structural features of Mc
45 using the semiempirical method AM1 for the planar 45PL (D2h) and nonplanar 45A
(C2v), 45B (C2h) configurations and crystalline solvate 45*DMF, as well as the transitory
45PL, D2h, ∆Hf = 259.8 kcal·mol –1 45A, C2h, ∆Hf = 250.4 kcal·mol –1
45B, C2v, ∆Hf = 242.5 kcal·mol –1 45*DMF, C1, ∆Hf = 200.0 kcal·mol –1
Figure 21 Spatial structure of symmetric benzene Mc 45 according to the data of quantum chemical
calculations by the AM1 method.
state 45TS between configurations 45A–45B.

Models of molecules, as well as the calculated values of the heats of formation of pla-
nar 45PL (D2h) configuration and nonplanar configurations 45A,B and solvate 45*DMF
are given in Fig. 21.
In the case of planar configuration 45PL (D2h), hydrogen atoms in the internal coor-
dination sphere are spatially close. In particular, the distance between hydrogen atoms of
benzene nuclei is 2.17 Å, and between hydrogen atoms of imino groups 2.59 Å. The spatial
closeness of atoms in the internal sphere leads to their strong mutual repulsion. The mole-
cule is stabilized as the result of its removal from the plane of the benzene and isoindole
fragments.
Two stereoisomers were found which conformed to the potential energy surface min-
ima: chair 45A (C2h) and saddle 45B (C2v). The formation energy gain is 9.4 and 17.3
kcal·mol –1, respectively, as compared with the planar configuration 45PL, symmetry D2h.
Thus, the most energetically advantageous is isomer 45A with the saddled configura-
tion, in which benzene nuclei are removed from the plane, determined by the exocyclic ni-
trogen atoms, by 42.8°. The isoindole fragments are inclined to the opposite side by an angle
of –21.2°. The same configuration is fixed in solvate 45*DMF, which is consistent with
the XDA data for 45a.
The calculated value of the energy barrier between conformers 45A => 45B is equal
to 0.6 kcal·mol –1, and for the inverse transition this characteristic corresponds to 8.5
kcal·mol –1. The so small conformation-transition energies commensurable with the energy
of the hydrogen bond suggest that similar conversions can occur in solvents. Herewith, de-
pending on the nature of the solvent, one or another conformation can be stabilized. This
conclusion drew confirmation from a recently published work [82], which, using tert-
butyl-substituted Mc as an example, considered the effect of solvents by means of the 1H
NMR method.
Thus, symmetric-structure Mc are structurally nonrigid molecules capable of existing
as various conformers separated by low activation barriers. The predominant content of this
or that species in crystalline state and in solution is determined by the character of intermo-
lecular interactions, the significant of which is specific solvation.
It should be noted that unsubstituted Mc are hard to dissolve in organic solvents. How-
ever, the solubility of Mc can be significantly increased by introduction of bulky substitu-
ents, e.g., tert-butyl groups.
Symmetric-structure tert-butyl-substituted Mc 46–50 were synthesized in accordance
with Scheme 19 by the interaction of 4-tert-butylphthalodinitrile or respective alkoxy- or
three-link compounds with aromatic diamines in ethylene glycol or butanol. It proved the
most expedient to synthesize compounds 46–50 from 4-tert-butylphthalodinitrile via re-
spective alkoxy compounds, without isolating the latter from the stock (Scheme 12).
OAlk N R N
CN AlkONa,
AlkOH R(NH2)2 R(NH2)2
N NH HN
CN
NH NH HN
R(NH2)2 R(NH2)2
N R N N R N Ln
M(CH3COO)2
NH HN N M N
N R N N R N
46 – 50 46a-c – 50a-c M = Cu (a);
Co (b);
N NH N N Ph N N (1-Nph ) Ni (c)
R=
N N N N
46 47 48 49 50
Scheme 12
At the introduction of tert-butyl groups, the character of Mc electronic spectra remains
almost the same. Usually, a small long-wavelength shift of the bands is observed, with ab-
sorption in the region of 300–450 nm preserved.
The experimental values obtained by means of NMR spectroscopy are consistent with
these data. Proton signals of imino groups are in the region of 10.2–12.6 ppm. The presence
of signals of intracyclic protons in a weak field is experimental proof of the absence of a
unified macrocyclic conjugation system in compounds 46–50.
The conclusion of the absence of a unified aromatic system in the macrocycles is con-
sistent with the results obtained in studies of a germanium complex of the symmetric pyri-
dine macroheterocycle by 1H NMR spectroscopy [84]. A later work [85] registered signals
of intracyclic protons in a weak field at 15.7 and 15.2 ppm (CDCl3) in an 1H NMR spectrum
of symmetric-structure tert-butyl-substituted Mc with fragments of 1-dodecyl-1,2,4-triaz-
ole containing two nitrile groups in one of the isoindole fragments.
Respective metal complexes were obtained by the interaction of compounds 46–50

with excess anhydrous acetates of respective metals in butanol (Scheme 12) [83]. The ther-
mal stability of the compounds increases in complexation. The complex of macroheterocy-
clic compound with fragments of 1-phenyl-1,2,4-triazole 49a with copper [83] proved to
be the most thermally stable; the maximal exo effect (depletion of mass, 86%) in air for it
is observed at 602°C.
Introduction of tert-butyl groups leads to a decrease of stability of Mc in proton-donor
media [86]. This, apparently, can be explained by that the introduction of two tert-butyl
groups possessing a pronounced +I effect into the Mc molecule is accompanied by an in-
crease of electronic density on exocyclic atoms of nitrogen, which facilitates protonation
and subsequent breakdown of molecules.
In complexation, stability of tert-butyl-substituted compounds increases by approxi-
mately an order of magnitude. While for compound 49 the efficient rate constant of destruc-
tion is equal to 0.11×10 –3 s –1, for its complex with copper this value is 0.47×10 –4 s –1 [86].
Complexation is accompanied by a decrease of solubility of tert-butyl-substituted Mc
in organic solvents.
6 ABABAB-type Macroheterocyclic Compounds

A macroheterocyclic compound based on 2,5-diamino-1,3,4-thiadiazole was first synthe-
sized in 1970s [87, 88]. By analogy with macrocycles known at the time, that compound
was presented as a symmetric ABAB-type compound. However, studies performed simul-
taneously and independently by two research groups [17, 89] showed it to be a new class
of ABABAB-type compounds, the basis of the structure of which is a macrocyclic system
consisting of six alternating substituted pyrrole and 1,3,4-thiadiazole fragments linked by
aza bridges. These systems have a formally conjugated internal macroring containing 30
π-electrons, which conforms to the Hückel’s rule.
tert-Butyl-substituted macroheterocyclic compound 51 was synthesized by the inter-
action of equimolecular amounts of 2,5-diamino-1,3,4-thiadiazole and 4-tert-butylphthalo-
dinitrile in phenol, or 5-tert-butyl-1,3-diiminoisoindoline in 2-ethoxyethanol (Scheme 13).
The presence of bulky tert-butyl groups in the molecule was a cause of a sufficiently good
solubility of 51 in organic solvents, which enabled its chromatographic purification.
CN
N N
3 PhOH N
3
CN H2N NH2 S
S N
N N N
N H
N
N H S
N
NH N H
N N N
EtOEtOH N
N N S
3 NH 3 N
H 2N NH2
S
NH
Scheme 13
Condensation of equimolecular amounts of 3,4-bis(4-tert-butylphenyl)pyrroline-

2,5-diimine with 2,5-diamino-1,3,4-thiadiazole during the boiling of the stock for 24 h
yielded compound 52 with an increased coordination cavity; the compound incorporates
substituted pyrrole fragments into the macrosystem [90] (Scheme 14).
N N N
NH S H S
N N
N N i N N
3 NH + 3 N N
H2N S NH2 H H
NH N N
N N
N S N
52
Scheme 14
The structure of Mc was established using the methods of electron, IR, 1H and 13C
NMR spectroscopy, as well as mass spectrometry data.
Thus, the PMR spectrum of compound 51 measured in deuterated chloroform, along
with signals in the region of 1.40–1.25 ppm, which characterize absorption of protons of
tert-butyl groups, and signals in the region of 7.94–7.61 ppm, which are evoked by absorp-
tion of protons of benzene nuclei of isoindole fragments, yielded a singlet in the region of
12.35 ppm, which vanishes upon addition of deuterated water and can be assigned to the
absorption of protons of imino groups. The location of the latter absorption in the weak field
indicates the nonaromatic character of the macrocycle.
It should be emphasized that under similar conditions, an ABAB-type macrocycle is
formed from 3,5-diamino-1,2,4-triazole. The cause of so significant differences in the struc-
ture of the condensation products is the nature of the thiadiazole fragment. As shown by the
XDA data of 3,5-diamino-1(H)-1,2,4-triazole [91] and 2,5-diamino-1,3,4-thiadiazole [92],
the lengths of S-C bonds in the thiadiazole cycle are much larger than the respective N-C
lengths in the triazole cycle. As the result, the angle between the C-NH2 bonds in the mol-
ecule of 2,5-diamino-1,3,4-thiadiazole proves to be 12.5° larger than that in the molecule
of 3,5-diamino-1(H)-1,2,4-triazole, which predetermines the possibility of forming macro-
heterocyclic compounds of larger size in the case of 2,5-diamino-1,3,4-thiadiazole.
The spectral curves obtained for solutions of Mc 51, 52 (Fig. 22) in chloroform have
an unusual appearance and are practically the same. Two most intensive absorption bands
are observed in the region of 392 and 413 nm for 51 and at 428 and 452 nm for 52, as well
as two bands of average intensity respectively at 463 and 501 nm and at 515 and 552 nm.
The presence of the main absorption in a so short-wavelength region (390–450 nm) indi-
cates the nonaromatic character of the macrocycle.
The features of the electronic and geometric structure of ABABAB-type macrohetero-
cyclic compounds were studied using semiempirical quantum chemical methods [90].
Analysis of the potential rotation energy surface of thiadiazole nuclei has shown that these
1.5
D
S
N 392 413
N
N N N
N H
N 428 452 S
N
N H S N
N N N N
N N H
H
1 N N
N
N
N H
N
S
S N
N
N H
N N N
N
S
N
0.5
0
250 350 450 550 650
Wavelength, nm
Figure 22 Electronic absorption spectra (solvent, CHCl3) of 51, c = 2.63 g-mol·l –1; of 52, c =
1.65·10 –5 g-mol·l –1.
Figure 23 A model of Mc 51 (the geometric parameters optimized by the AM1 method).
molecules are structurally nonrigid. The most preferable configurations are nonplanar ones;
in them, sulfur atoms of thiadiazole fragments are oriented outside. Six bridge atoms of ni-
trogen lie in one plane, whereas thiadiazole nuclei as well as pyrrole or isoindole nuclei are
out of this plane. The result is an umbrella-like configuration (Fig. 23).
The so unusual structure of Mc, containing three thiadiazole fragments and three pyr-
role or isoindole fragments, assumes the occurrence of unusual coordination properties as
compared with porphyrazines and phthalocyanines. These Mc proved to be capable of
forming stable complexes of composition 3:1. What is more, these metal complexes could
be synthesized both by the interaction of metal-free compounds with salts of respective met-
als (stepwise synthesis) and by heating a mixture of initial substances in the presence of a
metal salt (template synthesis).
The complexes were synthesized by the interaction of metal-free Mc with metal salts,
taken in a molar ratio of 1:3–5, in DMFA at 100°C. As complex formers, metals were taken
which have relatively small covalent radii (Ni, Cu, Co) and are widely used for the synthesis
of metal complexes based on macroheterocyclic compounds.
3
R 3 R
S N R
N N N N
N
R M
N M N S 3X
N N R N
N N N M N
N N
S N M S S N
N 3X R
N N R
N N
M M
N N
N N
R N S N R
R=tBu, 51a M=Ni, X=Cl (OH) 52a M=Ni, X=OAc (OH)

51b M=Cu, X=OAc (OH) 52b M=Cu, X=OAc (OH)
51c M=Co, X=OAc (OH) 52c M=Co, X=OAC (OH)
Metal complexes 51a–c and 52a–c are formed with good yields. Their structure is
confirmed using mass spectrometry, electron and IR spectroscopy, as well as flame ioniza-
tion spectroscopy. Mass spectra of compound 52a are observed to have peaks correspond-
ing to a complex with three metal atoms (Fig. 24). The isotopic composition of the signals
for the complex obtained completely coincides with that calculated theoretically.
Rather unexpected was also the fact that, besides clusters of signals corresponding to
the structure of compounds with three metal atoms [3+3+3M]+, the mass spectra of the
complexes obtained have intensive peaks corresponding to structures [3+3+3M+O]+, in
which one atom of oxygen is present instead of acetate or chloride anions.
Figure 24 MALDI-TOF. Isotopic distribution of peaks in molecular ions of compound 51a; a,

experimental; b, theoretical.
Complexation leads to a significant change of the spectral pattern – to the disappear-

ance of a characteristic splitting of the most intensive bands in the region of 400 nm, ob-
served in metal-free precursors, and to the appearance of broadened absorption bands.
OR H2N
CN N N
NC SAlk
H2N S NH2
CN
OR NC SAlk
Ni(OAs)2*4H2O BuOH EtOEtOH Ni(OAs)2*4H2O
RO
RS
S N OR SR
N S N
RO N N N N
N
X
Ni
N N N N
Ni N
N Ni X X S RS X N
N S
Ni N N Ni X X N
RO N N N N RS Ni
OR N N N N
S N N
RO S N SR
53 R = C5H11 55 R = C4H9
54 R = C10H21 56 R = C18H17
X = OAc(OH) 57 R = C12H25
X = OAc(OH)
Scheme 15
The method of stepwise synthesis failed to produce ABABAB-type macroheterocyclic

compounds based on 3,6-dialkyloxyphthalodinitriles. Nickel complexes 53, 54 were ob-
tained by the interaction of 2,5-diamino-1,3,4-thiadiazole with 3,6-dipentoxyphthalodini-
trile or 3,6-didecyloxyphthalodinitrile in the presence of nickel acetate in boiling butanol
or amyl alcohol for 20 h. This approach also proved efficient for the synthesis of metal com-
plexes of Mc with pyrrole nuclei 55–57 (Scheme 15).
7 Coordination Properties
Macroheterocyclic compounds of ABABAB type – structural analogues of hexaphyrin
[93] – are interesting objects for studies by coordination chemistry methods. As per the
classification proposed in [94] for monomacrocyclic ligands, compound 58 in accordance
with the type of heteroatoms should be assigned to the group of mixed polythiaazamacro-
cycles with a set of donor atoms SnNm. However, the specific structure, namely the “extra-
cyclic” arrangement of atoms of S and the formation of the internal “cavity” by three pyrrole
and six thiadiazole atoms of N, provides macroheterocyclic compound 52 with properties
of an N-donor ligand (“polyazamacrocycles with donor atoms Nm”).
Lomova et al. [95] were the first to determine the acid-base characteristics and stability
of the ABABAB-type macrocycle to the action of acids by the example of macroheterocy-
clic compound 52 constructed from six successively alternating fragments of thiadiazole
and 3,4-bis(4-tert-butylphenyl)pyrrole linked by six aza bridges (McH3).
Owing to the presence of six 4-tert-butylphenyl substituents, compound (McH3) 52
dissolves in nonpolar organic solvents, thus forming true solutions with a concentration of
(0.1–10)·10 –5 mol·l –1. In transition from neutral organic solvents to individual and mixed
protolytic solvents, the EAS of compound 52 are observed to have noticeable changes,
ooooooo
N N N
N N N N
N H N N
S S
S H S
N N
N N
N N
H H N N
N N
H H
S N N
N N N N
N N N N
S
58 52
which depend on the concentration of acid (Fig. 25). Experimentally, by dilution of solu-
tions with respect to acid, these changes in a benzene–AcOH medium were shown to be
reversible. The protonation process of McH3 in a benzene–AcOH proton-donor medium at
different temperature and acidities (H0 = 4.44–6.61) was studied by the method of spectro-
photometric titration (Fig. 26). It was found experimentally that protonation is a two-stage
process (equations (5) and (6)). Respective protonation constants K1 and K2 were found by
the method of least squares using Microsoft Excel by the mass action law equation (3) made
up for a three-component system with account for the proportionality of concentrations and
optical densities (A0, A∞ and Ap), the neutral and protonated species of McH3 and their equi-
librium mixtures at the working wavelength of 449 nm.
McH3 + AcOH ⇔ (McH3)H+ ·AcO – , (5)
(McH3)H+ + AcOH ⇔ (McH3)H22+ ·2AcO – , (6)
A 1
2
400 500 600 λ, nm
Figure 25 Electronic absorption spectra of McH3 in a benzene–AcOH medium, CAcOH, %: 1, 5; 2,

95; 3, 100. CMcH3 = 1.47·10 –6 mol·l –1.
A
0.08
0.06
0.04
0.02
4.2 5.2 6.2 7.2 H0
Figure 26 A curve of spectrophotometric titration of McH3 in benzene by acetic acid at 298 K, the
first stage; working wavelength, 449 nm.
log I
1.2
tan α = 1
0.6
0
5.5 6 6.5 7
-0.6 H0
-1.2
Figure 27 Dependence of log I on H0 for McH3 in a benzene–acetic acid medium, the first stage
(I = (Ap – A0)/(A∞ – Ap), correlation coefficient 0.99, tan α = 1.15.
Ap − A0
A∞ − A0 1
K= × . (7)
Ap − A0 ⎛ Ap − A0 ⎞
n
1− 0
A∞ − A0 ⎜⎜ H 0 − CMcH3 × A − A ⎟⎟
⎝ ∞ 0 ⎠
Here n is the number of protons attached in one stage. The numerical values of K (Ta-
ble 6) and n, equal to 1 at both stages, were found by optimizing the dependence in coordi-
nates of equation (7) reduced to a linear form: log(Ap – A0)/(A∞ – Ap) – H0 (Fig. 27). The
Table 6 Equilibrium constants K1 and K2 of the McH3 protonation
reaction in the benzene–acetic acid system at various temperatures.
T, K K1, l·mol –1 K2, l·mol – 1
298 0.14
308 0.23
321 0.36 1.3
-ln K1 3
0
3.1 3.2 3.3 3.4
1/T 10-3
Figure 28 Dependence –lnK1 on 1/T for McH3 (benzene–acetic acid).
521 nm
1
2
λ, nm
Figure 29 Electronic absorption spectra of McH3 in 18.1 M H2SO4: 1, initial; 2, in 48 h; 3, after

heating up to 353 K.
thermodynamic characteristics of protonation were determined from the temperature de-

pendence of K (Fig. 28). The thermodynamic parameters of reaction (1) are equal to: ∆H° =
31.6 kJ·mol –1, ∆S° = –90.2 J·mol –1 ·K–1.
Analysis of the values of equilibrium constants (Table 6) and thermodynamic param-
eters shows that compound 52 is a weak base, exceeding amphoteric H2O by only an order
and being by 10 orders weaker than NH3 (pK1 at 298 K are, respectively, equal to 0.85, 1.7
and –9.5 [96]). The protonation reaction of McH3 by the first stage in a benzene medium
is an endothermic process characterized by a not too high negative value of ∆S°. The latter
corresponds to a reaction of formation of charged associated particles more solvated as
compared with initial particles (equation (5)).
In sulfuric acid with concentrations higher than 17 mol ·l–1 at 298 K, we observe a new
protonated species (McH3)Hnn+ (n > 2) with an additional blurred band in the EAS with
λmax 521 nm (Fig. 29), which is confirmed by the shape of the EAS in chloroform after the
reprecipitation of McH3 from sulfuric acid. At concentrations of the acid smaller than
17 mol ·l–1, there occurs the destruction of McH in the course of its dissolution, which is
also confirmed by the analysis of the recrystallization products from the medium of H2SO4.
The low stability of (McH3)Hnn+ does not make it possible to study quantitatively the
1/A 120
1
90
60
2
30
3
0 40 80 120 160
τ, min
Figure 30 Dependence of 1/A on time (τ) for McH3. C 0H2SO4, mol/l: 17.1 (2); 17.4 (1, 3). T, K: 343
(1); 333 (2); 324.2 (3).
protonation equilibrium and to determine how many protons are attached additionally in
changing from AcOH to H2SO4. However, it can be assumed that in a sulfurous solution
for (McH3)Hnn+ n = 3 (for a solution in AcOH, n = 2), because up to 100% H2SO4 the EAS
with the band of 521 nm does not undergo changes, i.e., no new protonated species are ob-
served.
The destruction kinetics of (McH3)Hnn+ was studied quantitatively in H2SO4 with a
concentration of 17.1–18.1 mol ·l–1 at temperatures of 324–343 K. The kinetic parameters
of the destruction reaction are presented in Table 7. The order of the reaction with respect
to the concentrations of McH3 (Fig. 30) and H3O+ were found experimentally (Fig. 31) and
Table 7 Efficient rate constants (kef), energy (E) and entropy (∆S #) of activation of the McH3
destruction reaction in concentrated H2SO4.
CH2SO4, keff ×103, s–1 ·l·mol –1 E, ∆S #,

mol · l – 1 kJ·mol–1 J · mol–1 · K–1
324.2 K 333 K 343 K
17.1 0.50±0.05 3.7±0.4 19.0±0.2 179(a) 234(a)

17.4 0.37±0.02 3.1±0.3 13±1 175(a) 224(a)
18.1 0.080±0.007 0.52±0.05 2.4±0.2 146(a) 121(a)
k×105, s–1 ·l4 ·mol –4

17.1–18.1 168 157
0.189 1.32 5.74
(a) Efficient
values of E and ∆S # at a fixed concentration of H2SO4.
are equal to 2 and 3, respectively. Kinetic equation (8) is treated with account for the run
(in the reaction system) of equilibrium protonation reaction of (McH3)H22+ and water and
the irreversible destruction of the macrocycle (equations (9)–(11)).
–dCMcH3 /dτ = k × C 2McH3 × C 3H3O+, (8)

log [H3O+]
0.7 0.9
-1
1
2
-3
3
-5
tan α = 3
log ke -7
Figure 31 Dependence of log keff of the destruction of McH3 on log[H3O+]. T, K: 343 (1); 333 (2);
324.2 (3).
KK31
(McH3)H22+ + H3O+ (McH3)H33+ + H2O, (9)
KK41
H2O + H2SO4 H3O+ + HSO4– , (10)
(McH3)H33+ + H3O+ + H2So4 → thiadiazole and pyrrole derivatives. (11)
The experimental rate constant k will be expressed as k = k1 × K3 × K4 –1.

With account for the results of studies of McH3 acid-base properties, it was suggested
that in the transitory state of limiting stage (11) McH3, protonated most probably in two
opposite, the most basic, meso-atoms of N and in equilibrium with its triple-protonated spe-
cies (McH3)H33+, is further attacked by proton-donor particles, which are not only H3O+
but also the uncharged H2SO4 molecule. Interaction with the latter is electrostatically more
advantageous for protonated McH3, and its equilibrium concentration is several times high-
er than that of H3O+. The difficulty of splitting the macrocycle conforming to the aroma-
ticity rule is expressed in a very high activation energy (Table 7), which is two times higher
than that for phthalocyanine [97]. Solvation effects are not observed to promote the destruc-
tion process, as far as it can be judged from the positive ∆S # (Table 7). The destruction rate
of MH3 at standard temperature is several orders lower as compared with aromatic phtha-
locyanine. Thus, in 17.1 M H2SO4 keff is, respectively, equal to 0.17·10 –5 s –1 ·l·mol –1
(found by extrapolation by the data of Table 7) and 0.122·10 –3 s –1 ·l·mol –1 [97].
The kinetics of the reaction of MCH3 with nickel cations was studied by the spectro-
photometric method in a DMFA medium at temperatures higher than 348 K, using the
change of the optical density of the solution at the working wavelength of 420 nm. The ef-
fective (experimental) rate constants keff (Table 8) of the reaction carried out at an excess
of the nickel salt Ni(OAc)2 with respect to McH3 were calculated from the first-order equa-
tion (12), in which A0, Aτ, A∞ are the optical densities of the solutions at a working wave-
length at times 0, τ and at the end of the reaction. The choice of parameters of the
coordination reaction is determined by the possibilities of the spectrophotometric determi-
nation of the rate with a satisfactory accuracy (Table 8).
1 A0 − A∞
keff = ln . (12)
τ Aτ − A∞
Table 8 Efficient rate constants of the complexation reaction of nickel(II) with macrocyclic ligand
52 in DMFA(a).
CNiAc2 ×104, T, K kef ×105, s–1

mol · l – 1
1.47 348 4.5±0.1

353 2.2±0.1
358 0.76±0.08
1.58 353 2.5±0.1
358 3.7±0.1
1.70 353 3.4±0.2
358 4.6±0.2
1.87 348 4.1±0.1
353 4.3±0.2
358 5.2±0.3
2.06 348 3.7±0.1
353 7.4±0.4
358 12.0±0.6
(a) Concentration of McH3 did not exceed (2±5) · 10–6 mol · l –1.
The polydentate ligand with an extended coordination cavity and a large amount of
electron-donor heteroatoms – potential coordination cores – features a rather complex ki-
netics of complexation reactions. In the course of conversion, the character of the McH3
absorption spectral curve changes significantly and, according to the data of Fig. 32 and
[98], coincides in the end of the reaction with the spectrum of a complex compound of the
3:1 metal–macrocyclic ligand composition.
The logarithm log keff linearly correlates with the value of log[Ni(OAc)2] (equation
A
0.4 1
0.3
2
0.2
0.1
3
0.0
400 450 500 550 600 λ, nm
Figure 32 Electronic absorption spectra of McH3 in DMFA (1), of the complexation product of
McH3 with Ni(OAc)2 in DMFA (2) and, after isolation from the reaction mixture, in CHCl3 (3).
log ke
-4.0
1
-4.5
-3.81 3.78 3.75 log CNi(OAc)2
Figure 33 Dependence of log keff on logCNi(OAc)2 for the reaction of macrocyclic compound McH3
with Ni(OAc)2 in DMFA. T, K: 353 (1), 358 (2) (ρ equals 0.93 and 0.99, respectively).
(13), Fig. 33) only within a narrow range of concentration of the salt (1.58–1.87)·10 –4
mol·l –1, where the order close to two is observed with respect to [Ni(OAc)2]. Respectively,
only for this interval of reaction mixture concentrations with respect to the nickel salt we
can write down kinetic equation (14). At Csalt close to 1.47·10 –4 and 2.06·10 –4 mol·l –1
the order is, respectively much lower and higher than two.
logkeff = logkeff + nlog[Ni(OAc)2], (13)
–dCMcH3 /dτ = k·CMcH3·Csalt 2. (14)
Experimental kinetic equation (14) is treated with account for the reversible and irre-
versible reactions (equations (15)–(18)), where acetate ion is dropped in writing the formu-
la of the salt):
KK11
(McH3)2 2 McH3, (15)
kK2 1
McH3 + Ni2+ [NiMcH2]+…H+, slow, (16)
KK31
[NiMcH2]+ + Ni2+ [Ni2McH]2+…H+, (17)
kK41
[Ni2McH]2+ + Ni2+ [Ni3Mc]3+ + H+, slow. (18)
The rate of the limiting stage (14) is equal to k4 ·С[Ni2McH]2+…H+ ·СNi2+ or, after ex-
pressing the concentration [Ni2McH]2+…H+ with account of equilibrium (17): k4 ·K3 ·
С[NiMcH2]+…H+ ·(СNi2+)2. The latter coincides with the right-hand side of experimental
equation (18) owing to the equilibrium (19) and a fast irreversible transition of McH3 into
[NiMcH2]+…H+ (equation (16)). Thus, the experimental rate constant is k = k4 ·K3.
The value of K1 is first determined experimentally in this work and calculated by equa-
tion (19): K1 = (1.6±0.18)·105 mol –1 ·l. The equilibrium with constant K1 (equation (15))
proves kinetically significant, because it provides for the presence of McH3 molecules in
DMFA solutions during the complexation in predominant concentrations as compared with
the dimer (McH3)2.
Ap − A0
A∞ − A0 1
K= (19)
Ap − A0 0 Ap − A0
1− Cs − CMcH
A∞ − A0 3 A − A
∞ 0
(in the equation, C0McH3 and Cs are the initial concentrations of, respectively, McH3 and
DMFA; A0, Ap, A∞ are the optical densities at a working wavelength of 420 nm for solutions
of (McH3)2, equilibrium mixture at a definite concentration of DMFA and McH3).
It is characteristic that the second order with respect to the salt is disturbed at the broad-
ening of the salt concentration interval. The assumed scheme of stepwise reactions explains
this regularity. When the consentation of the salt is increased, stages (16) and (17) merge
into one equilibrium stage, which in the expression of the concentration [Ni2McH]2+…H+
leads to the third order with respect to the salt. In contrast, at low CNi2+ the coordination
reaction does not proceed to the end, and stage (16) becomes slow and limits the entire pro-
cess. Respectively, the order with respect to the salt becomes equal to unity.
The high sensitivity of the reaction to the concentration of the salt enables an assump-
tion on the detailed mechanism of limiting stage (18) as a dissociative one according to the
Langford–Gray classification [98]. The temperature dependence of the rate constants is
nonlinear, which does not make it possible to obtain the value of activation energy and, re-
spectively, entropy for the reaction. If the narrow temperature range (353–358 K) is used,
one can obtain the activation energy, regularly decreasing with the salt concentration rise
from 82 down to 25 kJ·mol –1. The value of ∆S # in this case changes from –109 down to
–266 J·(mol·K) –1. Large negative values of activation entropy (–109 to –266
J·(mol·K) –1) indirectly confirm the statement of the dissociative mechanism of reaction
(14), thus indicating a more ordered transitory state of the reacting system as compared with
the initial state. Indeed, the polar transitory state [[Ni2McH]2+ ·[Ni(OAc)2(DMFA)3]] #, as
the state formed from the pentacoordinated nickel complex (the dissociative mechanism of
reaction (14)) and from the macrocyclic ligand polarized by the N–H bond, is, probably,
more solvated in the medium of the main solvent, DMFA.
Thus, most probably, the nickel coordination reaction of the macrocyclic ligand with
an extended coordination cavity McH3 proceeds as follows: the macrocycle is readily co-
ordinated with one nickel ion, the second ion is coordinated with greater difficulty, and the
reaction does not proceed to the end (equilibrium (17) is established), the third nickel ion
is coordinated with even greater difficulty, also, apparently, reversibly, however, due to the
low rate of the direct reaction, no equilibrium is achieved under the conditions of the ex-
periment. The stepwise entry of metal cation into the coordinating space of McH3 opens
prospects for finding conditions of synthesizing heteronuclear complexes with ligands of
similar structure and for their practical applications.
8 Studies of the Practically Valuable Properties of Macroheterocyclic

Compounds and their Metal Complexes
By the time macroheterocyclic compounds were discovered [72, 73], phthalocyanines had
already been sufficiently well studied and found wide use as pigments and dyes [5]. For this
reason, the first studies mainly concerned the coloristic properties of Mc [99, 100]. It is not
hard to note that synthesized Mc were primarily tested for the same practically valuable
properties revealed in phthalocyanines. This is due both to similarities in their structure and
to the fact that macroheterocyclic compounds were, as a rule, studied by the same investi-
gators who studied phthalocyanines. And, though in most cases the presence of such prop-
erties was confirmed, Mc, as a rule, proved less efficient than phthalocyanines [101–106].
A true qualitative breakthrough in the approach to the assessment of the properties of
Mc was the discovery of their ability to catch and deactivate free radicals. A groups of Rus-
sian scientists [46] has found that metal-containing macrocycles possess a thermo- and
light-stabilizing activity, whereas respective phthalocyanines catalyze the destruction of
polycapramide. At present, the copper complex of the triisoindolebenzene macrocycle is
manufactured on a industrial scale under the trade mark of Stabilin-9 [47].
Apparently, the same quality is the basis of stabilization of siloxane rubbers (SKT)
[107], as well as of flame retardation of polyorganosiloxane polymers [108] by metal com-
plex Mc.
The development of new approaches to the synthesis enabled a structural modification
of Mc, which led to the discovery of new practically valuable properties, in particular, liquid
crystalline [60], nonlinear-optical [109], as well as the ability to form ordered
Langmuir–Blodgett layers [55].
Analysis performed has shown that macroheterocyclic compounds and their complex-
es with metals are substances possessing a whole range of practically valuable properties,
the study of which is far from completion.
8.1 Biological properties

One of the factors that impede studies of their properties is the low solubility of unsubsti-
tuted Mc in water and in organic solvents. As we reported above, introduction of bulky sub-
stituents led to a significant increase of solubility and, because of this, opened broad
prospects for studies of the properties, biological properties including, of this class of com-
pounds.
At the first stage, we studied the antimicrobial activity of a number of tert-butyl-sub-
stituted Mc and their metal complexes.
The antimicrobial activity was assessed by the germination of test cultures (Escheri-
chia coli str. 676,0 and Staphylococcus aureus Losmanov) and the diameter of the lysis
zone. tert-Butyl-substituted Mc and their complexes with copper, which include 1,2,4-tri-
azole fragments, proved to possess moderate antimicrobial properties.
A qualitatively new step in studies of the biological properties of Mc was a systemic
approach, which consisted in predicting the range of biological activity with the view to re-
veal promising structures and perform laboratory tests of chosen substances.
Prediction of the kinds of activity for a number of macroheterocyclic compounds, their
metal complexes, as well as initial substances for their synthesis was performed using the
PASS program [110]. A fragment of the calculated spectrum of biological activity is pre-
sented in Table 9.
Table 9 Prediction of the range of biological activity of Mc, their metal complexes and initial
substances for their synthesis.
Compound Coef. Type of activity
Anti- Nucleo- Carcino- Anti- Anti-

tumour phil.reag. genic bacter. viral
exchange
N NH Kconf 14.8 20.8 – 16.9 21.3
H2N N NH2 Keff 2.0 6.1 – 1.8 5.4

I(a)
N N Kconf 30.2 – 1.3 20.7 11.8
H2N S NH2
Keff 4.2 – 3.2 2.2 3.0
II
N NH
N N N
Kconf 15.7 18.9 – – 17.6
NH HN
III Keff 4.0 2.4 – – 5.2

NH HN
N N
N S N Kconf 12.1 – – 25.8 12.3
NH HN
Keff 1.7 – – 9.2 3.1
NH HN IV
N NH
N N N
Kconf 14.8 28.9 – 15.4 6.6
N Cu N
Keff 2.0 8.5 – 5.5 43.7
N N N
V
N NH
N NH
N N N
Kconf 14.8 28.9 – 15.4 2.8
N Co N
Keff 2.0 8.5 – 5.5 3.3
N N N
VI
N NH
Table 9 Prediction
(continued)of the range of biological activity of Mc, their metal complexes and initial
substances for their synthesis.
Compound Coef. Type of activity
Anti- Nucleo- Carcino- Anti- Anti-

tumour phil.reag. genic bacter. viral
exchange
N NH
N N N
Kconf 14.8 28.9 – 15.4 6.6
N Ni N
Keff 2.0 8.5 – 5.5 43.7
N N N
N NH
VII
N N
N S N
Kconf 19.7 21.1 – 25.3 11.8
N Zn N
Keff 1.3 6.2 – 9 4

N N N
VIII
Note: For reading convenience, in this section we use different numeration of compounds.
As it follows from the data presented in Table 9, for all tested compounds one should
expect the manifestation of an antitumour activity, which should decrease in the following
sequence: II > III > I = V = VI = VII > IV > VIII.
In this connection, chemical compounds presented in Table 10 were tested for the an-
titumour activity on a model of transplantable tumour L-1210.
Table 10 Indices of antitumour activity of compounds at repeated intraperitoneal injection of DBA
with L-1210 to female mice.
Compound No Injection Daily dose, Total dose, mg ALE, ALEI,

protocol, mg/kg days %
days
1 3 2 4 5 6
Control 2–3 – – 5.3±0.2 –

I 2–3 150 300 5.4±0.4 1.8
Control 2–6 – – 10.0±0.4 –
I 2–6 50 250 9.8±0.3 –2.5
(continued)
Table 10 Indices of antitumour activity of compounds at repeated intraperitoneal injection of DBA
with L-1210 to female mice.
Compound No Injection Daily dose, Total dose, mg ALE, ALEI,

protocol, mg/kg days %
days
1 3 2 4 5 6
Control 2–3 – – 5.3±0.2 –

II 2–3 150 300 6.0±0.1 13.2
Control 2–6 – – 10.0±0.4 –
II 2–6 50 250 10.5±0.17 5.0
Control 2–6 – – 8.3±0.5 –
III 2–6 250 1250 8.0±0.2 –3.6
Control 2–6 – – 8.3±0.5 –
IV 2–6 250 1250 4.0±0.1.1 –51.8
Control 2–3 – – 5.3±0.2 –
IV 2–3 40 80 4.8±0.1 –10.3
Control 2–6 – – 8.3±0.5 –
V 2–6 125 625 7.0±0.6 –15.6
Control 2–3 – – 5.3±0.2 –
VI 2–3 250 500 3.6±0.2 –32.1
Control 2–6 – – 10.0±0.4 –
VI 2–6 50 250 9.6±0.4 –4.0
Control 2–3 – – 5.3±0.2 –
VII 2–3 250 500 5.8±0.2 9.4
Control 2–3 – – 5.3±0.2 –
VIII 2–6 250 1250 6.6±0.6 –20.5
Control 2–6 – – 8.3±0.5 –
IX(a) 2–6 250 1250 5.4±0.1.3 –34.9
Control 2–3 – – 5.3±0.2 –
IX 2–3 100 200 4.8±0.9 –9.4
(a) IX, macroheterogenic compound of symmetrical structure with fragments of 1,2,4-triazole.
ALE, average life expectancy; ALEI, average life expectancy increase.
Preliminarily, maximum tolerated doses (MTD) were determined for each compound,
based on which daily therapeutic doses were chosen. The values of MTD made it possible
to assign all tested compounds to toxicity class IV (low-toxic compounds) [111].
Experimental studies of the antitumour activity were carried out under conditions of
intraperitoneal introduction of preparations. The results of this part of work are presented
in Table 10.
As it follows from the table, compounds 3,5-diamino-1,2,4-triazole (II) and the nickel
complex of symmetric-structure macroheterocyclic compound with fragments of 3,5-di-
amino-1,2,4-triazole (VII) exhibit a moderate antitumour activity with respect to lymphoid
leukemia L-1210.
In the sequence of triazole-containing compounds studied, two of them (3,5-diamino-
1,2,4-triazole and the nickel complex of Mc with fragments of 1,2,4-triazole) possess a
moderate antitumour activity. By the value of the increase of the average life expectancy,
these compounds can be arranged in the following sequence: I > VII > III > V > VI > IX.
Thus, the antitumour activity of triazole derivatives studied depends both on the struc-
ture of the organic moiety of the molecule and on the nature of metal complex former. In-
troduction of metal into the Mc molecule significantly increases the value of ALEI, thus
weakening the negative effect of descriptors present in the Mc molecule. The nickel com-
plex of Mc VII proved the most efficient in the sequence of all macrocycles and their com-
plexes studied, while the cobalt and copper complexes exhibited no activity with respect to
leukemia L-1210.
In the sequence of 1,3,4-thiadiazole derivatives, no manifestation of antitumour activ-
ity was found for any of the compounds studied, and the TZP with a fragment of 1,3,4-thi-
adiazole IV was even capable of significantly initiating the development of the disease
(Table 10). 2,5-Diamino-1,3,4-thiadiazole I itself exhibited a weak activity with respect to
leukemia L-1210, which is consistent with the literature data [112].
The results obtained served as the basis for continuation of works on the assessment
of the antitumour and antimetastatic activities of compounds II and VII on a solid LLC
(Lewis-lung carcinoma) tumour resistant to all currently known antitumour preparations.
The results of studies are presented in Table 11.
Table 11 Indices of antitumour and antimetastatic activity of compounds in female mice C57BL/6
in repeated injection.
Group Daily dose, Injection Tumour weight, g Number of metastases

mg/kg protocol,
days Mtm % inhibition Mtm MII, %
Control – – 9.16±0.57 0.0 5.0±0.2.2 –

II 50 6–14 9.80±0.43 –7.0 9.3±0.4.9 –59.4
VII 50 6–14 9.91±0.58 –8.2 5.8±0.3.6 –16.0
As indicated by the data presented, the compounds studied exhibited no specific ac-
tivity with respect to LLC tumour.
Thus, of the compounds we studied, only 3,5-diamino-1,2,4-triazole and the nickel
complex of Mc with fragments of 3,5-diamino-1,2,4-triazole exhibited a moderate anti-
tumour activity on an L-1210 lymphoid leukemia model, which gives grounds for directed
synthesis and search for more active compounds among their analogues (1,2,4-triazole de-
rivatives) for subsequent works on the development of antitumour means.
In conclusion, it should be noted that accumulated material is too little to judge on con-
crete structure–property dependences. At the same time, the already available results
enable a statement to be made that Mc are promising compounds for the search of substanc-
es with interesting biological properties.
The work was supported by the grant 05-03-33003a from the Russian Foundation for Basic
Research.
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7 The Photochemical Aspect of
Reactions of Flavonols with
Molecular Oxygen
E.A. Venedictov
Institute of Solution Chemistry, Russian Academy of Sciences,
1 Akademicheskaya Street, Ivanovo, 153045, Russia
email: eav@isc-ras.ru
This chapter presents the results of research into the kinetics of photochemical
reactions of biological pigments and related compounds with molecular oxygen.
Introduction
Flavonoids belong to the most widespread natural compounds [1–10]. Their sources are
mainly higher plants. They also occur in algae, lichens, mosses, fungi, ferns and some rep-
resentatives of insects and microorganisms. They are found in roots, wood, leaves, flowers,
fruits. These compounds are present in various cell organelles of plants. They are revealed
in a broad colour palette of the flora and are represented in a whole range of products of
plant raw material processing (juices, wine, oil and extracts).
Owing to the ability of flavonoids to pigment plant tissues, protection of plants from
the damaging action of solar UV radiation was historically considered to be their most sig-
nificant function [6–12]. Subsequently, this point of view found its experimental proof.
The rise of flavonoid biosynthesis in response to an increase of the level of UV radiation,
and the localization of these compounds mainly in the upper cell layer of plant tissues are
considered at present as an adaptation mechanism of plants to their new habitat conditions
[13–15].
In reality, the physiological functions of flavonoids are more diverse. They are be-
lieved to be involved in electron transfer processes, stimulate plant growth and protection
from harmful biological effects occurring with participation of pathogenic organisms and
insects [1–10, 16].
272 E.A. Venedictov
Making up sometimes up to 5% plant biomass, flavonoids are an essential part of our

diet. For this reason, much attention is given to the studies of their pharmacological prop-
erties [1–10, 17]. In particular, these compounds have been known since long time ago to
have antiischemic, antiinflammatory, fungicidal and anticarcinogenic activities. At present,
they are considered as a broadly accessible and potent means capable of not only preventing
but also curing many kinds of illnesses. There is a strongly pronounced antioxidant activity
of their action behind all the diversity of biological and pharmacological properties of fla-
vonoids [17–20].
It has been no secret since long time ago that active species of oxygen and its metab-
olites are responsible for virtually all negative processes occurring in the living organism
on the cell level [21–28]. They are also the main cause of spoilage of many foodstuffs. This
field of biochemistry has been given special attention; a large body of information has been
accumulated over two recent decades in favour of the ability of flavonoids to inhibit oxida-
tion processes and to reduce the risk of damaging essential biomolecules in stress situations
[26, 34, 35], as well as to react with many forms of active oxygen species [18–20, 26,
29–33].
Along with the antioxidant properties, in recent years flavonoids were noted to possess
a prooxidant activity [20, 36, 37], though the ability of their particular representatives to be
included into photodynamic processes has been known since long ago [9, 38].
Flavonols are of special interest, as according to modern views they are the main com-
ponents of the photoprotector system of plants and the most potent antioxidants.
This review deals with some aspects of the relation between the structure of flavonols
and their photochemical reactions with oxygen. It should be noted that ideas and assump-
tions presented here are far from being those, which could be taken for granted.
1 Structure of Flavonols
Flavonoids form a large family of natural O-heterocyclic colorants, the main structural unit
of which is 2-phenyl-4-benzopyrone (flavan). For this reason, flavonoids can be considered
as its derivatives. One of the classifications [10] divides these compounds into ten main
classes: catechins, leucoanthocyanidins, flavanols, dihydrochalcones, anthocyanidins, fla-
vanonols, flavones, flavonols and aurons. The structural features of some of them are shown
in Fig. 1.
The structural unit of flavonols is flavone. For this reason, these compounds are often
considered as its derivatives. The structural types of some of them are given in Table 1.
A feature of flavones is the presence of a C2C3 double bond conjugated with the
π-electronic system of cycle B. Emerging cross conjugation between the carbonyl group
and cycles A and B [7] determines the character of their electronic absorption spectra.
Flavonols can contain from one up to six free OH groups [1–4, 7, 9]. Compounds with
etherified OH groups (OR) are known to exist, as a rule, at C-3, C-5 and C-7 carbon atoms.
Methyl and acetyl radicals often occur as substituting R groups, as well as residues of car-
bohydrates (glucoses, rhamnoses, galactoses and others). Flavonols containing 3-, 7- and
4′-OH groups are weak acids and show three stages of titration within the range of pKa
6.7–11.6 [8, 32, 39, 40]. Compounds with the 7-OH group possess the maximum acidity,
whereas the acidity of other OH groups is on the level of phenol and is to some extent related
to the structure of molecules. For derivatives of flavonol with ortho-substitution of hydro-
gen atom in cycle B the acidity of the 3-OH group decreases. The cause of this is nonco-
planarity of cycles B and C, which emerges due to steric hindrances and prevents the
The Photochemical Aspect of Reactions of Flavonols with Molecular Oxygen 273
3'
2' 4' H
8 1 B
O 2 5' O
7 1'
A C 3 6' H
6
5 4 OH
H H
O
I II
O O
OH OH
O
III IV
Figure 1 Structural formulas of some representatives of the family of flavonoids: flavone (I), cat-
echine (II), flavonol (III) and anthocyan (IV).
delocalization of excess electron charge on the C2C3 double bond. The 5-OH group is the
most hard to ionize due to the strong intramolecular hydrogen bond formed with its parti-
cipation [8, 39].
Owing to the structural features of flavonols, their 3- and 5-OH groups and the carbo-
nyl atom of oxygen mutually affect one another, as the result of which cyclic unstressed
intramolecular hydrogen bonds can emerge between them (an H bond) [1, 40–43], as
shown schematically in Fig. 2. These bonds are formed in noncoordinating solvents and
Table 1 Structural types of some essential flavones.
Flavone Substituents
3 5 6 7 8 1' 2' 3' 4' 5'
Galangine OH OH H OH H H H H H H
Fisetine OH H H OH H H H OH OH H
Chrysine H OH H OH H H H H H H
Apigenine H OH H OH H H H H OH H
Luteoline H OH H OH H H H OH OH H
Kampferol OH OH H OH H H H H OH H
Morine OH OH H OH H H OH H OH H
Quercetine OH OH H OH H H H OH OH H
Rhamnetine OH OH H M H H H OH OH H
Myricetine OH OH OH H H H H OH OH OH
Rutine P OH OH H H H H OH OH H
M, methoxy group; P, rutinose residue.

274 E.A. Venedictov
O O
O
O O O
H
H
I II
Figure 2 Structure of 3-hydroxyflavone (I) and 5-hydroxyflavone (II) with intramolecular H bonds.
O O
hν
O O
O OH
H
N T
Figure 3 Structure of normal (N) and tautomeric (T) forms of 3-hydroxyflavone.
open partially or totally in proton-donor and proton-acceptor media capable of forming

thermally more stable intermolecular H bonds, as well as, apparently, in self-association of
these compounds in highly concentrated liquid and solid solutions [44]. Formation of in-
tramolecular H bonds explains many properties of flavonols, including photochemical
properties.
Under the action of light, such compounds undergo isomerization [40, 42, 47–69].
Formation of the phototautomeric configuration (Fig. 3) is achieved owing to the total intra-
molecular transfer of proton due to the increase of acidity of 3- or 5-OH groups in excited
state [39].
Theoretical studies predict the formation of an intramolecular H bond between oxygen
atom of the 3-OH group and hydrogen ortho-atom of benzene cycle B of flavonols [45]. A
greater extent of coplanarity of cycles B and C is assumed to be achieved owing to this and,
as a consequence, an enhancement of intramolecular H bond of the 3-OH group and carbo-
nyl atom of oxygen according to the feedback principle [40].
Another important structural element determining the physicochemical properties of
flavonols is cycle B. It is capable of undergoing a turn around the C1′C bond, thus contrib-
uting to a change of the extent of conjugation of π-electron systems of cycles B and C. There
are no reliable data yet on its equilibrium position relative to the plane of the rest of the mol-
ecule. At the same time, theoretical studies show cycle B to be removed from the plane of
cycles A and C [45, 46]. The angle of turn of cycle B is from 16 to 43° [46]. Nevertheless,
this geometry [45] does not completely restrict the conjugation of π-electron systems of cy-
cles B and C. For compounds containing the ortho-substituted cycle B, the advantageous
OH OH
OH OH
HO O HO O
M+n
OH O
OH O OH O
M+[n-1]
I II
M+n
M
O
O
HO O
O
OH O
M+[n-1]
III
Figure 4 Structural images of the coordination compounds of quercetine (I): complexes of the com-
position 1 : 1 (II) and 1 : 2 (III).
conformation due to steric reasons is the strongly nonplanar conformation with all ensuing
consequences. According to theoretical studies [57], the close-to-coplanar conformation of
cycles B and C seems to be achieved only in excited state.
Cycle B plays the role of an electron-density donor with respect to γ-pyronic cycle C.
This is indicated by the results of theoretical studies, which show a high density of electrons
at C-2 carbon atom of flavonols [45]. This is also testified by the data of PMR studies show-
ing an increase of aromaticity of cycle C [57]. Intramolecular charge transfer is enhanced
in excited state, if electron donors are substituents in cycle B [68].
One of the consequences of the structural features of hydroxyflavones is their capabil-
ity of coordination interactions with metal ions [4, 70–72]. Complexation can occur with
participation of carbonyl oxygen and 3-OH or 5-OH group, as well as involving ortho-OH
groups of cycle B. This property is the most pronounced in natural flavonols, which can
have up to three coordination sites (Fig. 4), the filling of which depends on the nature of
metal and specific conditions of the medium.
Complexation of flavonols can occur not only in the main state but also in excited state
[68], both from normal and tautomeric forms, thus enhancing intramolecular charge trans-
fer from cycle B to cycle C, which, in turn, would lead to an increase of the stability of the
complex. The ability of flavonols to bind heavy metal ions is believed to be one of the key
mechanisms of their antioxidant action [16, 18–20, 34].
276 E.A. Venedictov
2 Spectral Luminescent Properties of Flavonols
Flavonols are weakly-coloured compounds, which absorb light in the UV range of the solar
spectrum. Owing to the presence of two cross-conjugated systems, the electronic spectra of
these compounds consist of two groups of close-intensity bands lying in the range of
240–270 and 320–380 nm and related to the electronic transitions of ππ*type in the “ben-
zoyl” (cycle A) and “cinnamoyl” (cycles B and C) fragments of molecules, respectively
[2, 4, 7, 8, 17]. The first group of bands coincides with the mixed range of action of the le-
thal effect of UV radiation on living organisms, which is due to the absorption of proteins
and nucleic acids [23]. The second lies in the region of near UV light considered to have an
indirect damaging effect [23, 73] associated with the photochemical properties of natural
and artificial photosensitizers, which are either products of cell metabolism or get into the
organism with food or drugs.
Electronic spectra bear an imprint of the structural features of flavonols. Here we shall
give only the general ideas of the structure–property and medium–property relationships.
More profound views can be found in [2, 4, 7, 8, 17, 38, 39, 52, 66, 68].
Available data indicate that an increase of the number of OH groups in cycle B induces
a batochromic shift of the long-wavelength band and an increase of its absorption coeffi-
cient. This fact can be explained within the framework of an extended system of conjugated
bonds, including those due to superconjugation with participation of OH groups. As for the
short-wavelength band, the rise of the number of OH or OCH3 groups in this cycle usually
does not lead to a significant change of its position.
The successive increase of the number of these groups in cycle A leads to a similar
effect. However, it is less pronounced.
The spectra of many flavones change at high pH due to the ionization of “acidic” OH
groups. Herewith, a batochromic shift of the maxima of the bands and the redistribution of
their intensities are observed.
The strongest changes in the absorption spectra of flavonols are related to the 3-OH
group. Its elimination is accompanied by a hypsochromic shift of the absorption bands. A
similar change is observed at the substitution of hydrogen atom of this group by methyl rad-
ical. The effect of methylation of OH groups on the absorption spectrum of quercetine is
demonstrated by the data of Table 2.
The effect of a solvent on the spectra of flavonols is expressed to a lower degree [52].
No unequivocal relation between the parameters of the spectra and the nature of a solvent
Table 2 Parameters of the first band in the absorption spectra of quercetine and its derivatives in
different solvents [86].
Flavonoid λ max, nm (ε, cm –1 ·l·mol –1)
pyridine alcohol acetone benzene
Quercetine 379 (21700) 374 (20900) 369 (21000)

3,5,7,3′,4′-Pentamethoxyflavone 339 (18200) 340 (21900) 332 (19000) 330 (19600)
5-Hydroxy-3,7,3′,4′-tetramethoxy- 360 (19100) 352 (21900) 350 (18900) 351 (19200)
flavone
3-Hydroxy-5,7,3′,4′-tetramethoxy- 360 (22300) 367 (20300) 351 (19800) 356 (22000)
flavone
Mg-Quercetine 474 (21230
has been revealed. Nevertheless, it is believed that the effect of the medium reflects effects
of general and specific solvation.
The strongest changes in the absorption spectra of hydroxyflavones are caused by
complexation with metal ions, which leads to a batochromic shift of both bands. Herewith,
the complexation involving the 3-OH group is characterized by a maximal effect (Fig. 6).
Additional complexation via the ortho-site of cycle B (Fig. 4) enhances this effect.
Thus, it is seen that the diversity of flavone structures provides for the maximum effect
of absorption and reflection of solar UV radiation at the most harmful directions of its
action.
In contrast with flavone [42, 67, 69, 74], flavonols possess fluorescence. This is ex-
plained by the fact that in substitution of hydrogen atoms of flavone at C-3 carbon atom by
an OH group the excited triplet nπ* level of the molecular proves much higher than the ex-
cited singlet level of ππ* type and renders no effect on its radiative ability [67, 74].
Fluorescence of flavonols is of dual character. It is considered as a radiation from two
excited singlet states with energies of 69–71 and 53–55 kcal/mol, respectively [39, 40, 50,
56]. Short-wavelength luminescence is determined by the properties of normal molecules,
and long-wavelength luminescence is due to the properties of their tautomers formed as the
result of reversible intramolecular phototransfer of proton from the 3-OH group to carbonyl
oxygen.
Intramolecular proton phototransfer is an essential feature of flavonols. It occurs dur-
ing a time shorter than 8×10–12 s and is controlled by structural factors and, in particular,
specific properties of the medium [40, 51, 52, 56, 59, 64, 66]. Proton phototransfer can be
due both to the properties of singlet and triplet stages depending on the structure of a fla-
vonol. As a rule, it takes place in compounds containing a 5-OH group [57, 63, 69].
Intramolecular proton phototransfer determines the short lifetime of an excited singlet
state of flavonols, thus limiting its chemical potentialities. However, it does not totally sup-
press the interconversion of excited singlet molecules into triplet ones. The latter are char-
acterized by a lifetime of 7.2–7.5 µs and energy of 61–53. 4 kcal/mol [60, 65].
Fluorescence of tautomers of flavonols is characterized by a relatively large lifetime,
which has some dependence on the nature of substituents, their number and substitution site
in cycle B and is from 1.3 up to 4.4 ns [39, 48, 50, 56, 65]. The lifetime of the triplet state
of tautomers depending on the structural features of flavonols is from 9 up to 28 µs [49, 56,
60, 65]. The energy level of this state is within the limits of 31–33 kcal/mol [65].
In contrast to fluorescence [49], oxygen efficiently quenches triplet states of flavonols
[57, 60, 62]. Its removal from solution is an obligatory condition for studies of their prop-
erties. Studies have shown that the rate constants of quenching of triplet molecules by ox-
ygen are close to diffusion rates. In n-heptane, for instance, for the main and tautomeric
forms of unsubstituted flavonols they are 2.1×109 M–1 s–1 and 2.7×109 –3.2×10 9 M–1 s–1
[60, 62], respectively.
Unlike flavonols containing no 5-OH group, quercetine does not luminesce in practice
and gives no long-lived triplet states in direct excitation [57]. One of the causes is a high
probability of internal radiativeless conversion of absorbed energy.
However, formation of a long-lived triplet state of quercetine can be caused by the
triplet–triplet transfer of energy [57]. In this case, sensitizers having a triplet level no less
than 57 kcal/mol transfer their energy to the triplet level of the main form of quercetine with
a rate constant exceeding 1.2×10 9 M–1 s–1. Then there occurs rapid tautomerization of
triplet molecules as the result of intramolecular proton phototransfer from the 5-OH group
to carbonyl oxygen. This is one more significant fact, which indicates that quercetine is
278 E.A. Venedictov
capable of participating in quenching of excited triplet states of proteins and nucleic acids,
whose energy exceeds 57 kcal/mol [75].
The work [57] obtained the characteristics of the triplet state of the tautomer of quer-
cetine. It has an energy within the range of 17.1–24 kcal/mol and lifetime from 12.9 up to
18 µs depending on the solvent. The triplet tautomer of quercetine is quenched by oxygen.
In isopropanol, the quenching rate constant is 1.7×109 M–1 s–1.
Finally, it is necessary to note the luminescent properties of coordination compounds
of flavonols. Compounds with Mg2+, Zn2+, Cd2+, Al3+ and a number of other elements pos-
sess a high yield of fluorescence [68, 70, 71]. These results are interpreted from positions
of limitations imposed by complexation on intramolecular proton phototransfer [68].
Electron-donor substituents in para-position of cycle B increase the yield and lifetime
of fluorescence [68]. The latter is associated with the increase of the probability of transition
from an excited singlet state into the basic one with illumination and a reduction of the prob-
ability of internal radiativeless conversion. Introduction of a heavy atom leads to a reverse
phenomenon of the decrease of the quantum yield and a reduction of the fluorescence life-
time. As the rates of these transitions are relatively small, one should take into account the
possibility of formation of a long-lived triplet state of coordination compounds of flavonols
in absorption of light by them. As will be shown below, these conditions are realized also
in the case of quercetine complexes.
Nevertheless, unlike other flavonols, quercetine attracts attention by its capability of
highly efficient utilization of high-energy solar radiation.
3 Photoproduction of 1O2 (1∆g)

By their photochemical activity, flavonols can be conditionally divided into two groups,
one of which contains compounds only with a 3-OH group, and the other, compounds with
3- and 5-OH groups simultaneously. Flavonols of the first group exhibit a photochemical
activity [76] and enter into photochemical interaction with oxygen [49, 50, 60], undergoing
destruction, whose mechanism was discussed from various mechanistic positions, includ-
ing with attraction of 1O2 [60]. Natural representatives of the second group are photochem-
ically inactive.
Photoformation of 1O2 by compounds of the first group is supported by the results of
experiments on the sensitization of luminescence from 1∆g state of O2 in solution of un-
substituted flavonol [60]. In this case, the quantum yield of 1O2 is 0.18. As dual fluores-
cence of this compound is not quenched by oxygen [49], generation of 1O2 is associated
with the quenching by oxygen of its triplet states.
Unlike unsubstituted flavonol, quercetine does not sensitize photoformation of 1O2
[57]. These data are an additional argument in favour of the absence of photochemical ac-
tivity in this group of compounds [76].
A similar pattern was observed in studies of the photochemical properties of 3-
hydroxy-5,7,3′,4′-tetramethoxy- and 5-hydroxy-3,7,3′,4′-tetramethoxyflavones [77]. At
the same time, 3,5,7,3′,4′-pentamethoxyflavone was shown to be capable of photosensitiz-
ing the formation of 1O2. In this case, the quantum yield of 1O2 is 0.49. This is no wonder,
as it is known to exhibit a high photochemical activity [11, 12] and possess intensive phos-
phorescence, which indicates the formation of an excited triplet state [12]. These data show
that in such molecules as quercetine the photochemical inertness is related to both 3- and
5-OH groups.
Of special interest are the properties of triplet states and their role in the formation of
1
O2. This issue becomes especially topical when we speak of the triplet state of quercetine,
accounting for the special significance of this compound as a photoprotector. Though the
answer to the posed question requires special investigation, some conclusions can be made
based on the consideration of the quenching of triplet states by oxygen within the frame-
work of the formalism of the equilibrium formation of exciplex intermediate [78, 79].
3
S + O2 (3Σg– ) ↔ 1(S … O2) → S + 1O2 (1Σg+ and/or 1∆g),
↔ 3(S … O2) → S + O2,
↔ 5(S … O2).
Here 3S is the triplet molecule of a sensitizer; 1, 3, 5{S … O2 } are the singlet, triplet
and quintet states of the exciplex. In this scheme, the processes of conversion of singlet and
triplet exciplexes are considered to be the most probable, as they are allowed by the spin
selection rules. The theoretical values of statistical balances of these processes are 1/9 and
1/3, respectively. Their real values can be assessed from the ratio kQ /k diff, where kQ and
k diff are the rate constant of quenching of triplet molecules by oxygen and the diffusion rate,
respectively. For triplet states of the normal form of unsubstituted flavonol and its tautomer,
these values in n-heptane are 1/7 and 1/5, respectively. Comparing them with theoretical
values, it can be concluded that the probability of 1O2 formation at the expense of the triplet
state of the normal form is expected to be higher.
For a triplet state of tautomeric quercetine, the ratio kQ /kdiff is equal to only ~1/2 [57].
Evidently, in this case all triplet molecules are quenched by oxygen according to the mech-
anism of internal interconversion. Therefore, this process can be considered within the
framework of the photoprotector action of quercetine as an important link for transforming
the energy of the tautomer’s excited state to thermal energy.
Thus, from the data presented it can be concluded that natural flavonols containing a
5-OH group provide for highly efficient radiativeless dissipation of absorbed solar energy
without forming 1O2.
1
4 O2 Reactions
The first steps in this direction were made in [80], which showed the selective character of
the reaction of flavonols with 1O2, aimed to open the C2C3 double bond of cycle C.
The physical character of quenching 1O2 by quercetine was first demonstrated in [57].
The same work postulated the interaction mechanism determined by the formation of exci-
plex intermediate with reversible partial charge transfer, as it is proposed for other phenols
[25, 81, 82, 93–96]. Herewith, the possibility of not only the above-considered process was
shown, but also of oxidation of quercetine and electron-vibration transfer of energy with
excitation of the vibratory levels of its molecules. Therefore, the total rate of quenching in
this case can be presented by a sum of the rates of three elementary processes:
k t = k q + k r + k E-V,
where k t is the overall quenching rate constant; k q is the physical rate constant determined
by the formation of the exciplex intermediate; k r is the oxidation rate constant of flavonoids;
k E-V, the physical rate constant determined by electron-vibration energy exchange. The re-
sults of measurements and assessments of these constants are given in Tables 3 and 4.
280 E.A. Venedictov
Though for quercetine the value of k E-V is relatively small, it is still comparable with k r in
acetone. Therefore, it is natural that the expediency and reality of such a scheme for other
representatives of this family requires additional consideration and experimental proof.
Electron-vibration energy exchange should be expected exactly for molecules contain-
ing C–H and O–H bonds, as they have the highest vibratory states [57, 90–92]. The rate
constant of this process could not be measured, but yields to numerical assessment on the
basis of the additivity principle [90–92] by the formula
Table 3 Rate constants of interaction of flavonoids with 1O2 in different solvents.
Flavone Solvent kt × 10–5, kr × 10 –5, Reference

M –1 ·s–1 M–1 ·s–1
3-Hydroxyflavone CCl4 0.172 83

Benzene-h6 0.187 83
Acetonitrile 1.05 83
Methanol-h4 2.3 83
Pyridine 750 83
D2O (pD = 3) 2.5 83
D2O (1.0 M NaOH) 1900 83
3-Methoxyflavone CCl4 0.19 83
Benzene-h6 0.15 83
Quercetine Acetone 3.1 0.09 57
Acetone 0.26(a) 85–87
Diethyl ester 1(a)
Methanol-d4 24 8.9(a) 84
Dimethylformamide 53 49(a) 86, 87
Pyridine 156 117(a) 86, 87
Galangine Methanol-d4 12 7.4 84
Kampferol Methanol-d4 7.1 4.8 84
Fisetine Methanol-d4 31 11 84
Rutine Methanol-d4 16 1.1 84
Luteoline Methanol-d4 13 0.18 84
Chrysine Methanol-d4 2.4 0.06 84
Catechine Methanol-d4 58 84
3,5,7,3',4'-Pentamethoxy- Benzene-d6 35 1.4(a) 77, 87, 88
flavone Pyridine 0.5(a) 87
3-Hydroxy-5,7,3',4'-tetra- Benzene-d6 25 1.4(a) 87, 88
methoxyflavone Acetone 15(a) 87
Dimethylformamide 110(a) 87
Pyridine 250(a) 87
5-Hydroxy-5,7,3',4'-tetra- Benzene-d6 20 1.1(a) 87, 88
methoxyflavone Pyridine 0.9(a) 87
(a)The values of k were obtained relative to the reactivity of tetracene and were refined using its
r
values of kr [89] equal to 5.2×107 M–1 ·s–1 (pyridine), 7.7×107 M–1 ·s–1(DMFA), 2.2×107 M–1 ·s–1
(acetone) and 1.4×107 M–1 ·s–1 (benzene).
k E-V = mk C–H, arom + nk C–H, aliph + pk O–H .
Here k C–H, arom = 480 M –1 s–1, kC–H, aliph = 390 and kO–H = 2145 M –1 s–1 are rate constants
of 1O2 quenching by particular C–H (aromatic), C–H (aliphatic) and O–H bonds, respec-
tively [92]; m, n and p are the number of each of these bonds in the molecule. The values
of k E-V for some flavonoids are presented in Table 4. The same table gives the values of
k q. For compounds with a large number of OH groups, k E-V is only a small part of k q, not
exceeding 5%. At the same time, the values of k E-V and k q are commensurable for a whole
range of simplest hydroxyflavones. For this reason, the role of this process in 1O2 quench-
ing for them should not be underestimated.
The values of k q for some flavonoids are given in Table 4 together with their oxidation
potentials. The donor-acceptor nature of 1O2 quenching is confirmed by the dependence be-
tween k q and the oxidation potential, in which the rate constant increases when the oxida-
tion potential decreases. Therefore, this process can be believed to be controlled by charge
transfer and can be considered within the framework of exciplex intermediate formation.
Table 4 Parameters of the elementary reactions of 1O2 physical quenching by some flavonoids and
their oxidation potentials.
Flavone kE-V ×10–4 kq×10 –5(a), Eox(b) , V

M–1 ·s–1 M –1 ·s–1
Quercetine 1.5 15 0.60

Rutine 3 15 0.60
Kampferol 1 2.3 ~0.95
Catechine 1.5 ~58 0.57
(a) from
[84]; (b) from [32].
If we turn to the results of [84], it can be found that in the sequence of flavonols be-
tween the values of k q and k r a correlation takes place, which in methanol is described by
the equation
ln k r = (9.01 ± 0.93) + (0.34 ± 0.07) ln k q , r = 0.961.
Based on this, the following conclusion can be made [97]: exciplex formation is the
common stage of physical and chemical quenching of 1O2.
A study performed within the structure–property framework [84] made it possible to
single out a number of structural features affecting the quenching of 1O2 by flavonols. It
has been found that compounds with the catechol structure of cycle B are stronger quench-
ers (Table 3). The effect of the 3-OH group on the process occurring in methanol-d6 proves
to be relatively weak. This conclusion is supported by the data of an earlier work, which
studied the quenching of 1O2 by unsubstituted flavonol and 3-methoxyflavone in relatively
inert solvents, such as benzene and CCl4 [83].
The works [77, 86, 88] have shown that complete substitution of hydrogen atoms of
OH groups of quercetine by methyl radicals leads to a rise of efficiency of 1O2 quenching.
The presence of a free 3- or 5-OH group leads to an insignificant decrease of the quenching
rate (Table 3). An amplification of 1O2 quenching in these cases as compared with quercet-
ine should be, apparently, ascribed to a decrease of energy expenses for exciplex formation
owing to an increase of the density of π-electrons in molecules of these compounds due to
282 E.A. Venedictov
20
ln kr
10 1
2
0
0 20 40
DN
Figure 5 Dependence of the logarithm of the rate constant of oxidation of quercetine (1) and
3-hydroxy-5,7,3′,4′-tetramethoxyflavone (2) by singlet oxygen on the donor strength of the solvent.
the electron-donor effect of the methoxy groups. Herewith, the greatest contribution is, ap-
parently, made by methylation of ortho-OH groups of cycle B.
According to the data of [83, 86, 87], the efficiency of 1O2 quenching by flavonols ris-
es in highly specific media. Due to the absence of data in the case of elementary reactions
with unsubstituted flavonol, we carried out a simplified consideration of this phenomenon.
In the order of increasing k t, the solvents are in the following sequences: CCl4 ≤ benzene
< acetonitrile < methanol ≤ heavy water (pD = 3) < pyridine < heavy water (1 M NaOH)
for unsubstituted flavonol and acetone < methanol-d4 < N,N-dimethylformamide < pyri-
dine for quercetine, which give no grounds to relate the quenching rate to the polarity of
solvent. At the same time, it is evident that the efficiency of the process increases with the
rise of the ability of the solvent to form a strong intermolecular H bond with the OH group.
This is illustrated by the dependence of k t on its donor ability of its molecules (DN). For
unsubstituted flavonol, this type of dependence has the form:
ln k t = (9.04 ± 0.35) + (0.19 ± 0.02) DN, r = 0.988.
It is also confirmed for quercetine:
ln k t = (9.95 ± 1.60) + (0.21 ± 0.7)DN, r = 0.912.
Noteworthy is the qualitative and quantitative similarity of the correlation equations.

The coefficients at DN, which determine the slopes of the dependences, do not practically
differ from each other, though structurally flavonols differ significantly. This indicates that
1
O2 quenching in both cases experiences practically the same effect of the medium. Despite
the fact that k t is a complex value, for a simplified explanation of this fact it is necessary to
admit that the effect of the solvent is related to the presence, mainly, of the 3-OH group.
To understand the nature of this phenomenon better, we considered the features of the
behavior of quercetine in particular elementary reactions. The rate constant of physical
quenching changes in different media within one order of magnitude. The largest value of
k q was found in pyridine; the smallest, in acetone. Despite the fact that the change of k q
depends in a complex way on the solvent, nevertheless, there is a hint to a relation to its
donor properties. This makes us ascribe the observed effect to the solvation stabilization of
exciplex intermediate, the result of which is additional delocalization of the fractional pos-
itive charge emerging during its formation.
In contrast with k q, the value of k r in different media changes within broader limits.
The value of k r is related to the solvent in the same manner (Fig. 5). Unlike quercetine, ox-
idation of compounds without the 3-OH group experiences practically no effect of the me-
dium (Table 3). Therefore, the observed difference in the effect of the solvent is quite
naturally related to the solvation of the 3-OH group.
Undoubted is a symbateness in the change of k q and k r in these media, which is similar
to that considered above, though is less pronounced. This behaviour of k q and k r, evidently
corresponds to the case when the oxidation rate is controlled by the formation of exciplex
intermediate [98].
Another example is oxidation of 3-hydroxy-5,7,3′,4′-tetramethoxyflavone (Fig. 5).
The observed tendency for flavonols is apparently of general character. Still, it is not diffi-
cult to notice some difference. It is seen that the oxidation of 3-hydroxy-5,7,3′,4′-
tetramethoxyflavone possesses the least pronounced sensitivity to the solvent. To explain
this fact, it is necessary to admit that in this case the emerging cationic site in the formation
of exciplex intermediate is capable of more efficient intramolecular delocalization.
In [80], oxidation of flavonols by 1O2 molecules was demonstrated, unlike flavones,
to affect the C2C3 double bond. Herewith, the main product of oxidation is depside (Fig. 6,
VI). In the literature, two most possible mechanisms of this reaction are discussed (Fig. 6):
one assumes oxidation to proceed as an ene reaction (Fig. 6, III) with concerted addition of
1
O2 [80]; the other postulates the formation of a dioxetane intermediate (Fig. 6, IV) [84].
Based on the above analysis, it can be concluded that the primary intermediate of oxidation
O 1 O O
O2 O
O
O
OH OH O
O
O O H
O
I II III
H
OO
O O
O
I
O O
OH
O O
IV V
O
CO
O
COOH
VI
Figure 6 Hypothetic mechanisms of addition of 1O2 to flavonols.

284 E.A. Venedictov
k, s-1
100000
50000
0
0 0.00002 0.00004
c, mol/l
Figure 7 Effect of the concentration of Mg-quercetine on the rate constant of 1O2 luminescence
decay in pyridine (pulse fluorimetry method [88]; 1O2 photosensitizer, anthracene, λexc = 337 nm).
of flavonols is exciplex (Fig. 6, II). What is more, the difference in the kinetic behaviour of
flavonols and flavone suggests that its formation is related to the involvement of π-electrons
of the C2C3 double bond of cycle C.
Assessing on the whole the ability of flavonols to quench 1O2, it can be concluded that
the nature of the molecular environment is one of the most important factors making it pos-
sible to achieve the high efficiency of the process.
5 Photochemical Properties of Coordination Compounds of Quercetine

With many metal ions in stable degrees of oxidation (Mg 2+, Zn 2+, Cd 2+), quercetine forms
compounds of various compositions stable in the dark. The structures of these compositions
are shown in Fig. 4 (II).
Unlike quercetine, its coordination compounds efficiently quench luminescence of
1
O2 [85, 86]. For instance, Mg-quercetine presumably having the structure II (Fig. 4),
quenches 1O2 in pyridine with a rate constant of (9.3 ± 0.5)×10 8 M–1 s–1 (Fig. 7). Close
values were obtained for compounds with Zn2+ and Cd2+ [86].
In the presence of foreign sensitizers, photoexcitation leads to oxidation of coordina-
tion compounds [85, 86], which is related to the properties of 1O2.
Figure 8 presents the data showing the extent of difference in the rates of photosensi-
tized oxidation of Mg-quercetine and a typical 1O2 acceptor tetracene [99–101]. This result
is explained by a higher reactivity of the former.
Comparing the oxidation of these compounds, we can obtain k r for Mg-quercetine. In
pyridine, the ratio of their observed rate constants is 12. According to the principle of com-
peting reactions [102], this value reflects the ratio of their true rate constants (k r /k r,t).
Hence, knowing k r,t for tetracene, it is easy to find kr. Taking kr,t = 5.2×107 M–1 s–1 [89],
we get that k r = 6.2×108 M–1 s–1.
Based on k t and k r, we can find k q. Its value is 3.1×0 8 M –1 s–1, which is significantly
lower than k r. These data are in favour of the chemical mechanism of 1O2 quenching by
Mg-quercetine.
As seen from the data presented, the rate constants of the elementary reactions of 1O2
ln (A0/At)
1.2
0.6 1
2
0
0 30 60
Time, s
Figure 8 Kinetics of photosensitized oxidation of Mg-quercetine (1) and tetracene (2) in pyridine
during their joint presence in solution (1O2 photosensitizer: Pd-tetra(t-butyl)phthanocyanin, λex > 640
nm, A0 and At are the initial and current optical densities of the solution at the measurement wave-
lengths, respectively).
quenching found for Mg-quercetine are by approximately two orders of magnitude higher
than for quercetine (Table 3). We have here a rather interesting example of activation of
quercetine with respect to the reaction of 1O2, similar, apparently, to the activation of un-
substituted flavonol due to the ionization of the 3-OH group [83].
To date, there is sufficient proof of accelerated destruction of coordination compounds
of quercetine in the light [44, 86]. Consider the photochemical properties of these com-
pounds in greater detail by the example of Mg-quercetine. Experiments yield the following
results [86]. The range of action of photodestruction coincides with the absorption spectrum
of this compound. For this reason, it is natural to consider the photodestruction as a self-sen-
sitized photoreaction. Its yield depends on oxygen. The involvement of 1O2 in it is indicated
by the fact that para-hydroquinone inhibits the photoreaction with the efficiency, with
which it quenches the luminescence of 1O2 (Fig. 9). Hence, an important conclusion can be
made that Mg-quercetine is a photosensitizer of 1O2. As the lifetime of its fluorescence in
pyridine does not exceed 2×10–9 s, and the fluorescence is not quenched by oxygen, it can
be concluded that 1O2 is formed as the result of quenching of excited triplet molecules of
Mg-quercetine by oxygen.
We can not but mention another significant fact, namely, that the photooxidation
mechanism of coordination compounds of quercetine depends on the nature of a metal. For
instance, inhibition of Cd-quercetine photooxidation by para-hydroquinone is much less ef-
ficient [86]. In this case, the contribution of the reaction with participation of 1O2 is reduced
to 60–70%. It is evident that in the case of coordination compounds with heavy elements
other oxidation routes are possible in principle; oxidation of excited triplet molecules by
oxygen can be assigned to these routes [25, 99].
The determination of 1O2 quantum yield is more complicated. A low spectral resolu-
tion of the setup used in [86] did not make possible the direct proof of luminescence photo-
sensitization from the 1∆g condition of O2 in solutions of these compounds. To assess the
286 E.A. Venedictov
k, rel. units
1
1
2
3
0.5
0
0 0.0006 0.0012
c, mol/l
Figure 9 Effect of p-hydroquinone on the rate of self-sensitized photooxidation of Mg2+-quercetine

(1), tetracene (3) and 1O2 luminescence (2) in pyridine [86].
quantum yield of 1O2, we used the kinetic data of the direct photooxidation of Mg-quercet-
ine and tetracene, obtained under conditions of monochromic excitation by the light with
λ = 447 nm (Fig. 10). Consider this issue in more detail.
As shown in Fig. 9, the photooxidation reactions of Mg-quercetine and tetracene are
inhibited by para-hydroquinone with the same efficiency. These data indicate that oxida-
tion is related exclusively to 1O2. Therefore, photoconversion of these compounds can be
presented as the following scheme [97–101]:
hν 1 3
S → S→ S
3 –
S + O2 (3 Σ g ) → S + 1O2
1 –
O2 → O2(3 Σ g ) + hνlum, (krad)
1 –
O2 + Solv → O2 (3 Σ g ) + Solv + Q (ks)
1 –
O2 + S → O2 (3 Σ g ) + S (kq)
1O + S → oxidation products (kr),

2
where S, 1S and 3S is the photosensitizer in the basic and excited singlet and triplet states,
respectively; Q is the energy of 1O2 dissipated as heat; Solv is solvent; krad, ks, kq, kr are
rate constants of respective processes. Hence, we can write the following expression for the
oxidation rates for each of these compounds:
0.49
0.48 sequence 1
sequence 2
0.47
0.46
A
0.45
0.44
0.43
0 100 200 300 400
Time, min
Figure 10 Optical density changes in solutions of Mg2+-quercetine (1) and tetracene (2) at a wave-
length of 447 nm during the excitation with a monochromatic light (λ = 447 nm) in air-saturated pyri-
dine (initial concentrations of Mg2+-quercetine and tetracene are, respectively, 6.5·10 –5 and
1.1·10 –4 mol/l) [86].
k r γ I [ S ]τ
–d[S] ⁄ ( dt ) = ------------------------
-.
1 + k t [ S ]τ
Here γ is the quantum yield of 1O2; I, the intensity of the absorbed light flux; [S], concen-
tration of sensitizer; τ = 1/(krad + ks [Solv]), the lifetime of 1O2 in the solvent; kt = (kq + kr).
For tetracene, the term (1 + kt [S]t) in most common solvents can be accepted to be unity
[86, 89, 98].
If solutions of these compounds are illuminated under identical conditions, then for the
extent of their conversion not exceeding 15%, the relative value of γrel can be assessed from
the ratio
α = β γrel / (1 + kt [S]τ),
where α, β and γrel are the ratios of the observed rate constant, kr and γ for Mg-quercetine
to analogous values for tetracene; [S] is the initial concentration of Mg-quercetine.
It is seen from the data presented in Fig. 10 that in pyridine α = 1.8. The value of β,
according to the above-presented data, in the same solvent is equal to 12. Then, using τ =
18×10–6 s, for the concentration of Mg-quercetine equal to 6.5×10–5 mol/l, we obtain γrel
= 0.32. Taking for tetracene γ ~ 0.71 [103], we find that γ ~ 0.22. As is seen, this value is
sufficiently high and comparable with the value of γ for unsubstituted flavone, but is lower
that the value of γ for 3,5,7,3′,4′-pentamethoxyflavone. Therefore, complexation, by elim-
inating the effect of the hydrogen atom of the 3-OH group and restricting the effect of the
5-OH group on the photophysical properties of quercetine, contributes to the photoproduc-
tion of the triplet state of quercetine capable of exciting O2.
288 E.A. Venedictov
Summing up these results, it can be inferred that an unequivocal conclusion on the pho-
tochemical inertness of quercetine with respect to oxygen can not be correct without ac-
counting for the effect of complexation with a whole range of metal ions on this property.
6 Conclusion
At present, there is no doubt that flavonols, depending on conditions, can play the role of
both photoprotectors and photosensitizers. This confirms the reality of information men-
tioned at the beginning of the chapter on the photodynamic properties of some flavonoids.
Realization of photochemical reactions is determined, apparently, by whether conditions
and structural changes favour these processes.
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8 Solvation of Drugs as a Key
for Understanding
Partitioning and Passive
Transport Exemplified by
NSAIDs
German L. Perlovich1,2 and Annette Bauer-Brandl2

1Institute
of Sciences, 153045 Ivanovo, Russia
glp@isc-ras.ru
2University of Tromsø, Institute of Pharmacy, Breivika,
N-9037 Tromsø, Norway
annetteb@farmasi.uit.no
Passive transport properties of drug molecules are of utmost importance for their
pharmacological and biopharmaceutical effectiveness. Diffusion in different
media and through lipid bilayers are in many cases the rate-determining steps for
the distribution in the body. In the present review, an attempt is made to
demonstrate the importance of solvation of drug molecules for the diffusion and
partition/distribution in phases of different lipophilicity. Different approaches
known in the literature to describe solvation of compounds with flexible
conformation are discussed as well as the experimental methods to directly
measure the energy of solvation. NSAIDs are chosen as an example of a class of
drugs of different molecular structures that have already been studied thoroughly
in many aspects. Thermodynamic characteristics of solvation of the drug
molecules yielded by independent classical experimental methods (Gibbs energy,
enthalpic and entropic terms of Gibbs energy) are used in order to better
understand the diffusion and distribution properties. Correlations between in vitro
data (the partition coefficient, enthalpy of solvation) with biopharmaceutically
relevant characteristics (plasma half-life) are also discussed.
292 German L. Perlovich and Annette Bauer-Brandl
Keywords: NSAID, sublimation, solubility, solvation, calorimetry, thermodynamic func-

tions, driving forces, crystal lattice packing architecture, hydrogen bond networks topo-
logy, plasma half-life, passive transport, distribution / partitioning.
Introduction
Combinatorial chemistry, high throughput screening and in vitro receptor affinity studies
nowadays confront the developer of new drug formulations with a huge number of potential
new drug substances. Although receptor affinity is in many cases undoubtedly a key issue
for potent drugs, other factors may be equally important for the usefulness in vivo, such as
solubility, partitioning behavior, absorption properties, active and passive transport prop-
erties and biodegradation. In many cases, unfortunately, these important aspects are studied
only late in the drug discovery / drug development process. Therefore, as most new drug
candidates are in the first place barely tested in vitro, one faces this enormous amount of
promising new drug substances of high receptor affinity with horrible physicochemical ma-
terial properties. In many cases the substances are of extremely low solubility in any of the
physiological media resulting in tremendous absorption and distribution problems. This
should be a serious drawback for a drug candidate to become a useful drug preparation, be-
cause it is hard to compensate such bad properties even by using the most advanced drug
delivery systems. It would be much more efficient and economical to pick those candidates
with less difficult properties right in the beginning of the drug development process. There-
fore, adding studies of those physicochemical and biophysical characteristics of the com-
pounds, which may be biopharmaceutically relevant to the selection procedure and the
optimization process of new drug candidates at an early phase of development, is an impor-
tant issue today. It is the purpose of the present review to draw attention to physicochemical
properties of importance in this context, and to conclusions that may be drawn from them.
1 Partitioning and diffusion

In the past, relative measures of the lipophilic–hydrophilic properties of drug molecules,
like the logarithm of the partition coefficient between water and octanol, log P, have been
used extensively [1]. The log P is a relative measure of the lipophilic–hydrophilic balance
of a compound and characterizes the equilibrium of the partitioning process in a
water–octanol system resulting from respective driving forces. It is a thermodynamic mea-
sure for processes that, however, are kinetically controlled in vivo: most of the transport and
delivery processes in a biological environment take place under essentially non-equilibrium
conditions and in non-homogeneous media. Water/octanol partitioning is frequently used
as a simple model to mimic the distribution of drug substances in biological membranes,
which is important because the pharmacokinetic properties of drug molecules are strongly
dependent on their interaction with biological membranes. The conditions of absorption and
distribution in biological systems are various and complicated, just to name the difference
between trans- and paracellular pathways from the point of view of their
lipophilic–hydrophilic composition [2–4]. Moreover, questions connected with the mech-
anism of diffusion into biological membranes (e.g., what controls the size of activation vol-
ume for passive transport? what is the nature and dimension of the units that are transferred
by the diffusion process: is it the drug molecule or the drug molecule + its solvation shell?)
have widely been outside the focus of discussion up to now [2, 4, 5, 6–8].
Solvation of Drugs as a Key for Understanding Partitioning and Passive Transport 293
boundary of phases
phase 1 phase 2
a
partitioning
Figure 1 Schematic illustration of the partition process.
1.1 The partitioning (distribution) process
The partitioning between water and octanol is the phase transition of the solute molecule
across the partition boundary of the two separated phases (water and octanol) due to the
thermodynamic driving force, which is defined by the difference of the chemical potentials
of solute molecules dissolved, respectively, in water and in octanol. However, if one sticks
to the thermodynamic formalism, a lot of questions appear about a model to describe the
elementary act of the partitioning process. First of all, it is obvious that the solute molecules
in the solution interact with their nearest neighbor solvent molecules in a stronger way com-
pared to molecules further out in the bulk. Therefore, a so-called solvation shell is created
around the solute molecule. Strength and nature of the drug–solvent interactions within this
solvation shell are non-homogeneous (non-uniform): there are weak van-der-Waals inter-
actions as well as stronger donor–acceptor interactions, hydrogen bonds (as the extreme
case of a donor–acceptor interaction) and electrostatic interactions. Since most of the drugs
are compounds with the ability to create hydrogen bonds, the molecules are solvated in so-
lutions, particularly in aqueous solutions, by strong hydrogen bonds. During the partition-
ing/distribution process, the molecule solvated in water must rebuild this solvation shell
into the octanol solvation shell (resolvation). One can imagine several variants of the resol-
vation mechanism which differ by the height of the activation energy for the partitioning
(Fig. 1):
(a) Complete destruction of the water solvation shell and creation of the new shell in the
octanol phase. This case would be preferred with weak drug–water hydrogen bonds;
the fraction of the hydrogen bond energy in this case would be much less compared
to the van-der-Waals energy, in the case of big and topologically complicated

molecules.
(b) Partly destroying the solvation shell during the phase transition step. For example,
those water molecules forming strong hydrogen bonds with the drug molecule are
kept within the solvation shell, and others are destroyed and replaced by interaction
with octanol molecules.
(c) The complete solvation shell transits without changes from the one phase into the
other.
The last two variants ((b) and (c)) will be realized if the energy fraction of the hydrogen
bonds is comparable with the van-der-Waals energy.
Using the proposed classification, the effective size of molecules taking part in the par-
titioning process varies in size of the aggregate (solvation shell). As noted above, the main
driving force of the partitioning process is the difference of the chemical potentials of the
solute molecules in the considered phases, whereas the height of the activation barrier is
determined both by thermodynamic and kinetic factors [9]. There is mutual solubility of
water and octanol (which is important in the considered water–octanol systems), which will
influence the height of the barrier of activation, and the size of the solvation shell as well.
It may therefore be assumed that the height of the activation barrier of partitioning widely
depends on the solvation characteristics of the solute (drug molecule).
1.2 Influence of the solution pH on the partition/distribution coefficients

Numerous drugs contain ionogenic functions and, dependent on the pH value of the solution
and the strength of the acid, may exist in various degrees of ionization. In the literature a
whole set of studies has been devoted to the investigation of the correlation between the pH
value and the distribution coefficients (D = P7.4) and/or the partition coefficients (P) [10,
11]. The equation relating the partition coefficient, P, (for the unionized form of the com-
pound) to the distribution coefficient, D, (for the ionized form) is well known:
log D = log P – log (1 + 10pH–pK), (1)
where pK is the acid strength.

The D values are always smaller than the P values. This means that the activation bar-
rier of partitioning is always higher for the ionic form of a drug compared with its nonionic
form. This can be explained by large energy expenses for the resolvation of molecules in
the ionic form during the transfer from the water into the octanol phase [2, 6, 7]. However,
some authors [12, 13] describe the partitioning process as the transfer of the solvated form
of the drug molecules from one phase into the other. This model concept seems reasonable,
particularly taking into consideration that the saturated octanol phase (as in the partitioning
experiments) contains more than 26 mol % of water [14]. Therefore, a close look on the sol-
vation of molecules is useful to understand the nature of partitioning and passive transport.
1.3 Diffusion of drugs

Since the lipid membrane is in the liquid crystalline state [15, 16], the mechanism of diffu-
sion of drugs into lipid membranes should be similar to diffusion processes into crystals
(amorphous state). It is well known that the diffusion of molecules/atoms is controlled by
Lateral diffusion coefficient, cm2/sec
Molecular weight, Da
Figure 2 Relationship between lateral diffusion coefficients and molecular weight of drugs. (Data
from Johnson et al. [5].)
defects of the medium (i.e. the lattice of the solid/liquid crystal). A better model concept
for the characterization of self-diffusion processes in the considered case could be the “void
volume” (which is often used for the description of diffusion processes in liquids) or va-
cancies (bi-vacancy etc., which is used for solid/liquid crystals) [17–19]. The elementary
diffusion step can be made if the following conditions are fulfilled: a) availability of a “void
volume” of the size being comparable to the size of the diffusing particle (molecule, aggre-
gate etc.); b) the activation energy of defects (“void volume”) should be enough in order to
remove it; c) the activation energy of the diffused molecule should be high enough in order
to partially destroy the interaction with the nearest neighbors (lipid matrix or octanol, for
example) and to remove the molecule from the region where the defect of the structure is
situated. Therefore, the activation energy of the diffusion process is a function of the Gibbs
energy of solvation:
∆G* = f (∆Gsolv). (2)
Consider the lateral diffusion coefficients of drug molecules into the lipid bilayers of
the stratum corneum (SC), which have been calculated from permeability values [5]. The
lateral diffusion coefficients versus the respective molecular weights are presented in Fig. 2
(data are obtained from the work by Johnson et al. [5]).
These authors paid special attention to the bifunctional size dependence of the trans-
port characteristics (diffusion coefficient) within SC. There is an overall dependence of the
lateral diffusion coefficient on the molecular weight of the compound. However, if one
picks compounds with a molecular weight of ≈ 120 Da (the dashed line in Fig. 2) as an ex-
ample, the ratio of the maximum and minimum diffusion coefficients (points 1 and 2) may
be approximately a factor of 200. This huge variation may be explained as follows. Firstly,
probably a number of different diffusion mechanisms take part in the overall diffusion (i.e.
different defects of the structure are present in the elementary diffusion step). Secondly,
aggregates (solvated drug molecules) of different size may take part in the elementary dif-
fusion act. Thirdly, the studied drugs (at pH = 7.4) are ionized to a different degree and
therefore build hydrated aggregates (solvates) of different sizes. The noted reasons are con-
sidered to be closely interrelated. Nevertheless, the conception of solvated drugs appears
useful for explanation of the transport parameters.
2 Solvation of drugs: relevance and theoretical approaches

The main qualification of drug molecules to passively penetrate through membranes by dif-
fusion is still acknowledged to be their lipophilic–hydrophilic balance. However, the dif-
fusion rate depends strongly both on the concentration gradient and the energetic
parameters of drug–membrane interaction. The diffusion process consists of a number of
elementary activation steps, each having its own defined energy barrier. These energy bar-
riers may be split up into two general forms: a) nonspecific drug–membrane interactions
(van-der-Waals interactions) and b) specific interactions (hydrogen bonding, donor–accep-
tor interactions). The overall diffusion characteristics are a function of both the absolute
strength of the mentioned interactions, and of the balance between them. In other words,
the solvation properties of drug molecules, i.e. an understanding of the nature of interactions
and the estimation of relative and absolute energetic terms thereof, are the key to understand
the mechanism of not only passive transport, but also the mechanism of drug–receptor
interactions.
2.1 The main definitions
The solvation of one mole of solute molecules in the solvent can be defined as the total
change of the standard thermodynamic functions (∆G0, ∆H0, ∆S0) of the compound when
transferring it from the gas phase (ideal gas, single molecules without interaction) into the
solvent. The thermodynamic cycle of solvation is illustrated by Fig. 3, from which it follows
that
0 0 0
∆Ysolv = ∆Ysol − ∆Ysub , (3)
where ∆Y 0 is the standard change of any of the thermodynamic functions of the solvation
(∆Y 0solv), dissolution (∆Y 0sol) and sublimation (∆Y 0sub) process. Therefore, the following
equations may be defined:
∆G0solv = ∆G0sol – ∆G0sub , (4)
∆H 0solv = ∆H 0sol – ∆H sub

0
, (5)
T∆S 0solv = T∆S 0sol – T∆S 0sub . (6)
In order to study the solvation process, which is experimentally not accessible, we need to
investigate the other two processes: sublimation and dissolution.
Gas
Solution
Solid Dissolution
state
Figure 3 Thermodynamic cycle of the solvation process.
Accessible surface
........
........... .......
..........
............
............. ...........
.............
..............
............... ..............
...............
...............
.............. ...............
............... Solvent probe
.............. ..............
.............
...........
......... ...........
.........
Excluded (Molecular) surface
Figure 4 Schematic picture illustrating an accessible surface (volume) and an excluded (molecular)
surface (volume) introduced by Lee and Richards [20, 21].
2.2 Models describing the solvation of molecules

More than 30 years ago, Lee and Richards [20, 21] introduced the definitions of accessible
volume and excluded volume (surface), which turned out to be very useful for describing
various solvation effects. Pharmaceutics and medical chemistry are not exceptions to the
broad use this approach has found, and therefore in the following paragraph we pay more
attention to definitions and applications of this approach.
The van der Waals molecular surface (SW) is the envelope surface of a set of inter-
secting spheres with given atomic radii centered on the nuclei of selected atoms of the mol-
ecule. The van der Waals volume (V W) is the volume enclosed by the van der Waals surface.
The accessible molecular surface (S acc) is the surface defined by the center of the solvent,
considered as a rigid sphere (probe sphere), when it rolls around the van der Waals surface.
The accessible molecular volume (V acc) is the volume enclosed of the accessible molecular
surface. The excluded (molecular) surface (S exc) is the surface envelope of the volume ex-
cluded to the solvent, considered as a rigid sphere, when it rolls around the van der Waals
surface. The excluded (molecular) volume (V exc) is the volume enclosed by the excluded
(molecular) surface (Fig. 4).
As a rule, the presented approach has been used for calculation of hydration energy,
where a water molecule is used as a “probe” solvent. Since the size of the water molecule
is much smaller compared to a drug molecule, the assumption that it is spherical in shape
is reasonable. In some cases the noted approach is also applied to big solvent molecules
(benzene, n-hexane), the size of which is comparable to the size of the solute molecule. In
such cases various modifications to the model are being made: an ellipsoid (or cylinder, or
any other geometric body) is used to describe the solvent molecule. It should be kept in
mind that a lot of substances are topologically complicated and their conformation is flex-
ible. The calculation of excluded (molecular) and accessible volumes (surfaces) of such
molecules is very sensitive to the conformational state they are in. Therefore, different ways
to calculate the mean statistical molecular conformational states have been suggested. For
example, let us consider some modifications of the Lee’s and Richards’s approach:
a) The hydration shell model

Kang et al. [22–25] developed a model providing the opportunity to estimate Gibbs ener-
gies of hydration of conformationally flexible solute molecules. The main basic points of
this approach are the following: the free energy of conformation i, ∆Gt(i), in solution can be
represented by the equation:
∆Gt(i) = ∆E (i) + ∆Gh(i) , (7)
where ∆E (i) is the conformational energy, and
∆Gh(i) = Σk (Vwa,k(i ) ·∆gh,k) (8)
is the free energy of hydration of the solute molecule in conformation i, expressed as the
sum of the free energies of hydration of all its constituent atoms or groups k. Vwa,k(i) is the
water accessible volume of group k in a given conformation i of the compound, and ∆gh,k
is the free energy density of hydration of group k.
In polyfunctional molecules, a correction term is added to account for the polarization
of the hydration shell around polar groups by the presence of neighboring polar groups. The
free energy of hydration of a compound is obtained as a Boltzmann weighted average over
all conformations. In order to estimate ∆gh,k, free hydration energies of various classes of
compounds have been used, which have been averaged for definite atoms (functional
groups).
b) The dynamic polar molecular surface area (PSAd ) model

Out of the numerous useful models used to describe and predict various pharmacokinetic
and pharmacodynamic properties of drugs, the model with the PSAd predictor should be
mentioned, which has been developed by Palm et al. [7, 8] during the past years. The mo-
lecular surface area is divided into a polar fraction (nitrogen atoms, oxygen atoms and the
hydrogen atoms attached to these heteroatoms) and a nonpolar fraction. The dynamic aver-
age of the surface properties is then calculated from all low-energy conformations accord-
ing to a Boltzmann distribution at 37°C. In a first step, the conformational analysis of the
drug has been carried out in vacuum, chloroform and water by Monte Carlo simulations.
The low-energy conformations were exclusively chosen for further analysis. The authors
noted that for conformational flexible molecules in vacuum and chloroform there is a wide
spectrum of low-energy conformational states, whereas in water (hydrogen bonding sol-
vent) the number of these variations is restricted. Therefore, water renders an essential in-
fluence on the stabilization of the conformation state of the drug molecule. As a next step,
-3.5
-4.0
-4.5
logPc [cm⋅c-1]
-5.0
-5.5
-6.0
-6.5
-7.0
30 40 50 60 70 80 90 100 110 120
PSAd, water [Å2]
Figure 5 Relationship between log Pc, the Caco-2 cell monolayer permeability coefficients and
PSAd,water, dynamic polar molecular surface area of β-adrenoreceptor antagonists. (Data from Palm
et al. [8].)
different low-energy conformations are compared. It appeared that the dispersion of these
values was from 6 to 54% of the dynamic average (the last value corresponds to conforma-
tionally flexible drugs). The correlation analysis shows that there are linear dependencies
between the logarithm of the Caco-2 cell monolayer permeability and PSAd values of water
(Fig. 5) for a number of β-adrenoreceptor-blocking agents that have been chosen as a sub-
ject of investigations.
The models presented above demonstrate attempts of estimating the thermodynamic
functions of solvation of solute compounds. There may be experimental data on the solva-
tion of some organic substances; however, for drug substances such data are practically ab-
sent. This is an essential obstacle for the development and improvement of models
describing the transport characteristics of drugs from the point of view of the solvation phe-
nomenon. In recent publications [26–31], in contrast to the equilibrium partitioning ap-
proach in water–octanol systems, we have tried to define an absolute lipophilicity scale for
drug molecules of the group of non-steroid anti-inflammatory drugs, NSAIDs. The work is
based on the analysis of drug solvation characteristics, where the experimental data were
obtained using classical thermoanalytical methods: sublimation, solution calorimetry and
the solubility method. This enables to us compare the solvation characteristics of a wide
range of compounds using quantitative parameters derived from experimental data. The
main results of the noted studies will be presented below.
3 Experimental methods to measure solvation characteristics and choice

of subjects
3.1 Sublimation experiment

In order to study thermodynamic characteristics of the sublimation of molecular crystals,
usually the Knudsen effusion method is used. In principle, this technique monitors the rate
of vapor loss through a small orifice under conditions of free molecular diffusion by
gravimetry.
However, during the last ten years, a transpiration (inert gas flow) method to investi-
gate the sublimation process of various molecular crystals [32, 33] and NSAIDs [27–31]
has been developed. The method has a number of advantages in comparison to effusion
method. Firstly, the transpiration method works over a wider temperature interval, which
increases the number of experimental data points and promotes a statistically better inter-
pretation of the results. Secondly, the transpiration method works at lower temperatures (in
comparison to the effusion method), which decreases the risk of chemical decomposition
of the substance. It should be noted that both sublimation methods have practically not been
used for drug substances except for the above mentioned and the following works: investi-
gation of (±)-ibuprofen sublimation by Ertel et al. [34] applying the effusion method; and
studies on the polymorphism of caffeine, theophylline and carbamazepine by Griesser et
al. [35] using the transpiration method.
In brief, the transpiration method can be described as follows. A stream of an inert gas
passes above the sample at a constant temperature and at a known slow constant flow rate
in order to achieve saturation of the carrier gas with the vapor of the substance under inves-
tigation. The vapor is condensed at some point downstream, and the mass of sublimate and
its purity determined.
The velocity of the carrier gas stream through the sublimation chamber should be cho-
sen very carefully in order to establish and maintain the thermodynamic equilibrium be-
tween the solid and the gaseous state of the substance. The apparatus needs to be tested
before starting the exact measurements by determining the relation between P and ν and
choosing the gas flow velocity value adequate to the appearance of a plateau on P = f(ν)
curve. The velocity of carrier gas flow for the considered compounds was 1.8 dm3/h.
The equipment is calibrated using the benzoic acid standard substance. The standard
value of sublimation enthalpy obtained should be in good agreement with the value recom-
mended by IUPAC of ∆H 0sub = 89.7±0.5 J·mol –1 [36]. In order to gain valid results, the
saturated vapor pressures should be measured 5 times at each temperature with the standard
deviation being within 3–5%. The experimentally determined vapor pressure data can be
described in (ln P; 1/T) coordinates by eq. (9):
ln P = A + B/T. (9)
The value of the enthalpy of sublimation is calculated by the Clausius–Clapeyron equation:
∆H Tsub = –RT2 ·∂(ln P)/∂(T), (10)
whereas the entropy of sublimation at a given temperature T was calculated from the fol-
lowing relation:
∆S Tsub = (∆H Tsub – ∆G Tsub)/T, (11)
where ∆G Tsub = –RT·ln(P/P0) and P0 = 1.013·105 Pa.

In order to improve the extrapolation to room conditions, heat capacity (Cpc298 value)
of the crystals was measured using a calibrated sapphire standard (Perkin Elmer). Heat ca-
pacity was introduced as a correction for the recalculation of the ∆H Tsub value at 298 K
(∆H 298
sub value), according to eq. (12) [64]:
298 = ∆H T + ∆H
∆Ηsub T 298
sub cor = ∆H sub + (0.75 + 0.15·Cpc )·(T – 298.15) (12)
3.2 Method of isothermal saturated solubility
In order to investigate solubility, usually the method of isothermal saturation is used. This
is a routine method and has extensively been described in the literature (for example [37]).
The Gibb’s energy of solution is derived from equilibrium conditions, i.e. solubility:
∆G 0sol = – RT·ln (X2), (13)
where X2 is the mole fraction of solute molecules.

The standard values of solution enthalpies for slowly dissolving compounds can be de-
rived from the temperature dependencies of solubility by the equation:
∆H 0sol = –RT2 ·∂(ln X2)/∂(T), (14)
However, in this case the thermodynamic functions of ∆G 0sol and ∆H 0sol have self-
consistent experimental errors and their respective dependence is difficult to interpret. In
order to make discussion of the data more correct, one needs to measure ∆H 0sol values by a
direct method (for example, calorimetry).
3.3 Isothermal calorimetry
Isothermal calorimetry provides the opportunity to measure heat effects of dissolution in

various solvents with an accuracy of approximately 0.1% (depending on the total heat ef-
fect). As already has been mentioned above, it is important that this method and the isother-
mal saturated solubility are independent experiments. Therefore, also the experimental
statistical errors obtained by the two methods are independent and not correlated. This fact
enables us to analyze enthalpy entropy-compensation effects [38] both with respect to the
solubility of definite compounds in a series of solvents, as well as in the form of a series of
solutes in a particular solvent. The noted techniques have been discussed in more detail in
[26, 27].
3.4 Choice of drugs
Non-steroidal anti-inflammatory compounds (Table 1) are destined for solvation studies

due to the following reasons. First of all, for the named substances, a lot of solubility, par-
titioning, distribution, X-ray structure, pharmacokinetic and pharmacodynamic experimen-
tal data are available in the literature. This fact essentially facilitates analysis and
interpretation of the obtained results within a logical line. Secondly, the drugs have different
basic structures: phenyl derivatives (BA, ASA, IBP); biphenyl derivatives (DIF, FBP); ben-
zophenone substituted (KETO) and naphthalene substituted (NAP). Moreover, all the con-
sidered drug substances are carboxylic acids. In conclusion, comparison of solvation
characteristics of a wide spectrum of compounds using quantitative thermodynamic param-
eters derived from experimental data is possible.
Table 1 Structural formulas of NSAIDs studied.
Compound Structural formula Compound Structural formula
Phenyl derivatives Biphenyl derivatives
Benzoic acid COOH Diflunisal (DIF) COOH

(BA)
F OH
Acetylsalicylic COOH O Flurbiprofen (FBP) CH3

acid (ASA) O C CH3
C H
COOH
F
Ibuprofen H CH 3
(IBP)
CH 3 C CH 2 C H
CH 3
COOH
Benzophenone derivative Naphthalene derivative
Ketoprofen O CH3 Naproxen CH3

(KETO) (NAP) H
C C H C
COOH COOH
H3CO
4 Crystal structures of NSAIDs
4.1 Description of hydrogen bond networks topology by graph set assignment

Hydrogen bond networks play an important role both in the creation of architecture and in
the energetic parameters of crystal lattices. Therefore, before we analyze packing architec-
tures of the drugs under investigation, let us consider an approach describing the topology
of hydrogen bond networks by means of graph set assignment introduced by Etter [58].
Graph sets describe the topological nature of the hydrogen-bond pattern and the num-
bers of donors and acceptors involved while highlighting the common features of molecular
aggregates that are not addressed by empirical formulae or by symmetry considerations of
the crystallographic space group. A graph set is specified as Gda (r), where G is the pattern
designator, a and d are the numbers of acceptors and donors, respectively, that are used in
forming the hydrogen-bond pattern, and the degree r is the total number of atoms involved
in the pattern. The pattern designator G describes the pattern of hydrogen bonding and can
be one of four types: S, C, R and D. As illustrated in Scheme 1, S (or self) denotes an in-
tramolecular hydrogen bond, C refers to infinite intermolecular hydrogen-bonded chains,
R refers to intermolecular rings, and D refers to non-cyclic intermolecular diads and other
finite hydrogen-bonded sets.
R R
R H
C C
O O O O O O
H R
H H H
O H R
O O S
R R O O
C R
S(6) C(2) R D
R2 (8)
2
Scheme 1
A hydrogen-bond pattern containing one unique type of hydrogen bond (distinguished

from other types of hydrogen bonds by chemical nature and/or the symmetry relation of the
donor and acceptor atoms used in the hydrogen bond) is referred to as a motif. Although
each example in Scheme 1 contains only one motif, a graph set may define a single motif
or it may define a pattern containing two or more motifs. A listing of all the hydrogen-bond
motifs present in a given structure is known as the first level network. Hydrogen-bond pat-
terns containing two different motifs are called second level networks.
4.2 Analysis of packing architectures of NSAIDs crystal lattices

The parameters of NSAIDs crystal lattices received from diffraction experiments are sum-
marized in Table 2.
Two molecules in both crystal lattices of (+)- and of (±)-Ibuprofen form a cyclic dimer
through hydrogen bonds of their carboxylic groups (Fig. 6a,b). However, in the unit cell of
(+)-IBP, both molecules are in the S configuration, and they are in different conformational
O2B C9B C12B

C6A O1A C8B
C12A C5A C1B C2B C7B
C11A C10A C10B C11B
C4A C2A C4B
C7A C1A C5B
C8A C13B
C3B C6B
C13A C9A C3A
O2A O1B
(a)
O1
C6 C5
(b) C12
C11 C10 C7 C4 C2 C1
O2
b C8 C9 C3
C13
c
a PowderCell 2.0
Figure 6 A perspective view of the cyclic dimer of (+)-Ibuprofen (a) and (±)-Ibuprofen (b); frag-
mentation of the molecule for calculations.
304
Table 2 Crystal lattice parameters of some NSAIDs(a).
(±)-IBP (+)-IBP ASA FBP DIF KETO NAP
Crystal data Shankland et al.(b) Perlovich et al.(c) Kim et al.(d) Perlovich et al.(e) Kim et al.(f) Briard et al.(g) Kim et al.(h)
Crystal system monoclinic monoclinic monoclinic triclinic monoclinic triclinic monoclinic
Space group P21/c P21 P21/c P1 C2/c P1 P21
a, Å 14.397 (8) 12.456 (4) 11.430 (1) 9.299 (3) 34.666 (6) 13.893 (8) 13.375 (5)
b, Å 7.818 (4) 8.0362 (11) 6.591 (1) 12.721(4) 3.743 (1) 7.741 (3) 5.793 (2)
c, Å 10.506 (6) 13.533 (3) 11.395 (2) 5.786 (5) 20.737 (4) 6.136 (2) 7.914 (3)
α, ° 90.00 90.00 90.00 82.99 (4) 90.00 89.61 (3) 90
β, ° 99.70 (3) 112.86 (2) 95.68 (1) 107.28 (4) 110.57 (2) 94.56 (4) 93.91 (3)
γ, ° 90.00 90.00 90.00 106.94 (3) 90.00 88.78 (4) 90
Volume, Å3 1165.6 (11) 1248.2 (5) 854.2 (4) 624.7 (6) 2519.4 (4) 657.639 611.7 (3)
Z 4 4 4 2 8 2 2
Dcalc, g⋅cm–3 1.175 1.098 1.400 1.293 1.324 1.28 1.25
Radiation pulsed neutron Mo Kα Cu Kα Mo Kα Mo Kα Mo Kα Mo Kα
T, °C –173 25 (2) 25 (2) 25 (2) 25 (2) 10–33 10–33
R[F2 >2σ (F 2)] 0.077 0.0385 0.046 0.0378 0.045 0.0661 0.042
Graph set assign. R 22 (8) R 22 (8) R 22 (8) R 22 (8) R 22 (8); S (5) R 22 (8) C (4)
(a) In brackets, standard deviations; (b)(CSD-IBPRAC02) Ref. [59]; (c)Ref. [31]; (d) Ref. [60]; (e) Ref. [50]; (f)Ref. [61]; (g)(CSD-KEMRUP) Ref. [62];
(h)
(CSD-COYRUD11) Ref. [63].
states, in contrast to (±)-IBP, where the dimer is formed by hydrogen bonds across the cen-
ter of inversion (space group P21/c), one molecule being R and the other S. Therefore, the
topology of the hydrogen bonds networks of both ibuprofens can be described by R22 (8)
graph set assignment.
Some special characteristics of the conformational states and geometry of hydrogen
bonds of both (+)- and (±)-IBP are presented in Table 3. As the two molecules of (+)-IBP
are situated in the asymmetric unit, the geometries of the two hydrogen bonds are not equiv-
alent: one of them is shorter than the other. Comparison of the data leads to the following
conclusions: a) the angle of the hydrogen bonding of the racemate is more planar in com-
parison to the enantiomer; b) the D…A distance of one of the hydrogen bonds of the (+)-IBP
is approximately equal (within experimental errors) to the analogous value of the enanti-
omer, whereas the second hydrogen bond is longer; c) the H…A distance of the (+)-IBP is
the average of the analogous values of the enantiomer. The conformational states of the
enantiomer molecules in the asymmetric unit cell are different. As it follows from Table 3,
one of the two molecules in the (+)-IBP asymmetric cell has approximately the same con-
formational state as in the racemic IBP crystal.
Table 3 Some parameters, which characterize the conformational states and hydrogen bond
geometry of (+)- and of (±)-ibuprofen molecules in the crystal lattice.
(+)-IBP (±)-IBP
A B
∠C5-C4-C2-C3 [°] 144.4 (4) –29.1 (4) 140.9 (4)

∠C7-C10-C11-C12 [°] –67.9 (5) 68.0 (5) –67.3 (4)
∠C4-C2-C1-O1 [°] 81.7 (4) –83.5 (3) 88.7 (3)
O2-C1 [Å] 1.219 (3) 1.226 (3) 1.222 (3)
O1-C1 [Å] 1.302 (4) 1.302 (4) 1.305 (3)
C1-C2 [Å] 1.496 (5) 1.518 (4) 1.509 (3)
Hydrogen bond geometry
D⎯H…A D⎯H [Å] H…A [Å] D…A [Å] D⎯H…A [°]
(+)-IBP (A) O1A-H1AO…O2B 0.94 (5) 1.73 (6) 2.651 (4) 169 (5)
(+)-IBP (B) O1B-H1BO…O2A 1.07 (5) 1.58 (5) 2.634 (4) 168 (4)
(±)-IBP O1-H1O…O2(a) 0.963 (13) 1.664 (10) 2.627 (7) 179.5 (7)
(a) Symmetry
code: 1-x,1-y,1-z
The packing architectures of flurbiprofen, ketoprofen and acetylsalicylic acid crystal

structures are shown in Fig. 7a,b and c, respectively. Both FBP and KETO structures have
the same symmetry of the crystal lattices – triclinic, whereas ASA has the monoclinic sym-
metry. However, the molecules of outlined compounds form dimers packing architecture.
Therefore, the hydrogen bond networks can be described by R22 (8) graph set assignment.
It should be noted that for FBP and KETO the dimers in the crystal lattices have conforma-
tional states like a “chair”.
Projection of the naproxen crystal lattice along the OX axis is presented in Fig. 8a. In
contrast to the previous structures, the hydrogen bonds create infinite chains (helicoids)
(a) (b)
(c)
Figure 7 The packing architecture of flurbiprofen (a), ketoprofen (b) and acetylsalicylic acid (c)
crystal structures.
(a) (b)
Figure 8 The packing architecture of naproxen (a) and diflunisal (b).
with graph set assignment C (4) along the two-fold screw axis. The packing architecture of
diflunisal is shown in Fig. 8b and may be characterized by motifs R22 (8); S (6).
with graph set assignment C (4) along the two-fold screw axis. The packing architecture of
diflunisal is shown in Fig. 8b and may be characterized by motifs R22 (8); S (6).
Based on the carried out analysis, it can be concluded that the topology of hydrogen
bond networks in the crystal lattices of the outlined class of compounds differs. For exam-
ple, ASA, (+)- and (±)-IBP, KETO and FBP molecules are packed with only dimer aggre-
gates with the graph set assignment R22 (8). The DIF molecules in the crystal lattice create
additional multicenter hydrogen bonds (combination of inter- and intramolecular hydrogen
bonds). Due to this fact, the topology of hydrogen bond networks has a higher level, includ-
ing several motifs: R22 (8); S (6). NAP molecules form only a C (4) motif. In order to find
out correlations between the crystal lattice parameters and the thermochemical and thermo-
dynamic characteristics, let us consider some experimental methods and approaches which
give an opportunity to analyze the outlined functions correctly.
5 Thermochemical and thermodynamic properties of NSAIDs
5.1 Thermodynamic characteristics of sublimation of NSAIDs

The saturated vapor pressure data and thermodynamic functions of sublimation, fusion and
vaporization processes calculated from the experimental data are summarized in Tables 4
and 5, respectively.
Table 4 Coefficients and correlation parameters of the regression equation ln (P [Pa]) = A + B/T
for the studied NSAIDs.
ASA (+)-IBP (±)-IBP FBP DIF NAP KETO
[t1–t2](a) 40.0–89.0 32.0–45.0 40.0–67.0 68.5–104.5 76.0–137.0 68.0 –124.0 68.0–91.5

A 38.2±0.2 38.1±0.2 40.4±0.2 33.8±0.2 36.4±0.2 39.7±0.2 33.0±0.2
–B 13190±65 12920±60 13927±73 13040±60 14400±800 15431±65 13250±60
r (b) 0.999 0.999 0.999 0.9998 0.9997 0.99987 0.999
σ (c) 3.74·10–2 9.1·10–3 2.54·10–2 1.62·10–2 3.72·10–2 3.22·10–2 1.35·10–2
F (d) 41808 48662 36648 52994 36261 58613 47818
n(e) 23 14 25 10 16 15 16
(a)experimental temperature interval, °C; (b)pair correlation coefficient; (c)standard deviation; (d)cal-
culated Fisher distribution value; (e)number of experimental points.
The Gibbs energy of the sublimation process at room temperature can be separated into
relative fractions of the enthalpic and entropic terms by the following parameters:
298 298 298

ς H = (∆H sub /(∆H sub + T ∆Ssub )) ⋅ 100%, (15)
298 298 298

ς TS = (T ∆Ssub /(∆H sub + T ∆Ssub )) ⋅ 100%. (16)
Results of these calculations are also shown in Table 5. It is not difficult to see that in
all studied cases the sublimation process is an enthalpy controlled process because the en-
thalpy exceeds the entropy by a factor of approximately 1.8. However, the relative fraction
308
Table 5 Thermodynamic characteristics of processes of sublimation, fusion and vaporization of NSAIDs.
BA(c) ASA(d) (+)-IBP(g) (±)-IBP(g) FBP(i) DIF(i) NAP(l) KETO(n)

298
∆ G sub [kJ·mol–1] 34.4 43.6 41.6 44.2 53.3 57.6 58.5 57.0
298
∆ H sub [kJ·mol–1] 90.5± 0.3 110.2±0.5 107.7±0.5 116.0±0.6 110.2±0.5 120.1±0.6 130.1±0.5 111.9±0.5
298
T⋅∆ S sub [kJ·mol–1] 56.1 66.6 66.1 71.8 56.9 62.5 71.6 54.9
298
∆ S sub [J·mol–1 ·K–1] 188 ±2 223±2 222±2 241±2 191±1 210±2 240±2 184±1
ςH [%](a) 61.7 62.3 62.0 61.8 65.9 65.8 64.5 67.1
ςTS [%](a) 38.3 37.7 38.0 38.2 34.1 34.2 35.5 32.9
T (f) [°C] 122.3±0.5 141.0±0.5(e) 50.3±0.4(h) 74.0±0.4(f) 113.5±0.2(k) 212.8±0.2(j) 154.4(m) 93.9±1.3(o)
298 (e) (h) (f) (k) (j) (m)
∆ H fus [kJ·mol–1] 18.1±0.2 30.2±0.2 15.4±0.4 23.1±0.4 26.4±0.5 35.9±0.2 31.5±2.1 21.0±0.8(o)
298
∆ H fus [kJ·mol–1](q) 13.6 21.7 14.2 19.8 20.4 22.0 22.0 17.1
298
∆ S fus [J·mol–1 ·K–1](b) 45.8 72.9 48 67 68.3 73.9 74 57
298
∆ H vap [kJ·mol–1] 76.9 88.5 93.5 96.2 89.9 98.1 108.1 94.8
(a)ς 298 298 298 298 298 298

H = (∆ H sub /(∆ H sub + T∆ S sub ))·100%; ςTS = (T∆ S sub /(∆ H sub +T∆ S sub ))·100%;
(b)∆S
fus = ∆Hfus/T(f);
(c)Ref.
[32]; (d)Ref. [26]; (e)Ref. [47]; (f)Ref. [48]; (g)Ref. [31]; (h)Ref. [42]; (i)Ref. [27];
(j)Ref.
[49]; (k)Ref. [50];
(l)Ref.
[29]; (m)Ref. [51]; (n)Ref. [28]; (o)Ref.[52]
(p)∆ H 298 298 298
vap =∆ H sub −∆ H fus
(q)∆ H 298 T T
fus = ∆ H fus – ∆ S fus (Tm – 298.15)
of the enthalpic term varies between 61.7% for benzoic acid and 67.1% for ketoprofen. For
a better comparison, benzene [39], biphenyl [39], benzophenone [40] and naphthalene [41]
as the non-substituted analogues of the studied NSAIDs (which in their crystal lattices ex-
clusively interact non-specifically by van-der-Waals forces) were investigated. The value
of solid state benzene ςH (Ben) = 53% differs significantly from the benzene derivatives of
the NSAIDs (ςH (BA) = ςH (ASA) = ςH ((+)-IBP) = ςH ((±)-IBP) ≈ 62%). The analogous val-
ue of biphenyl ςH (BiPh) = 60% differs also from the biphenyl substituted compounds:
ςH (FBP) = ςH (DIF) ≈ 66%. The same situation is observed for naphthalene ςH (Naph) =
59% and naproxen ςH (NAP) ≈ 65% and also for benzophenone ςH (BenzPhen) = 62% and
ketoprofen ςH (KETO) ≈ 67%. The causes of the non-systematic share between enthalpy
and entropy with respect to the skeletal structures are supposed to be a different distribution
of the total crystal lattice energy between van-der-Waals interactions and hydrogen bonds
depending on the functional groups in the molecules.
5.1.1 Differences of racemate and enantiomer ibuprofen crystal lattices

It is interesting to note that the difference between the Gibbs energies of (±) and (+)-IBP at
25°C is 2.6 kJ·mol –1 (Table 5). This value practically coincides with the value of heat fluc-
tuation, RT = 2.5 kJ·mol –1. This fact once more confirms that the problem of separation of
the enantiomers is very delicate. Using the thermodynamic cycle (whilst neglecting the dif-
ferences of the heat capacities between the racemate and chiral substances which is sup-
posed to be very small), the thermodynamic functions of evaporation of both (±)- and
(+)-IBP may be estimated as follows:
298 298 298

∆H vap = ∆H sub − ∆H fus , (17)
298 298 298

∆S vap = ∆Ssub − ∆Sfus . (18)
298 for (±)-IBP is slight higher

It is not difficult to see (Table 5) that the value of ∆H vap
than that for (+)-IBP. The standard value of the entropy of sublimation of (±)-IBP at 298 K
exceeds that of (+)-IBP by 19 J·mol –1 ·K –1. The difference between the entropies of fusion
at 298 K between the racemate and the enantiomer is also 19 J·mol –1 ·K –1 (Table 5). There-
fore, the entropy of evaporation for both compounds considered coincides at 174
J·mol –1 ·K –1. If we take into account the thermodynamic cycle presented in Scheme 2 then
the difference between the entropies of (±) and (+)-IBP crystal lattices may be calculated as:
∆∆S = ∆Ssub (± ) − ∆Ssub (+ ) − R ln 2 = 13 J ⋅ mol-1 ⋅ K -1 . (19)
This value quantifies the difference between the entropies of the crystal lattices of the
racemate and the enantiomer and is only caused by particularities of the respective crystal
lattice structures.
For a deeper understanding of the nature of interaction of IBP molecules both in the
racemate and the enantiomer in the crystal lattice, the packing energies are calculated. For
this purpose X-ray data for (+)-IBP obtained by us [31] and the neutron diffraction data for
(±)-IBP from Shankland et al. [59] (refcode CSD – IBPRAC02) were used. Molecular crys-
tals consist of discrete molecules, which interact with each other by intermolecular
oooooooo
chiral gas racemic gas

Rln2
∆Ssub((+)-IBP)
∆Ssub(IBP)
∆∆S
1/2 (+)-IBP
1/2 (-)-IBP
chiral crystal racemic IBP
crystal
Scheme 2
non-bonded interactions. Therefore the crystal lattice energy, Elatt, may conditionally be di-
vided into three main terms: van der Waals, E vdw; electrostatic (Coulombic), E coul; and hy-
drogen bonds energy, E HB:
E latt = E vdw + E coul + E HB . (20)
The results of calculations of the energetic terms of the (±)- and the (+)-IBP crystal
lattices for both the Mayo et al. [65] (M) and Gavezzotti [66] (G) force field are presented
in Table 6. As it follows from Table 6, the van der Waals terms of both (+)- and (±)-IBP
are approximately equal if calculated by the same force field. It should be noted that this
value is slightly higher (absolute value) for the G force field compared to the analogous val-
ue for the M force field. In contrast, the terms related to energies of hydrogen bonds show
an opposite trend: for the G force fields the term on approximately 3 kJ·mol –1 less than for
the M force field. The opposite trends sum up to approximately the same total values of the
Table 6 The calculation results of the various energetic terms of (+)- and (±)-ibuprofen crystal
lattices obtained by the two types of the force fields (Mayo et al. [65] and Gavezzotti et al. [66]).
Terms(a) (±)-IBP (+)-IBP ∆((±)-(+))
Gavezzotti et al.
Evdw –78.0 (71.0) –78.8 (72.8) 0.8
Ecoul –2.8 (2.6) –2.5 (2.3) –0.3
EHB –29.0 (26.4) –26.9 (24.9) –2.1
Elatt –109.8 –108.2 –1.6
Mayo et al.
Evdw –71.7 (65.2) –73.1 (68.2) 1.4
Ecoul –2.8 (2.5) –2.5 (2.1) –0.3
EHB –35.5 (32.3) –31.9 (29.7) –3.6
Elatt –110.0 –107.5 –2.5
298
H sub 116.0 ±0.6 107.7±0.5 8.3
(a)[kJ⋅mol–1];
in brackets E term/Elatt in % is presented.
- (+)-IBP
- (±)-IBP
40
30
[%]
vdw
EX-X / E
20
10
0
H-H C-H H-O C-C C-O O-O
Types of interactions
Figure 9 The energetic terms of different types of nonbonded van der Waals interactions of the (+)-
and (±)-ibuprofen crystal lattices (Mayo et al. force field).
crystal lattice energies obtained by the two force fields considered. It should be mentioned
that for (±)-IBP the ratio between hydrogen bonding energy and common crystal lattice en-
ergy is sensitive to the choice of the force field: 26.4% for the G force field and 32.3% for
the M. The same tendency is observed for (+)-IBP: 24.9% for G and 29.7% for M. It should
also be noted that the van der Waals term of (+)-IBP is higher (absolute value) compared
to the (±)-IBP irrespective of the force field used. Therefore, it may be assumed that for
(±)-IBP the loss of van der Waals energy (in comparison with (+)-IBP) is compensated by
hydrogen bonding energy. Probably, these two energies are competing when enantiomer
and/or racemate crystals are growing.
Comparative analysis of the energetic terms of different types of nonbonded van der
Waals interactions for the crystal lattices considered (Mayo et al. force field) was carried
out. The results thereof are presented in Fig. 9. As it follows from Fig. 9, both for (+)- and
(±)-IBP, the dominating contributions are interactions between: C–C > C–H > C–O. More-
over, a transition from (+)- to (±)-IBP makes the relative contributions of the following
terms decrease: C–H, C–C and H–O, while a slight increase of the H–H, C–O and O–O
interaction is noted.
Because the contribution of the C–H interactions for both compounds contribute by
more than 25% to the common energy of the nonbonded van der Waals interactions, it
should be expected that if the positions of the hydrogen atoms in the unit cells could be re-
solved more accurately by neutron diffraction experiments, the accuracy of the final result
should be essentially increased. Lacking these data, we tried to estimate the influence of the
C–H distance on the van der Waals term of the crystal lattice energy. For this purpose, in
the calculation procedure, only the C–H distance is changed (from 0.95 to 1.20 Å), and the
same coordinates of the other atoms in the unit cell retained. The results of the calculation
are presented in Fig. 10. In Fig. 10 the filled symbols mark the E vdw values corresponding
to the C–H bonds obtained from diffraction experiments. From Fig. 10, it follows that with-
in the noted variation interval of the C–H bond lengths, the E vdw values for (+)-IBP are
changed within 4.5 kJ·mol –1, whereas for (±)-IBP – within 17.6 kJ·mol –1. This fact con-
firms once more that the van der Waals energy of the enantiomer is approximately two times
less sensitive to C–H bond variations (6.2%) in comparison to the racemate (24.4%). This
-54 (±)-IBP
-56
-58
-60
Evdw [kJ⋅mol-1] -62
-64
-66
(+)-IBP
-68
-70
-72
-74
0.95 1.00 1.05 1.10 1.15 1.20
RC-H [Å]
Figure 10 The dependence of the van der Waals term of the crystal lattice energy on the length of
the C–H bond (the filled symbols mark the Evdw values corresponding to the C–H bonds obtained
from the diffraction experiments).
fact also stresses that the estimation of crystal lattice energy for (+)-IBP does not too much
depend on the accuracy of hydrogen atoms coordinates (by X-ray or neutron diffraction).
5.2 Thermochemical characteristics of NSAIDs

The thermodynamic functions of fusion and vaporization processes for the NSAIDs studied
are presented in Table 5.
In the next step of the investigations we tried to analyze the influence of the topology
of hydrogen bond networks on the thermodynamic functions of the fusion process. The de-
pendence of the fusion entropies on the fusion enthalpies of the compounds discussed is
shown in Fig. 11.
It should be noted that the drugs with the complex structure of hydrogen bond network
deviate from the common trend line, with a tendency of decreased fusion entropy. Probably,
the hydrogen bond networks are kept maximally in the liquid state after the fusion process.
Substances situated on the trend line, probably, realize the degrees of freedom completely
(if one takes into account the hydrogen bond networks in the melt). The compounds, which
deviate from the trend line to the side of fusion entropy increase, are conformationally more
flexible in comparison with the others. Probably, in the fusion process, as the hydrogen
bonds are broken off, additional degrees of freedom appear due to conformational (struc-
tural) disordering.
In order to find out a regularity between the characteristics of the fusion process and
crystal lattice parameters, we calculated the free molecular volumes, V free, of the consid-
ered drugs in the crystal lattices. For this purpose, the following algorithm was used. The
unit cell volumes, Vcell, were estimated based on X-ray diffraction experiments. After them,
the van der Waals molecular volumes, V vdw, into the crystal lattice by GEPOL package [67]
and Kitaigorodsky radii [68] were calculated and the free volumes were derived by the
equation:
V free = (Vcell – Z·V vdw )/Z, (21)

Figure 11 Dependence of fusion entropy on fusion enthalpy of drugs (the graph set assignment is
shown under the drug name). DIF: Diflunisal; NAP: Naproxen; IBP: Ibuprofen; ASA: Acetylsalicylic
acid; FBP: Flurbiporfen; Keto: Ketoprofen; SA: Salicylamide; i-OH-BA: i-hydroxybenzoic acids;
Me-, Et-, Pr-, BuPB: Parabens.
Figure 12 Dependences of melting points on molecular free volumes in crystal lattices (symbols
correspond to Fig 11).
where Z is the number of molecules (structural units) in unit cell.

Dependences of the melting points on molecular free volumes in the crystal lattices of
drugs under investigation are presented in Fig. 12. It is not difficult to see a regularity: an
increase of V free values leads to a decrease of the melting temperature, with an essential
exception for naproxen due to structural disordering of the molecules in respective crystal
lattices.
6 The difference between partitioning and distribution of NSAIDs from

the thermodynamic point of view
The thermodynamic cycle of the relationships between the thermodynamic parameters of a
drug molecule HD and its dissociate D – + H + is shown in Scheme 3. The thermodynamic
parameters of solution and solvation are presented in Tables 7 and 8.
HD(gas)
∆Hsolv
∆Hsub ∆Hsolv
HD(buffer pH 2.0)
∆Hsol
∆Hdep
∆Hsol
HD(solid) (D- + H+)
(buffer pH 7.4
Scheme 3
Table 7 Coefficients and correlation parameters of the regression equation ln (X2) = A + B/T for
the studied NSAIDs in buffers with pH 2.0 and pH 7.4.
(+)-IBP (±)-IBP FBP DIF NAP KETO
[t1°C – t2°C](a) 20.0 − 42.0 20.0 − 42.0 20.0 − 42.0 20.0 − 42.0 20.0 − 42.0 20.0 − 42.0
pH 2.0
–A 2.3±0.2 1.0±0.3 –2.9±0.2 6.0±0.4 2.33±0.02 –1.8±0.2
–B 2874±76 3518±101 5195±65 2582 ±118 3367±6 4123±48
r(b) 0.9990 0.9988 0.9998 0.9969 0.9999 0.9998
σ(c) 1.45·10–2 1.24·10–2 1.24·10–2 2.27·10–2 1.09·10–3 9.26·10–3
N(d) 5 5 5 5 5 5
pH 7.4
–A 6.0±0.2 4.1±0.2 8.8±0.2 8.6±0.2 9.08±0.04 4.4±0.1
–B 1670±55 2345±48 811±54 952±62 628±13 1929±32
r(b) 0.9984 0.9994 0.9934 0.9938 0.9994 0.9996
σ(c) 1.06·10–2 9.22·10–3 1.04·10–2 1.19·10–2 2.41·10–2 6.22·10–3
n(e) 5 5 5 5 5 5
(a) experimental temperature interval; (b) pair correlation coefficient; (c) standard deviation;
(d) number of the experimental points
Table 8 Thermodynamic characteristics of solubility and solvation processes of some NSAIDs in aqueous buffers at pH 2.0 and 7.4 at 25 °C.
(+)-IBP (±)-IBP DIF FBP KETO NAP
pH 2.0
0
∆G solv [kJ · mol–1](a) 29.7 31.7 (34.85)(e) 36.3 36.0 (35.7)(e) 29.8 (28.5)(e) 33.8 (33.72)(e)
0
∆H solv [kJ· mol–1] 23.9±0.6 29.3±0.8 (9.6)(e) 21.5±1.0 43.2±0.5 (12.5)(e) 34.3±0.4 (26.4)(e) 28.0±0.1 (21.3)(e)
0
T∆S solv [kJ·mol–1] –5.8 –2.4 –14.8 7.2 4.5 –5.8
0
∆S solv [J·K–1 ·mol–1] –19.5±2.1 –8.0±2.7 –49.6±3.4 24.1±1.7 15.1±1.3 –19.5±0.4
0
– ∆G solv [kJ · mol–1](b) 11.9 12.5 21.3 17.3 27.2 24.7
0
– ∆H solv [kJ · mol–1] 83.5±1.1 86.5±1.2 97.8±1.6 65.2±1.0 75.8±0.9 100.3±0.6
0
– T∆S solv [kJ · mol–1] 71.6 74.0 76.5 47.9 48.6 75.6
0
– ∆S solv [J·K–1 ·mol–1] 240.1±3.7 248.2±4.0 256.6±5.4 160.7±3.4 163.0±3.0 253.6±2.0
ςHsolv [%](c) 53.8 53.9 56.1 57.6 60.9 57.0
ςTSsolv [%](d) 46.2 46.1 43.9 42.4 39.1 43.0
pH 7.4
0
∆G solv [kJ· mol–1](a) 28.8 29.8 29.2 28.7 27.0 27.7
0
∆H solv [kJ·mol–1] 13.9 ± 0.5 19.5±0.4 7.9±0.5 6.7±0.5 16.0± 0.3 5.2± 0.1
0
T∆S solv [kJ·mol–1] –14.9 –10.3 –21.3 –22.0 –11.0 –22.5
0
∆S solv [J· K–1 ·mol–1] –50.0±1.0 –34.5±1.5 –71.4±1.7 –73.8±1.7 –36.9±1.0 –75.5±0.4
0
– ∆G solv [kJ·mol–1](b) 12.8 14.4 28.4 24.6 30.0 30.8
0
– ∆H solv [kJ·mol–1] 93.5±1.0 96.3±1.0 111.4±1.1 101.7±1.0 94.1±0.8 123.1±0.6
0
– T∆S solv [kJ·mol–1] 80.7 81.9 83.0 77.1 64.1 92.3
0
Solvation of Drugs as a Key for Understanding Partitioning and Passive Transport
– ∆S solv [J⋅K–1 ·mol–1] 270.7 ±3.3 274.7 ±3.3 278.4±3.7 258.6±3.4 215.0±2.7 309.6 ±2.0
ςHsolv [%](c) 53.7 54.0 57.3 56.9 59.5 57.1
ςTSsolv [%](d) 46.3 46.0 42.7 43.1 40.5 42.9
0 0 0 0 0 0
315
(a)accuracy,
2 %; (b)Ref. [77]; (c)ςH = (⏐ ∆H solv ⏐/(⏐ ∆H solv ⏐+⏐ T∆S solv ⏐)) ·100%; (d)ςTS = (⏐ T∆S solv ⏐/(⏐ ∆H solv ⏐+⏐ T∆S solv ⏐)) ·100%; (e)Ref. [53].
6.1 Solvation characteristics of dissociated and non-dissociated (+)- and (±)-IBP
The respective solution enthalpies, ∆H 0sol , were calculated using the van’t Hoff relation-
ship, which – in contrast to other works [69] – was found to be satisfactorily linear. Further
it was found that the dissolution of ibuprofen in the buffers, both the racemate and the pure
enantiomer, is endothermic (Table 8). Moreover, the values of the entropy of the dissolution
process, ∆S 0sol , (calculated from solubility and enthalpy) are negative. Probably, while a
molecule is transferred from the solid state into the solution, some structure is built in the
solvation shell and in the surrounding solvent, overcompensating the increase in entropy
caused by the dissolution due to a “hydrophobic effect” [70–73] of solvation.
Analysing the solvation process in more detail (Table 8), in both buffers the solvation
is found to be exergonic, and Gibbs energy of solvation, ∆G 0solv , to comprise negative val-
ues for both its enthalpic and entropic terms, ∆H 0solv , and ∆S 0solv . In the current case, the
main driving force of solvation is enthalpy, which is regarded as a “classical” hydrophobic
interaction as the mechanism of solvation [70–73]. The significance of entropy for the sol-
vation process, which is decreasing and working in the opposite direction by probably cre-
ating solvent cages around the solute molecules, is not much smaller than enthalpy at room
temperature.
When further comparing the racemate with the pure enantiomer, all the (absolute) val-
ues of the thermodynamic solvation functions for (+)-IBP are slightly smaller (taking into
account the experimental errors) compared to (±)-IBP, for both the dissociated and non-dis-
sociated form. This means that the solvation of racemic IBP molecules is slightly stronger
compared to the pure enantiomer. This behaviour is probably connected with a difference
in the molecular association states for the racemate and the pure enantiomer in solutions.
The molecules may be exposed to interaction of neighboring molecules /solvation shells
present in the buffers, like dimers (or multimers). These would probably be of different
symmetry in the case of the racemate compared to the pure enantiomer (similarly to the
symmetry of dimers in crystals). Different symmetry determines small variations of the
structure of the solvation shells and their thermodynamic characteristics: the more symmet-
rical (±)-IBP dimer has a stronger ability of solvation. However, the solubility is consider-
ably higher (by approx. a factor of 2) for the pure enantiomer than for the racemate, which
means that the distance to neighboring molecules is smaller. It is difficult to decide whether
this effect accounts for the difference in solvation energy, considering the generally very
low solubility.
Comparison of the enthalpy values in the respective solutions of different pH, as pre-
sented in Scheme 3 and Table 8, enables one to calculate the enthalpy of deprotonation of
the molecules. Deprotonation is exothermic, the absolute value of enthalpy of protonation
/deprotonation is 10 kJ·mol –1, coinciding within experimental error both for (+)- and
(±)-IBP.
It should be noted that the ionic state of molecules in general is more important for the
solvation thermodynamics compared to non-ionic molecules (Table 8). This difference is
higher than the differences between the racemate and the pure enantiomer at the same pH.
The total solvation abilities of (+)- and (±)-IBP for the different states of protonation over-
0 0 0 0
lap: | ∆Ysolv ((+)-IBP)| < | ∆Ysolv ((±)-IBP)| < | ∆Ysolv ((+)-IBP-)| < | ∆Ysolv ((±)- IBP-)|. Partic-
ularly for the entropy, the protonated / deprotonated state of the molecule is of significance.
Therefore, it may be speculated that also the interaction of the solvated IBP-molecules with
membranes and receptors is widely dependent on the properties of the surrounding medium
in terms of pH, possibly also on ion strength and composition.
- pH 2.0
- pH 7.4
60
50
40
ζΗ [ %]
30
20
10
0
(+)-IBP IBP DIF FBP KETO NAP
Figure 13 Comparative analysis of the relative enthalpic parameter, ςH, of the studied drugs for
buffers with pH 2.0 and 7.4.
6.2 Solvation characteristics of dissociated and non-dissociated forms of the other

NSAIDs
As it follows from Table 7 and as is shown in Table 8, dissolution in aqueous buffers both
at pH 2.0 and pH 7.4 is endothermic for all drugs studied. This is evidence for solvation
enthalpies not overweighing the respective crystal lattice energies (as was discussed above
for IBP). The entropies of dissolution are, as a rule (with the exception of FBP and KETO
at pH 2), negative. Therefore, the degree of order in the solvation shells and in the solvent
structure increases (i.e. there is a hydrophobic effect). In all cases, the enthalpic term of the
Gibbs energy of solvation exceeds the entropic term. The relative enthalpic parameter
(ςHsolv) is between 53.8% (for (+)-IBP at pH 2.0) and 60.9% (for KETO at the same pH).
The comparative analysis of ςHsolv values of the studied drugs for the two buffers consid-
ered is shown in Fig. 13. It is not difficult to see that the noted parameter is sensitive to the
variation of buffers only for KETO, DIF and FBP. It should be mentioned that for every
drug under investigation and for the two buffers considered the solvation process is an en-
thalpy-driven process.
It is interesting to note that the enthalpy of transition from pH 7.4 to 2.0, ∆Htr (pH 7.4
→ pH 2.0), which characterises the protonation process, is endothermic in all cases. The
values vary from a minimum for (±)-IBP (9.8 kJ·mol –1) to a maximum for FBP (36.5
kJ·mol –1) by a factor of more than three. All these values exceed by far the enthalpy of
proton ionisation in dilute aqueous solutions, which have been reported for some aromatic
acids [74]. Probably, in the present case, the solvation effects play an essential role in the
transfer of molecules from one buffer to the other. It can be assumed that fluorine atoms (as
an electron acceptor) in molecules of diflunisal and flurbiprofen induce an essential redis-
tribution of the electron density from the COO – group to the phenyl ring (by conjugation
effects). As a consequence thereof, solvation effects are increased by both specific and
non-specific interaction (electrostatic interactions and hydrogen bond energy terms). Prob-
ably, the outlined effect of F atoms is the cause of an extraordinarily large increase in sol-
5.5
(+)-IBP IBP
5.0
KETO FBP
4.5
NAP
pKa
4.0
3.5
DIF
3.0
0 1 2 3 4 5 6 7 8
∆Gtr(pH 7.4 → pH 2.0) [kJ⋅mol ]

-1
Figure 14 Dependence of pKa value on the transfer of Gibbs energy of solvation from the buffer at
pH 7.4 to the buffer at pH 2.0; ∆Gtr(pH 7.4 → pH 2.0) = ∆G0solv
pH2.0 – ∆G0 pH7.4.
sol
ubility with pH that is found for DIF and FBP (Table 7) compared to the other compounds
studied.
It is not difficult to see a regularity between transfer energy ∆Gtr (pH 7.4 → pH 2.0)
values and pKa (Fig. 14): the weaker the acid, the lower the value of the driving force for
the transfer process (and the easier it is to protonate the respective base). It should also be
noted that a compensation effect is observed between the thermodynamic functions of
transfer, which can be described by the following equation:
T∆Str = (–8.2 ± 0.2) + (0.8 ± 0.1) ·∆Htr , (22)
σ = 2.29; r = 0.970; F = 64.4; F 2.5%

tab = 9.365; n = 6.
In other words, the entropic term of the Gibbs energy is 0.8 times less compared to the
enthalpic term.
It should also be kept in mind that determination of pKa values of poorly soluble drugs
is a delicate experiment, and the values may differ considerably according to the method
used. In the case of IBP, for example, using apparent pKa values in different solvent/water
mixtures and extrapolating to 0% solvent content, the pKa value varies between 5.2 and 4.3
[75]. The pKa values reported for other NSAIDs vary within the similar ranges [76]. In the
present study, this was not taken into consideration, neither for correlation analysis nor in
Fig. 14, because it would not affect the key messages anyway.
6.3 Solvation characteristics of transfer process of dissociated and non-dissociated

molecules from buffer to n-octanol
Taking into account the thermodynamic data of solvation in octanol measured earlier for
the NSAIDs discussed [77], it is possible to analyse the transfer of dissociated and non-dis-
sociated molecules from a respective buffer solution into the octanol phase. The thermody-
namic relations between the parameters are illustrated in Scheme 4. Knowledge about
hydration and solvation characteristics of drug molecules exclusively enables the use of an
absolute energetic scale. The discussed thermodynamic parameters together with related lit-
erature data are presented in Table 9.
Buffer pH 2.0
Buffer pH 7.4
T⋅∆Str
∆ Gt
T⋅∆Str ∆ Gt
∆Ht
n- Octanol
∆Ht
Scheme 4
From Table 9 and Scheme 4 it follows that the relationship between the outlined func-
tions can be described as follows:
Buffer pH 2.0 → octanol: ∆Htr < 0; ∆Str > 0; |∆Htr |<|T∆Str |
Buffer pH 7.4 → octanol: ∆Htr > 0; ∆Str > 0; |∆Htr |<<|T∆Str |.
Thus, the two types of the transfer processes (non-dissociated molecule to octanol
phase, and dissociated molecule to octanol) are basically different regarding their respective
driving force. Here, partitioning is a typically enthalpy-driven process, whereas the second
case, i.e. distribution of the charged form of the molecules, in contrast, is entropy driven
[70–73].
It has extensively been discussed in the literature how far water–octanol systems can
be used to predict properties of transport through biological membranes. One of the forcible
arguments of opponents of this approach was the different nature of the driving forces of
the processes: the octanol–water system was classified as enthalpy-driven, whereas the lip-
id phase–water system should have entropic driving forces [78]. Unfortunately, the works
devoted to studies of the partitioning/distribution processes analysed only the change in
Gibbs energy (log P, log D). This approach does not provide the opportunity to understand
the mechanism of the process. However, from the present results it follows that the basic
differences claimed to exist between the thermodynamics of the transfer in octanol–water
and lipid–water systems do not exist. The nature of the driving forces of the processes as
well as the ratio between enthalpic and entropic terms is determined by an eventual charge
of the drug molecule, and the energetic state of this molecule within the respective phases.
In buffer at pH 7.4, the charged drug molecule interacts stronger with the solvation shell
(by additional electrostatic interactions) compared to the uncharged molecule. As a conse-
quence, more energy costs are needed for resolvation of a charged molecule in comparison
with uncharged molecules at partitioning/distribution. Moreover, the costs in enthalpy for
resolvation of a charged molecule are not completely compensated by the solvation en-
thalpy in the octanol phase. This fact may be an essential argument for the assumption that
drug molecules may transfer (during partitioning/distribution processes) with partly
Table 9 Thermodynamic characteristics of the transfer process from n-octanol to buffer (pH 2.0 / pH 7.4) of some NSAIDs at 25°C.
320
(±)-IBP DIF FBP KETO NAP
n-Octanol
0
– ∆G solv [kJ·mol–1] 40.2(f) 49.3 47.1 50.4 48.0
0
– ∆H solv [kJ·mol–1] 90.9(g) 108.6 86.5 82.5 107.1
0
– T∆S solv [kJ·mol–1] 50.7(h) 59.3 39.4 32.1 59.1
octanol pH2.0
∆Y tr = ∆Y solv – ∆Y solv
∆Gtr [kJ·mol–1] –27.7 (24.35)(d) (–25.7)(e) –28.0 –29.8 (–28.0)(d) (–23.8)(e) –23.2 (–21.96)(d) (–17.8)(e) –23.3 (–23.8)(d) (–20.0)(e)
∆Htr [kJ·mol–1] –4.4 (6.7)(d) (–6.1)(e) –10.8 –21.3 (–4.6)(d) (–15.6)(e) –6.7 (–1.7)(d) (–5.2)(e) –6.8 (0.0)(d) (–13.3)(e)
T ∆Str [kJ·mol–1] 23.3 17.2 8.5 16.5 16.5
ςH [%](a) 15.9 38.6 71.5 28.9 29.2
ςTS [%](b) 84.1 61.4 28.5 71.1 70.8
octanol pH7.4
∆Y tr = ∆Y solv – ∆Y solv
∆Gtr [kJ·mol–1] –25.8 –20.9 –22.5 –20.4 –17.2
∆Htr [kJ·mol–1 5.4 2.8 15.2 11.6 16.0
T∆Str [kJ·mol–1] 31.2 23.7 37.7 32.0 33.2
ςH [%](a) 14.8 10.6 28.7 26.6 32.5
ςTS [%](b) 85.2 89.4 71.3 73.4 67.5
log P2.0 (c) 3.50 4.44 4.16 3.12 3.34
log D7.4 (c) 1.07 0.76 0.85 –0.25 0.33
pKa(c) 5.2 3.3 4.6 4.6 4.15
(a)ς (b) (c) (d) (e) (f) –1
H = (|∆Htr|/(|∆Htr| + |T∆Str|))·100%; ςTS = (|T∆Str|/(|∆Htr| + |T∆Str|))·100%; Ref. [54]; Ref. [53]; Ref. [55]; 42.6 kJ⋅mol , Ref. [56];
(g)
84.2 kJ⋅mol–1, Ref. [56]; (h)41.4 kJ·mol–1, Ref. [56]
1.2 (+)-IBP (±)-IBP
1.0
FBP
0.8 DIF
0.6
log(D7.4)
0.4 NAP
0.2
0.0
KETO
-0.2
-0.4
90 95 100 105 110 115 120 125
-1
-∆Hsolv [kJ⋅mol ]
Figure 15 Dependence of the buffer/octanol distribution coefficient in the form of log D7.4 on sol-
vation enthalpy in buffer pH 7.4, ∆Hsolv .
retained solvation shells. The volume and structure of the “removing/accompanying” shell
is determined by the ratio of all the thermodynamic parameters.
It is interesting to compare the solvation characteristics with the experimentally deter-
mined partitioning properties of the drugs taken from the literature (Table 9). The depend-
0
ence of log D7.4 plotted versus H solv (pH 7.4) is shown in Fig. 15.
As the absolute value increases, the log D7.4 value decreases (with an exception for
KETO). Probably, this regularity is connected with considerable energetic costs for molec-
ular resolvation at the transfer step from the buffer to the octanol phase during the distribu-
tion. Because for buffer pH 7.4 a compensation effect between the solvation functions is
observed, the character of the correlation dependencies between log D7.4 values and entro-
pic term does not change. It should be noted that the value for KETO does not deviate from
the log D7.4 versus ∆G0solv correlation as the ∆H0solv does (Fig. 16). This fact confirms that
the enthalpic and entropic terms are more sensitive to the nature of the occurring processes
compared with Gibbs energy [57].
As a consequence of this fact, also the widely studied (and in many cases relatively
poor) correlations between Gibbs energy of drug – cyclodextrin complexation and log P are
of limited value as a measure of hydrophobicity, because comparison of Gibbs energies of
two different processes (complexation and partitioning/distribution) does not consider their
driving forces [70]. It is obvious that a good correlation can only be expected to be observed
in cases where the values and signs of the enthalpic and entropic terms of both processes
are identical.
Finally, let us consider the distribution/partitioning process from the point of view of
solvation. In order to transfer a molecule from one phase (buffer) to another (octanol) it is
necessary to overcome a potential barrier, which is “hypothetical” and equals the solvation
enthalpy in the buffer (Scheme 5). The height of this barrier determines the kinetic param-
eters of the partitioning/distribution process. Obviously, the outlined process does not in-
tend to desolvate the molecule completely: the resolvation process presents itself as a
1.2 (+)-IBP (±)-IBP
1.0
FBP
DIF
0.8
0.6
log(D7.4)
0.4 NAP
0.2
0.0
KETO
-0.2
-0.4
10 15 20 25 30 35
-∆Gsolv(pH 7.4) [kJ⋅mol-1]
Figure 16 Dependence of log D7.4 on Gibbs energy of solvation in buffer pH 7.4, ∆Gsolv (pH 7.4).
complicated process including the simultaneous destruction of the old shell and the creation
of a new one. As a consequence of this competition, the height of the activation barrier de-
creases considerably. The value of the activation barrier may be estimated from kinetic pa-
rameters of the partitioning/distribution process. This may in the future be helpful for
further characterisation of biopharmaceutical properties of drug molecules.
Scheme 5
7 Correlation between biopharmaceutically relevant parameters and

solvation characteristics
More than twenty years ago, Ochs et al. [43] paid attention to the inverse proportionality
between the lipophilicity of drugs (salicylic acid, antipyrine and amitriptyline) and the time
interval until the peak concentration occurs in the cerebrospinal fluid (CSF) postdose. Since
then, numerous works have confirmed this correlation between lipohilicity and the distri-
14
DIF
NAP
12
10
PHLm [h] 8
4 FBP
IBP KETO
2
ASA
0
-110 -105 -100 -95 -90 -85 -80

Oct -1
∆Hsolv [kJ⋅mol ]
Figure 17 Dependence of mean plasma half-life values, PHLm, vs. solvation enthalpy in octanol,
oct .
∆Hsolv
bution into CSF. This relation prompted us to carry out a correlation analysis between the
mean plasma half-life values, PHLm, [44] for the NSAIDs under investigation and the ob-
tained thermodynamic functions in octanol as a model for a lipophilic compartment. As it
follows from Fig. 17, a common trend line is derived for the solvation enthalpies of the not-
ed drugs in octanol, ∆H oct solv: good solvation in octanol corresponds to a long plasma
half-life. All drugs studied are carboxylic acids – and, therefore, all of them have a high
plasma-protein-binding affinity; correlations between the plasma-binding capacity of drugs
and their lipophilic–hydrophilic properties (expressed as log P) have already been ac-
knowledged [45]. The specific interaction between plasma proteins and the (acidic groups
in the) drug molecules is complex as there are quite a number of binding sites of different
affinity and hydrophobicity [46]. However, the plasma half-life may from a general point
of view be looked at as the equilibrium between the binding and the re-distribution of mol-
ecules from the (hydrophobic) plasma binding sites into the surrounding aqueous phase,
where similar thermodynamic prerequisites apply as for partitioning.
From Fig. 17, it can be seen that both ibuprofen and acetylsalicylic acid have a lower
plasma half-life compared to flurbiprofen and ketoprofen. It may be supposed that this dif-
ference in behavior is related to their being close derivatives of benzene, whereas all the
other substances have two benzene rings. This would in particular indicate that the interac-
tions of major importance in the redistribution of the drug molecules from the plasma pro-
tein binding into the surrounding aqueous phase are van-der-Waals forces. On the other
hand, Fig. 17 can be expressed for flurbiprofen and ketoprofen in terms of a somewhat long-
er plasma half-life.
Anyhow, the present approach may be useful in estimating plasma half-life values
from thermochemical measurements of solvation. A connection between the quality of sol-
vation in octanol and the plasma half-life seems reasonable as there may be some basic sim-
ilarity in the nature of the drug–octanol and drug–plasma protein interactions.
Acknowledgments
This work was supported by Norges Forskningsråd, project number HS 58101, and a
personal grant for GP from the Russian Science Support Foundation.
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Окончательный вариант. Авторам: одобрить (или изменить) выделенное
красным -- как и всю главу в целом
На стр. 5 (в середине, тоже выделено красным) -- > was studied for days...
сколько дней ?
9 Biodamage of Materials:
Adhesion of
Microorganisms on the
Surface of Materials
K.Z. Gumargalieva1, I.G. Kalinina1, S.A. Semenov1

and G.E. Zaikov2
1N.N.
Semenov Institute of Chemical Physics, Russian
Academy of Sciences, 4 Kosygin Street, Moscow 119991,
Russia
2N.M. Emanuel Institute of Biochemical Physics, Russian
Russia; chembio@sky.chph.ras.ru
We studied adhesion interaction of the most widespread species of microscopic

fungi Aspergillus niger, A. terreus, Trichoderma viride and Penicillium funiculo-
sum with surfaces of materials (polymers, metals). The force of adhesion
interaction was measured by the method of centrifugal detachment. The adhesion
microorganism–metal surface macroscopic characteristics were obtained based
on the analysis of kinetic curves. By the example of A. niger, the stochastic nature
of adhesion of microbial cells was shown to be caused by the heterogeneity of
support surface and size of conidia; their distribution by forces of adhesion was
shown to obey the Gaussian law. The fungal cell-wall structure was found to
change as a function of age, and the change of force of adhesion interaction to
correlate with changes in the cell-wall structure. The dominating role in adhesion
interaction strengthening was established to be played by an increase of the
concentration of albuminous components in the cell surface layer.
Keywords: adhesion, adhesion interaction, adhesion force, polymer surfaces, microorgan-

ism, Aspergillus niger, A. terreus, Trichoderma viride, Penicillium funiculosum
328 K.Z. Gumargalieva, I.G. Kalinina, S.A. Semenov and G.E. Zaikov
Introduction
Problems set by Professor N.M. Emanuel in the field of applied works were characterized
by a fundamental approach. In particular, at the beginning of 1980s the issues of polymer
materials’ biodamage traditionally related to microbiology were included by N.M. Emanuel
into the chemical destruction section of polymer materials science. Subsequent investiga-
tions in this field showed active destructing agents of materials in interaction with micro-
organisms to be products of their vital functions – metabolites representing famous
chemical agents: water, salts, acids, alkali, enzymes, toxins, i.e., biodamage proceeds by
laws of chemical destruction [1–10].
Investigation of the results of biodestruction generally concerns two stages: adhesion
or attachment of microorganisms to the surface of material and growth of microbial biomass
as the result of substrate-support utilization. These two stages should predetermine further
processes of material degradation.
Given the current ecologically fraught environment, fast adaptation of various micro-
bial species to the changing environmental conditions and accumulation of various wastes
of synthetic origin, there is a requirement in research and development of both rapidly de-
grading materials and materials resistant to biodeterioration [11–12].
Studies of microbial cell adhesion are of great interest now, but in many cases these
investigations are “exotic” as far as the selection of microbial species is concerned. The au-
thors of [13] studied selective adhesion of extremophile cultures to micaceous plates coated
with polyethyleneimine. Using the method of scanning electron microscopy, adhesion of
extremophiles to a polyethyleneimine coating was found to be preferred in comparison to
that to a polylysine coating. The main physicochemical factors of material surface such as
hydrophobicity and roughness and the morphology of microbial cells are discussed in
[13–16]. These works and the current state of the problem confirm the necessity of inves-
tigating the nature of adhesion interaction on the material–microbial cell wall interface with
the view of a quantitative estimation of this process.
1 Results and discussion

Adhesive cells of microorganisms act as aggressive bioreagents by secreting exoenzymes
or other low-molecular-mass substances and producing the so-called biofilm. That is why
quantitative parameters of adhesion are determinative for rates of biofouling (biomass ac-
cumulation) and biodestruction.
Polymer surface–microorganism interaction was determined by the value of adhesion
force in relation to the time of contact at various external conditions (temperature, moisture)
to polymer materials of various degrees of hydrophilicity: polymethyl methacrylate, cello-
phane, polyethylene, acetylcellulose, epoxide resin and polyethylene terephthalate.
Conidia of the microscopic fungus Aspergillus niger were plated on surfaces of mate-
rials in doses from water suspension with a certain titre by a microbatcher. Then the conidia
were dried and aged under various thermo-moist conditions in the interval from 0 up to 24 h.
The value of the force of conidial adhesion to surfaces of polymer materials was determined
by the method of centrifuge detachment [17]. Changes of morphology, (shape) and size of
conidia were studied by the method of scanning electron microscopy (Tesla BS300). The
number of conidia remaining on the surface of material after the action of a centrifugal force
(angle rate of rotor ω = 15,000 revs for 15 min) was counted in Goryaev’s chamber.
Change in the number of residual A. niger conidia, γ = N/N0, after centrifugation in
Biodamage of Materials: Adhesion of Microorganisms on the Surface of Materials 329
100
5
Adhesion number, % 4
80 3
2
60 1
40
20
0
0 5 10 15 20 25 30 35 40 45 50
Time, h
Figure 1 Kinetic curves of adhesion of A. niger conidia to various polymer materials at a tempera-
ture of 22°C and relative moisture of 30%. 1, polyethylene; 2, epoxide resin; 3, polymethyl methacry-
late; 4, acetyl cellulose; 5, cellophane.
relation to the time of preliminary ageing under given thermo-moist conditions (T = 22°C,
ϕ = 30%) is presented in Fig. 1. Adhesion interaction of the polymer material–micro-
organism system is of kinetic character. Each polymer material–microorganism system is
characterized by the time of completion of adhesion forces’ formation between conidia and
the surface (approaching the plateau in the kinetic curve), which depends on external
conditions.
The dependences presented in Fig. 1 are well described by eq. (1):
ln γ/γ ∞ = – K ⋅ t , (1)
where γ and γ∞ are, respectively, the number of particles remaining on the surface of poly-
mer material after preliminary ageing for time t, and of those irreversibly adherent under
given conditions; K is the rate formation constant for adhesion forces between conidia and
the surface of material. The parameter values determined from the equation for polymers
studied and also the value of force per cell are given in Table 1. At ω = 15000 min –1 the
force influencing each conidium was equal to F = 1.2·10 –4 dyn/cell. As it is known [18]
the value of force under adhesion corresponds to γ = 50%. If we know F50, the time of the
kinetic curve approaching the plateau and the exponential course of kinetic curves of adhe-
sion, we may determine the values of adhesion forces for each polymer under given thermo-
moist conditions.
Thus, under fixed thermo-moist conditions each material may be characterized by a
proper set of values of K and F. With respect to the strength of A. niger conidium adhesion
Table 1 Parameters of adhesion of A. niger conidia to the surface of polymer materials.
Material k, sec–1 F, dyn/cell γ∞, %

Polyethylene 1.66·10–5 3.3·10–4 55
Epoxide resin 2.20·10–5 6.6·10–4 70
Polymethyl methacrylate 1.0·10–4 1.1·10–3 80
Cellophane 1.0·10–4 1.6·10–3 85
Acetyl cellulose 1.3·10–4 2.5·10–3 90
100
5
4
3
Adhesion number, %
80
60 2
1
40
20
0
0 10 20 30 40 50
Time, h
Figure 2 Kinetic curves of adhesion of A. niger conidia to polyethylene and cellophane at a relative
moisture of 30% and various temperatures: 1, polyethylene, T = 38°C; 2, polyethylene, T = 22°C; 3,
polyethylene, T = 10°C; 4, cellophane, T = 22°C; 5, cellophane, T = 10°C.
the investigated polymer materials can be arranged in the following sequence: polyethyl-
ene, epoxide resin, polymethyl methacrylate, cellophane, lavsan, acetyl cellulose.
As we mentioned above, polymer materials with various degrees of hydrophilicity
were used in experiments. It is obvious from Fig. 1 and Table 1 that hydrophilic polymers
have high values of adhesion force and formation rate constants, whereas for hydrophobic
polymer materials these values are significantly lower. Obviously, the values of K and F
are determined by the ability of material to strong adhesion of microorganisms in relation
to the moisture capacity of material.
For a more detailed understanding of the process, we investigated the influence of ex-
ternal operational factors on adhesion. The kinetic curves of adhesion on polyethylene and
cellophane at various temperatures of conidial ageing on the polymer surface are presented
in Fig. 2. It is obvious that with a temperature rise, adhesion of A. niger conidia to polymer
materials and the rate of adhesion bond formation are decreased, which is characteristic of
physical phenomena.
The values of the characteristics of adhesion interaction for polyethylene and cello-
phane at various temperatures and relative moisture are presented in Table 2.
As is obvious from Fig. 2 and Table 2, the adhesion force (F) and the constant of ad-
hesion force formation rate are observed at a 98% relative air moisture. Obviously, the
Table 2 Adhesion parameters of interaction for polyethylene and cellophane at various thermo-
moist conditions.
T, °C φ, % Polyethylene Cellophane
γ∞, % k, sec–1 F, dyn/cell γ∞, % k, sec–1 F, dyn/cell
0 70 2.20·10–5 3.0·10–4 – – –
10 30 85 1.66·10–5 5.2·10–4 90 1.0·10–4 2.5·10–3
98 100 9.70·10–5 1.9·10–3 – – –
22 30 55 1.66·10–5 3.3·10–4 85 1.0·10–4 1.6·10–3
38 30 50 1.66·10–5 1.3·10–4 – – –
120
Adheson number, % 100 3

80 2
1
60
40
20
0
0 10 20 30 40 50 60
Time, h
Figure 3 Kinetic curves of adhesion of A. niger conidia to polyethylene at 10°C and a relative mois-
ture of 0 (1), 30 (2) or 100% (3).
conidia of macroscopic fungi in an air medium interact with the surface of polymer material
at the expense of molecular forces and at the action of capillary forces of liquid condensed
in the gap between the conidium and polymer surface under the action of forces of Coulomb
interaction and other causes. In the presence of air moisture, the condensation of vapor oc-
curs between the conidium and polymer surface. Capillary forces are the greater, the higher
the surface tension of liquid whose vapor surrounds the area of interaction of conidia with
polymer surface, where the moistening of the polymer surface is better. The liquid interlay-
er between interacting objects (in this case between conidia and polymer surface) excludes
or to a great extent weakens the action of forces of electric nature. That is why the results
of experiments carried out with polyethylene at a fixed temperature 10°C but at various air
moistures (see Fig. 3 and Table 2) confirm the physical character of adhesion interaction of
the system. To confirm the assumption, adhesion interaction of A. niger on the surface of
polyethylene at 22°C and relative air moisture of 30% was studied during days on a raster
electron microscope. An increase of interaction time leads to a change of the geometric
shape of A. niger conidia and an increase of the contact area by almost 100%. Probably, this
fact may explain the kinetic character of conidial adhesion to polymer surfaces that allows
a quantitative description of microbial adhesion to various surfaces and its characterization
by kinetic parameters.
The process of microbial adhesion is also determinative in biofouling of metals in wa-
ter and air media [19–22]. Those experiments made use of conidia of Tr. viride fungi, which
occur in water media and possess optimal sizes for quantitative microscopic analysis. They
determined the force, causing the detachment of 50% of conidia from the general number
of cells attached to the metal surface, from the integral curves for the distribution of conidia
by adhesion forces characterizing the dependence of part of detached particles on the force
of detachment.
The force of detachment of 50% of conidia was calculated to be
F 50 = π 3 /675·R(ρc – ρav)·ω 2 ·r 3, dyn/cell, (2)
where ρav is the density of the medium (water); ρc is the density of conidia equal to 1.15
g/cm3, R = 4 cm is the radius of the centrifuge rotor, ω is the number of revolutions, r is the
100 1
2
Adhesion number, %
80 3
4
5
60 6
40
7
20
0
0 5 10 15 20 25
Time, h
Figure 4 Curves of the increase of adhesion of Tr. viride conidia to metals: 1, zinc; 2, copper; 3, alu-
minium; 4, nickel; 5, titanium; 6, tantalum; 7, molybdenum.
conidium radius equal to (3 ±0.3)·10 –4 cm. The adhesion of conidia to a metal surface was
characterized by two parameters γ∞ and F50. The curves of adhesion of Tr. viride conidia
to various metals are presented in Figure 4, the values of γ∞ and F50 are given in Table 3.
By adhesion parameters, all metals may be divided into two groups: the first is char-
acterized by F50 ≥ 10 –4 dyn/cell; the second, by F50 from 10 –5 up to 10 –7 dyn/cell.
Prehistory of surface plays a significant role; e.g., for nickel treated by cold plastic de-
formation γ∞ = 79±8%, and F50 = 2.0·10 –4 dyn/cell, and for nickel aged for 1 h at a tem-
perature of 700°C and pressure 10 –4 mm Hg, γ∞ = 50±2%, and F50 = 8.2·10 –4 dyn/cell.
Adhesion is observed to be minimal on gold, tungsten and molybdenum; the maximum
is on zinc, lead and copper. Aluminium and titanium occupy an intermediate position.
On the whole, the sequence of a limited number of adhesion is the following: lead,
zinc, copper, aluminum, nickel, silver, titanium, tantalum, platinum, molybdenum, gold,
Table 3 Adhesion parameters of Trichoderma viride conidia at T = 22°C and ω = 15000
revolutions in water medium to metals treated in solvent by a − boiling, b − at 22°C.
No. Metals a b
γ∞, % F50, % γ∞, % F50, %
1 Aluminum 95±5 – 82±5 (88B) 3,0·10–4

2 Tungsten 0 1.86·10–7 0 1.9·10–7
3 Gold – – 25±5 2.3·10–5
4 Copper 85±3 4.1·10–4 93±7 5.8·10–4
5 Molybdenum – – 34±5 7.4·10–5
6 Nickel 62±5 1.67·10–4 79±8 2.0·10–4
7 Platinum – – 53±5 2.2·10–4
8 Lead 96±2 7.00·10–4 100±1 7.0·10–4
9 Silver – – 78±5 5.0·10–4
10 Tantalum 63±8 2.97·10–4 65±10 2.5·10–4
11 Titanium 76±5 1.35·10–4 71±3 9.0·10–5
12 Zinc 100 – 97±3 6.7·10–4
100
60
4
40
3
20
2 5
1
0
10 15 20 25 30 35 40
Angle rate, min_1
Figure 5 Distribution of adhesion number (γ) of A. niger conidia in relation to the angle rate (ω) on
polyethylene (1, 2) and cellophane (3, 4) at 22°C and ϕ = 98%. 1 and 3, ageing time 4 h; 2 and 4, age-
ing time 24 h; 5, distribution of polystyrene particles in relation to ω in an integral form under the
same adhesion conditions.
tungsten; and by a decrease of values of force of 50% detachment of conidia: lead, zinc,
copper, silver, aluminium, tantalum, platinum, nickel, titanium, molybdenum, gold, tung-
sten.
Thus, Tr. viride conidia reveal a low level of adhesion to metals, which are not oxi-
dized under experimental conditions and do not form oxides at all, e.g., gold.
The results obtained testify that the main characteristic of adhesion – the force of ad-
hesion – increases in time reaching an equilibrium value and is determined by the nature of
material surface and type of microscopic fungi. The experimental fact of the distribution of
adhesion forces in relation to applied stress was also noticed. This effect was studied earlier
and probably is caused by both the heterogeneity of the surface of material and heteroge-
neity of conidia by size. The dependences of the number of A. niger conidia remaining on
the surface of a material after centrifugation (adhesion number γ ) on the angle rate of rota-
tion for polyethylene and cellophane at a temperature T = 22°C and ambient moisture ϕ =
98% are presented in Fig. 5. From the data obtained, using eq. (2) we calculated forces of
50% detachment of conidia, F50. The experimental results show that at time of ageing
conidia on a polymer surface for 4 and 24 h the adhesion force Fa (force of 50% detachment
of conidia F50) is increased for polyethylene 2 times, and for cellophane 1.5 times. This fact
obviously testifies to a significant difference in the constants of adhesion force formation
rates for the same system (Tables 1 and 2).
For a more detailed investigation of the dependences of the adhesion number on the
number of revolutions γ = γ (ω) (in implicit form it is the dependence of adhesion number
on applied force γ = γ (F) that represents the distribution by adhesion forces) the derivative
of this function was determined for polyethylene γ (F) = dγ /dF(F) (Fig. 6). The dependence
presented in Fig. 6 is described by an equation of the type corresponding to the Gaussian
distribution:
ϕ (x) = Aexp (– α x 2) in coordinates ϕ = ϕ (x), (3)
where A and α are constants.

Such a distribution may be caused by both heterogeneity of conidia by size and heter-
ogeneity of polymer surface. With this in mind, the size of conidia A. niger was determined
19 2
17
15
Y, units
13
11
9 1
7
5
3
0 0.005 0.01 0.015 0.02 0.025 0.03 0.035 0.04 0.045
F, dyn/cell
Figure 6 1, Distribution of A. niger conidia by forces of adhesion on polyethylene in differential

form; ageing time, 24 h; 2, distribution of polystyrene particles by adhesion forces in differential
form. T = 22°C, ϕ = 98%.
40
30
N
20
10
0
1 2 3 4 5 6 7
r, µm
Figure 7 Distribution of A. niger conidia by radius.
on a TESLA BS 300 raster electron microscope. The measured values of conidial diameters
on electron micrographs demonstrate a significant scattering by size (Fig. 7) that should in-
fluence the scattering of adhesion force values.
Let us show that the distribution of conidia by size contributes (i.e. is described by the
same function) into the distribution of adhesive conidia by force.
We present the derivatives of adhesion number by radius and force:
γ (r) = dγ /dr(r); γ (F) = dγ /dF(F).
We should prove that fromγ = γ (r) it follows that γ = γ (F). Let us write the distribution
by conidial size in the following form:
d γ /dr = dγ /dF dF/dr = Kdγ /dF , (4)
To determine the value of K, we present as dγ /dr from eq. (2):

100 2
80
1
60
40
20
0
0 10 20 30 40 50 60 70 80
Time, h
Figure 8 Kinetic curves of adhesion of polystyrene particles to polyethylene (1) and cellophane (2)
at T = 22°C, ϕ = 30%, ω = 15 000 min –1.
dγ /dr = 3F/r dγ /dF; K = 3F/r. (5)
It follows from the experimental data (the dependences F = F(r) and γ = γ (r)) that for
all values of radiuses (r) of conidia at the functioning force of detachment from F1 =
5.5·10 –3 dynes per cell up to F2 = 2.3·10 –2 dynes per cell (Fig. 6) the values in the
right-hand side of eq. (5) will be
3F/r dγ /dF = 4.6±0.3,
and in the left-hand side of eq. (5):
dγ /dr = 4.7±0.3.
Within the limits of experimental error these values are equal, i.e., the distribution of
conidia by size is one of the causes of the real distribution by adhesion force. The hetero-
geneity of support should also contribute to the distribution of particles by adhesion force.
To elucidate this, we carried out experiments with spherical polystyrene particles (di-
ameter d = 2.8 µm; Serva). The kinetic curves of adhesion and distribution of particles by
adhesion force obtained in the experiments are given in Fig. 8.
By their character, the kinetic curves and curves of distribution of conidia A. niger and
polystyrene particles are similar, although there are some differences in parameters of ad-
hesion interaction (Figs. 4 and 6). The constants of adhesion force formation for polystyrene
particles have high values in comparison with A. niger conidia. Moreover, the scattering of
adhesion force values γ in the case of polystyrene particles is significantly lower.
The results of the experiment suggest that adhesion interaction of system polystyrene
particles–polymer changes in time; the distribution of particles by adhesion forces is de-
scribed by the Gaussian equation. Consequently, the picture of the adhesion of polystyrene
particles and A. niger conidia to the polymer surface is analogous, which may be explained
for polystyrene particles by the following:
• an increase of contact area of polystyrene particles with polymer surface causes the
rise of adhesion force in time at the expense of deformation of the surface;
• capillary concentration of moisture in the contact zone also promotes an adhesion
force;
• energetic heterogeneity of the surface of polymer material causes the distribution of

polystyrene particles by adhesion forces.
Thus, the adhesion of A. niger conidia is of stochastic nature caused by their hetero-
geneity by size and the heterogeneity of the polymer support; the distribution of conidia by
adhesion forces obeys the Gaussian distribution. The quantitative characteristics of the in-
teraction of microscopic fungi with solid polymer surfaces may serve as an estimative cri-
terion of microdestruction processes in materials.
In fact, the adhesion process should also be determined by the parameters of the conid-
ial external wall (the cell-wall structure) and by the ability of conidia to secrete various or-
ganic substances (exoenzymes, organic acids, etc.) [9, 10]. To establish a correlation
between the adhesion properties of conidial microdestructors of polymer materials and the
structure of the cell wall of various ages of cultures, samples obtained by washing out from
the surface of conidia A. niger, A. terreus, P. funiculosum, Tr. viride agar, deposited on Syn-
por membrane filters (thickness of layer ~ 0.01mm) and dried in exsiccator over CaCl2 were
studied by the method of IR spectroscopy with Fourier analysis.
The concentration of analyzed substances was estimated according to Lambert–
Bouguer–Beer law with the help of the following equation:
C = D/l·E,
where l is the depth of beam penetration and E is the extinction coefficient.

Table 4 Characteristics of IR spectra of conidia of fungi of various ages.
Age of conidia, days

Microscopic
fungi ν, cm–1 3 15 30
D C, % D C, % D C, %
A. niger 1550–1560 0.01 5 0.08 58 0.07 56

1275–1278 0.84 71 0.20 34 0.20 34
831–835 0.78 24 0.14 8 0.15 10
A. terreus 1550–1560 0.10 37 0.07 30 0.08 32
1275–1278 0.54 47 0.56 54 0.55 50
831–835 0.48 16 0.50 16 0.52 18
P. funiculosum 1550–1560 0.05 5 0.01 1 0.02 2
1275–1278 0.57 70 0.36 73 0.35 71
831–835 0.57 25 0.39 26 0.40 27
Tr. viride 1550–1560 0.12 45 0.06 28 0.05 26
1275–1278 0.48 42 0.38 54 0.39 56
831–835 0.42 13 0.41 18 0.41 18
For comparative quantitative analysis of IR spectra, the absorption bands significantly

changing in intensity in the course of the growth process should be chosen. The following
bands were selected: 1545–1550 cm –1 (amide 2) – deformational vibrations of the
NH-group; 1275–1280 cm –1 – stretching vibrations P=O of phospholipids; 831–835 cm –1
– deformational non-planar vibrations of the CH-group of α-glycanes.
The values of optical densities and concentrations of analysed substances for four
types of conidia of microscopic fungi are presented in Table 4. In the case of some types of
c
1.0
0.7
a b
1.0
0.5 0.5
0.5
2000 1000 400

0.9
f
2000 1000 400 2000 1000
0.9
1.6 d e
1.0 0.5 0.5
2000 1000 400 2000 1000 2000 1000 400

1.4 1.0
g h
1.0
0.5
2000 1000 400 2000 1000 400
Figure 9 IR spectra of the surface layer of conidia: a, A. niger, 3 days; b, A. niger, 15 days, 30 days;
c, A. terreus, 3 days; d, A. terreus, 15 days, 30 days; e, Tr. viride, 3 days; f, Tr. viride, 15 days, 30 days;
j, P. funiculosum, 3 days; h, P. funiculosum, 15 days, 30 days.
conidia in the course of growth noticeable quantitative changes in the structure of cell walls
occur, and for other types significant changes were not noticed.
IR spectra of the surface layer of A. niger conidia at various development times are
presented in Fig. 9 as an example. The IR spectra of 15- and 30-day-old conidia coincide
almost completely, which is explained by the known factor of the decay of the metabolic
processes and by the cell quiescent state already after 13–14 days. So, for conidia of
Tr. viride type the concentration of amides in the surface layer of the cell decreased from
45% (age, 3 days) down to 26–28% (age, 15 days). A weak rise of the concentration of
Adhesion number, %. 100
80
60
40
20 2
1 3
0
0 5 10 15 20 25 30
Angle rate, min_1
Figure 10 Distribution of the adhesion number (γ) of A. niger conidia in relation to the angle rate
(ω) on polyethylene at T = 22°C and ϕ = 98%. The ageing time of conidia on the surface of polyeth-
ylene, 24 h. 1, 3-day-old conidia, 2, 15-day-old conidia, 3, 30-day-old conidia.
phospholipid (from 42 up to 56%) and glycane (from 13 up to 18%) components is ob-

served. Probably, the synthesis of these substances is continued for more than 3 days.
When considering the IR spectra of the surface layer of P. funiculosum and A. terreus
conidia, it is obvious that there are no significant changes in their spectra. Comparison of
the spectral characteristics of the walls of 3, 15 and 30-day-old conidia of these types of
microscopic fungi shows that the surface layers of these cells do not differ in chemical struc-
ture. The concentrations of amide, phospholipid and glycane compounds were practically
unchanged.
Analysis of the spectra of the surface layer of A. niger conidia of various ages shows
that at the stage of growth the concentration of albumin components is increased (from 5
up to 58%) and phospholipid (from 71 down to 34%) and α-glycane (from 24 down to 8%)
concentrations are decreased. Probably, albumin synthesis in the surface layer of A. niger
intensively proceeds for 15 days. In particular, this type of conidia was selected for inves-
tigation of their adhesion to polymer materials.
Integral distribution of A. niger conidia of various ages by adhesion forces to polyeth-
ylene at a temperature of 22°C and relative moisture of 98% is given in Fig. 10. As is seen,
the curves of 15- and 30-day-old conidia totally coincide. The force of adhesion of these
conidia is as follows:
F5015 = F5030 = (1.1±0.02)·10 –2 dyn/cell,
where F5015 and F5030 are the forces of 50% detachment (γ = 50%) of 15- and 30-day-old
conidia correspondingly, that is 3.4 times higher for adhesion force in the case of 3-day-old
conidia.
For 3-day-old conidia, the adhesion force is:
F503 = (3.2±0.2)·10 –3 dyn/cell.
Comparing the data on the adhesion of 15- and 30-day-old conidia with the IR spectra
of their surface layer, we can see that after 14– 15 days of cultivation of the fungi all pa-
rameters are stabilized, metabolic processes decay and cells pass into a quiescent state.
For small-size conidia (A. terreus, P. funiculosum) the concentrations of the main com-
ponents of the cell wall practically do not change, whereas for conidia of large size (T. vir-
ide, A. niger) such changes are observed (Table 4). Obviously, for conidia of small size the
surface layer of the cell is formed in the time not exceeding 3 days. For conidia of larger
size (for example, A. niger) the process of surface layer formation does not cease in 3 days
and continues for a longer time. At the first moment of adhesion, albumin macromolecules
evolve intensively [11]. For the case of A. niger conidia, the albumin component is in-
creased in the course of cell growth and their adhesive interaction also increases.
Thus, experimental material suggests that the adhesion force of conidia of microscopic
fungi in the course of their growth changes differently. It depends on the change of the con-
centration of albumin components in the surface layer of the cell having a mosaic structure.
In the cases when these changes are significant (predominantly for large-size conidia) the
force of adhesion interaction with surfaces increases and if there are no clear changes (pre-
dominantly for small-size conidia) the adhesion force does not change.
References
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Polymers, Moscow: Nauka (1982) (in Russian).
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Materials from Biodamage, N. Novgorod (1991) (in Russian).
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Appl. Phys. Colloids and Surfaces A: Physicochemical and Engineering Aspects, 249,
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Boston: Int. Union of Microbiol. Soc., 75 (1982) (in Russian).
10 Controlled Release of
Aseptic Drug from
Poly(3-hydroxybutyrate)
Films: A Combination of
Diffusion and Zero-order
Kinetics
R.Y. Kosenko1, Y.N. Pankova1, A.L. Iordanskii1*,

A.P. Bonartsev3, and G.E. Zaikov2
1Semenov
Institute of Chemical Physics, 4 Kosygin Street,
B-334, Moscow 119991 Russia
iordan@chph.ras.ru
2Emanuel Institute of Biochemical Physics, 4 Kosygin Street,
B-334, Moscow 119991 Russia
3Biological Department, Moscow State University,
6 Leninskie Gory, Moscow 119991 Russia
A polymer system based on biocompatible and biodegradable poly(3-hydroxy-

butyrate) (PHB) has been elaborated for controlled release of furacillin (Fr). The
kinetics of release from membranes of PHB loaded with 0.5–50 wt. % Fr into an
aqueous medium has been investigated by UV spectroscopy at 25°C. The release
profiles comprise diffusion and kinetic impacts.
The diffusion component of the release has been analyzed and the diffusi-
vity dependence on the drug concentrations has been determined. The release
kinetic constant is directly related to the hydrolytic destruction of PHB and the
dependence on the initial concentration of the drug. The destruction of PHB is
* To whom correspondence should be addressed

342 Controlled Release of Aseptic Drug from Poly(3-hydroxybutyrate) Films
clearly demonstrated by long-term experiments (after first week of release). These

results are required for further elaboration of novel drug delivery systems
including a combination of several drugs that will render a combined action on
tissues and organs of biological systems.
Keywords: poly(3-hydroxybutyrate) films, controlled release of drugs
Introduction
Dramatically increasing costs of hydrocarbon raw materials stimulate the development of
new polymers, which do not depend on production of oil and gas. Fermentative biosynthesis
of poly(3-hydroxybutyrate) (PHB) and its homologues poly(3-hydroxyalkonoates) (PHAs)
is based on using renewable substrates. Hydrocarbon wastes of food and wine/juice indus-
tries (sugars, molasses, starch etc.) are the basic “structural material” for bacterial PHB (and
PHA). Utilization of hydrocarbons in biosynthesis of PHA is ecologically efficient.
In the recent decade, PHB and its copolymers have begun to be productively used in
medicine [e.g., 1–3]. For example, composites of PHB are characterized by a high biocom-
patibility to bone tissues that enables their use as bioresorbable osteo implants [4]. Modified
PHB works as a highly effective scaffold in tissue engineering and promotes proliferation,
adhesion and production of cells [5, 6]. PHB–PEG blends [7] have a good hemocompati-
bility. It is reported [8] that, in contact with blood, surfaces of PHB and PHBcoHV films
do not activate hemostatic changes on the cell level. Great progress for poly(4-hydroxybu-
tyrate) (P4HB) is observed in cardio implantation [9]. Artificial heart valves produced by
stereolithography [10] with P4HB and controlled by x-ray tomography have demonstrated
a relevant combination of mechanical properties and hemocompatibility [11]. Fabrication
of a stent based on PHB has been reported [12].
For this reason, PHB and its derivatives can be considered to be new promising med-
ical materials for tissue engineering [13], design of osteoprostheses with replacement of
biodegradable material by germinated bone tissues [14], and hemocompatible coatings for
cardiovascular surgery [15]. Within the framework of this paper, it should be emphasized
that there is a broader area of PHB applications: design of matrices, reservoirs, and micro/
nano-particles for controlled drug release [16–18]. In this case, information on the biocom-
patibility, rate of resorption and diffusion characteristics of polymer systems is required.
The aim of this paper is to design and study a therapeutic PHB system loaded with an aseptic
(furacillin) and intended for the release of the drug into biological aqueous media.
Recently [19] we have shown that water diffusion in PHB films 40-60 µm thick is com-
pleted in several tens of minutes, whereupon the films absorbed the limiting equilibrium
concentration of water (ca. 1 wt. %). Structural relaxation in PHB under humid conditions
is achieved after a longer period of time (nearly 1000 min). We have investigated the kinet-
ics of the release for several tens of days, therefore, to a first approximation, water transport
phenomena in PHB are not essential. However, the long-term kinetics of drug release from
PHB films has an intricate form and requires special analysis for both diffusion modeling
and drug delivery applications.
1 Experimental
We used PHB from Biomer (Lot F16). The initial PHB powder was solved in chloroform
under long-term boiling. The hot polymer solution was filtered, and the molecular weight
R.Y. Kosenko, Y.N. Pankova, A.L. Iordanskii, A.P. Bonartsev, and G.E. Zaikov 343
of PHB was determined by the viscosimetry technique in accordance with the procedure
described in [20]. The averaged value of MW is 183.5×103 g/mol. As an aseptic drug we
used furacillin (MW = 198 g/mol)
The basic characteristics of the polymers include density = 1.25 g/cm3, Tm = 178°C,
Tg = 9°C, degree of crystallinity = 70% (determined by WAXS data).
Films of PHB containing furacillin were prepared in a two-stage procedure. 1 g of
powder was mixed with 50 ml of dioxan and boiled in a retort with reverse glass refrigerator
as well. Then after cooling and removing dioxan by a vacuum pump, PHB and Fr were
solved in chloroform, which was followed by casting the film. The thickness of cast PHB
films varied from 120±10 µm to 180±15 µm, and the concentration of loaded Fr changed
in the sequence 0.5 > 1.0 > 1.5 > 1.75 > 2.0 > 3.0 > 5.0 wt. %. The drug release profiles of
PHB were registered in water and phosphate buffer (pH = 7.4) by the UV technique using
a Beckman DU65 UV spectrophotometer at 25°C.

Typical kinetic profiles of Fr release from PHB films are shown in Fig.1. As is clear from
the graph, for PHB release systems loaded by the drug at concentrations exceeding 1% there
are no constant limiting values of equilibrium concentration that would be typical of a dif-
fusion picture according to Fick’s law. These kinetic curves are characterized by the initial
nonlinear range and final range where the drug release profile is linear relative to time (ze-
ro-order kinetics). Analysis of the data in Fig. 1 shows that the superposition of the proper
diffusion and a linear kinetic process defines the complicated character of release. Most
clearly the linear ranges are manifested after compilation of drug diffusion and are observed
for 8–10 days.
0.45
0.40 5
0.35
0.30
0.25 3
Dt
0.20
0.15
0.10 1
0.05
0.00
0 5 10 15 20 25 30
time, days
Figure 1 Kinetic profiles of aseptic release. The figures show the initial concentrations of the drug
(wt. %); Dt is the optical density of the drug in a liquid medium at 373 nm.
1.0 1
2
0.8
3
0.6
Gt/G00
0.4
0.2
0.0
0 5 10 15 20 25 30 35 40
(Time)1/2, (h)1/2
Figure 2 Impact of diffusion in the total drug release kinetics for PHB films (thickness, 180 µm).
Initial concentrations of the aseptic are (1) 1.75 wt. %, (2) 2.5 wt. %, (3) 3 wt. %; (5) 5 wt. %.
Based on aforesaid, the release profile is described by the following equation
∂Ct/∂t = D[∂2Ct/∂x2] + k, (1)
where D is the drug diffusion coefficient, cm2/sec; k is the kinetic constant of hydrolytic
destruction, sec –1; Ct is the concentration of the drug in the polymer, wt. %; x (cm) and t
(sec) are the coordinate and time of diffusion, respectively.
After subtracting the linear kinetic term (k · t) from the integral profile of release
Ct – kt ≡ Gt, (2)
the diffusion equation has the classical view
∂Gt/∂t = D[∂2Gt/∂x2]. (1a)
The solution of Eq. 1a provided that Gt/Goo > 0.5 has the classical Fick’s form
Gt/Goo = 1 – (8/π2⋅exp (–π2Dt/L2), (3)
where L is film thickness, cm; Gt is the concentration of the drug available for diffusion,
and other parameters were determined in Eq. 1.
Finding the logarithms of the left- and right-hand side parts of Eq. 3 and solving this
equation graphically in coordinates log (1 – Gt/Goo) – t:
log (1 – Gt/Goo) = log (8/π2 ) – π2Dt/L2 , (4)
we can calculate the diffusion coefficients of Fr in the PHB films. Numerical subtraction
of the linear contribution to the release kinetics from the total concentration of the drug
3.0
Mobile fraction of furacillin, wt. %

2.5
2.0
1.5
1.0
0.5
0.0
0 1 2 3 4 5
Total furacillin concentration, %
Figure 3 Relation between total and mobile concentrations of the drug in PHB films.
(shown in Fig. 1) was performed to separate the diffusion and kinetic components. After
computer calculation, the proper diffusion contribution is shown in Fig. 2. Here the results
are presented in classical diffusion coordinates: Gt/Goo – t1/2. We emphasize two features
of drug release profiles: 1) the linear character of the data in the above diffusion coordinates
holds within the limits 0 < Gt/Goo < 0.65 that attests a predominance of diffusion at the ini-
tial stage of release; 2) the slope of the lines in the same coordinates depends on the initial
concentration of the drug that demonstrates its concentration diffusivity dependence. Lim-
iting (equilibrium) values of the drug (Goo) dissolved in PHB films are needed to plot the
curves in Fig. 2. Additionally, these values show the portion of the drug taking part in mo-
lecular diffusion.
The isotherm of Fr sorption as the dependence of free diffusing concentration of the
drug on the total loaded drug concentration is shown in Fig. 3. From this figure it follows
that up to 3 wt. % both concentrations are related by a linear function. The deviation from
linearity is observed at the maximal concentration (5 wt. %) of the loaded Fr. At this point
the drug forms the proper phase in the polymer as yellow crystals, while the effect of phase
formation does not distort the manner of the kinetic curve (See Fig.2, the curve for 5 wt. %).
Summarizing the results presented in Figs. 1 and 2, we can estimate effective diffusion
coefficients for all initial concentrations of the drug. Figure 4 shows the concentration de-
pendence of diffusivities which has a maximum defined clearly in the drug concentration
area 1.0–1.5 wt. %. The rising branch of the curve D(C) results likely from disordering of
the PHB structure after drug loading. In contrast, the dropping branch is related to drug crys-
tal formation in the PHB matrix that occurs during the decrease of low-molecular-weight
component mobility. The formation of Fr crystals in PHB has been observed recently in our
work by Krivandin with WAXS technique [21].
Above we have mentioned that, simultaneously with the diffusion kinetics, the linear
kinetic process of Fr release is observed. In this case, the greater the initial concentration
of the loaded drug is, the higher the constant rate of drug release is. More informatively this
effect is shown in Fig. 5, where the exponential dependence of the degradation constant (k)
on the drug concentration is observed. Simultaneously with the measurement of the
3.0
Mobile fraction of furacillin, wt. %

2.5
2.0
1.5
1.0
0.5
0.0
0 1 2 3 4 5
Total furacillin concentration, %
Figure 4 Dependence of release destruction constant (k) on loaded drug concentration for PHB
films.
1.6
Drug diffusion coefficient × 1010, cm2/s
1.2
0.8
0.4
0.0
0.0 0.5 1.0 1.5 2.0 2.5 3.0
Aseptic concentration, wt. %
Figure 5 Diffusivity dependence on aseptic concentration in the drug release process.
concentration of the drug desorbing PHB from films, we have determined the loss of weight
for polymer samples. The gravimetric measurements have shown that the polymer sample
loses its weight also in accordance with the linear law. Initial polymer without the drug has
the stable weight throughout the entire time of release. Preliminary experiments show that,
in contrast to enzymatic biodegradation of PHB going on the polymer surface [19], hydro-
lytic destruction involves all accessible volume of PHB that is supported by the increase in
brittleness and the decrease in the strength of PHB films.
3 Conclusion
We proposed a new polymer system for aseptic controlled release that includes films of
PHB and furacillin. The release goes on simultaneously in accordance with the kinetic
(polymer degradation) and diffusion mechanism. The rates of the kinetic mode for release
obey zero degree curves relative to time. The diffusion mode, which determines the profile
of release in the initial range of time (about the first week), was analyzed in greater detail.
The dependences both of diffusion coefficients and kinetic constant (k) of release on the
drug concentrations were demonstrated. These results are requisite for the further develop-
ment of drug release systems with multi-component action when several drugs simulta-
neously have a local action on biological tissues and cells.
4 Acknowledgement
The authors thank the Biomer Company (Krailing FRG) for the donation of PHB. Funding
for this research was provided partly by the Russian Foundation for Basic Research with
grants Nos 06-04-49339 and 08-03-00929-a.
References
1. R.W. Lenz and R.H. Marchessault, Biomacromolecules, 6 (1), 1–8 (2005).
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4. Y. Liu and M. Wang, Current Appl. Phys., 7 (5), 547–554 (2007).
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ASAIO J., 46 (1), 107–110 (2000).
6. Biomaterials Science: An Introduction to Materials in Medicine, ed. by B.D. Ratner, A.S.
Hoffman, F.J. Schoen and J.E. Lemons, San Diego: Academic Press (1996).
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8. V.I. Sevastianov, N.V. Perova, E.I. Shishatskaya, G.S. Kalacheva and T.G. Volova, J. Biomater.
Sci. Polymer Ed., 14, 1029–042 (2003).
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10. R. Sodian, M. Loebe, A. Hein, D.P. Martin, S.P. Hoerstrup, E.V. Potapov et al., ASAIO J., 48,
12–16 (2002).
11. R. Sodian, S.P. Hoerstrup, J.S. Sperling et al., Ann. Thorac. Surg., 70, 140–144 (2000).
12. M. Unverdorben, A. Spielberger, A. Schywalsky et al., Cardiovasc. Intervent. Radiol., 25,
127–132 (2002).
13. G.T. Köse, S. Ber, F. Korkusuz et al., Biomaterials, 24, 4998–5007 (2003).
14. L.J. Chen and M. Wang, Biomaterials, 23, 2631–2639 (2002).
15. G.-Q. Chen and Q.Wu, Biomaterials, 26 (33), 6565–6578 (2005).
16. D.P. Martin, F. Skraly and S.F. Williams, US Patent 403242 (2003).
17. C.W. Pouton and S. Akhtar, Adv. Drug Deliver. Rev., 18, 133–162 (1996).
18. D. Sendil, I. Gürsel, D.L. Wise and V. Hasirci, J. Controlled Release, 59, 207–217 (1999).
19. A.L. Iordanskii and P.P. Kamaev, Polym. Science, 41B (1–2), 39–43 (1999).
20. R.H. Marchessault, K. Okamura and C.J. Su, Macromolecules, 3 (6), 735–740 (1970).
21. O.V. Shatalova, A.V. Krivandin, and A.L. Iordanskii, 6th Eur. Symposium on Polymer Blends.
Program and Abstracts. May 16–19, Max-Planck-Institut fur Polymerforschung (1999).
Авторам:
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11 Transport of Water
as a Structurally Sensitive
Process Characterizing the
Morphology of
Biodegradable Polymer
Systems
A.L. Iordanskii1, Yu.N. Pankova1, R.Yu. Kosenko1,

A.A. Ol’khov1 and G.E. Zaikov2
1N.N.
Semenov Institute of Chemical Physics, Russian
Russia
email: iordan@chph.ras.ru
2N.M. Emanuel Institute of Biochemical Physics, Russian
Russia
email: chembio@sky.chph.ras.ru
Investigations of water transport in polymer mixtures (where, as a consequence of

a partial compatibility of components, the influence of structural factors on the
mechanical, barrier and other operational properties is especially pronounced) take
on special significance. In this work, we try to show how the morphological charac-
teristics of polymer mixtures influence the water diffusion parameters and how the
latter correlate with the operational characteristics of mixtures by the example of
mixed compositions of a natural polymer product of bacterial vital functions,
poly(3-oxybutyrate) [PHB], and synthetic polymers of various polarities (hydro-
philicities).
Such investigations seem necessary for a detailed description of ecologically
compatible (bio)degradable systems functioning in water media.
350 A.L. Iordanskii, Yu.N. Pankova, R.Yu. Kosenko, A.A. Ol’khov and G.E. Zaikov
Keywords: diffusion, structure of polymers, morphology, biodegradation, bacteria, natural

and synthetic polymers
Introduction
Professor Nikolai M. Emanuel very attentively and, one could say, tremulously treated
young scientists. Direct contacts with Professor Emanuel were infrequent but became turn-
ing and, sometimes, key points of the scientific life of an inexperienced researcher. In our
case, his conceptions about diffusion processes playing not an accessory but a dominating
role in multi-phase biosystems served the basis of a scientific direction “Transport Phenom-
ena in Structurally Organized Polymer Systems”.
Why is it, in particular, diffusion? Why is it the diffusion namely of water? The human
habitat including its biological structures contains a considerable amount of water. Varia-
tion of its concentration and its functionalization determine all processes of vital functions
[1]. Concerning polymer systems, especially if they are used as biomedical systems (vas-
cular prostheses, matrixes for the controlled release of medicines, osteoprosthesis, new-
generation vascular stents, etc.) the necessity arises to obtain information about the state
water is in and the rate of its motion in intercrystal regions of polymer products [2].
From the fundamental point of view the diffusion of bipolar water molecules may be
considered as the motion of a specific probe extremely sensitive to polar functional groups
of macromolecules forming the polymer matrix. Moreover, water has a noticeable affinity
to various hydrophilic impurities occurring in the polymer at some concentration (remnants
of catalysts, plasticizers, monomers and other accompanying components of synthesized
materials or the residual content of albumens, lipids and slats in polymers of natural origin
[3]). Due to the small size of water molecules, such a probe overpasses nonpolar polymer
regions quickly enough. But its diffusion rate is significantly decreased in polar regions (for
example, near albumens’ and polyamides’ amine groups [4]) due to participation in the for-
mation and redistribution of hydrogen bonds in such hydrophilic polymers as cellulose,
PVA, etc. [5].
Investigations of water transport in polymer mixtures (where, as a consequence of par-
tial components’ compatibility, the influence of structural factors on the mechanical, barrier
and other operational properties is especially pronounced) take on a special significance. In
this work we try to show how the morphological characteristics of polymer mixtures influ-
ence the water diffusion parameters and how the latter correlate with the operational char-
acteristics of mixtures on the example of mixed compositions of:
• a natural polymer product of bacterial vital functions – poly(3-oxybutyrate) [P3HB],

• synthetic polymers of various polarities (hydrophilicities).
Such investigations seem necessary for a detailed description of ecologically compat-

ible (bio)destructible systems functioning in water media.
1 Experimental
Materials. A superfine powder of poly-3-oxybutyrate (PHB) was kindly provided by
Biomer (Krailing, Germany). For preparation of mixed films from the solvent the powder
was preliminary dissolved in chloroform and the polymer solution was filtered through a
glass Schott 160 filter. The solubility of initial polymer powder in chloroform turned to be
Transport of Water as a Structurally Sensitive Process 351
close to 0.01 k/ml. The thickness of films was 100±5 µm. X-ray crystallinity of polymer
was 70% and according to the data of the DSC method it was 75%.
After filtration, the molecular mass of polymer films was determined by viscosimetry
using the following equation [6]:
[η ] = 7.7·10 –5 M ω
0.82
, (1)
where [η ] is the intrinsic viscosity of the polymer solution in chloroform. The viscosity-
average molecular mass of polymer, MW, turned to be equal to 150·103 g/mol. Granular
low density polyethylene (LDPE) (State Standard 583-020) was characterized by a molec-
ular mass of 250·103 and density 0.92 g/cm3. In this work, we also used polyvinyl alcohol
(PVA) of Russian brand 8/27 with the residual content of non-saponated acetate groups
8.2% and the concentration of sodium acetate 0.04 wt. %. The molecular mass of PVA is
equal to 64,000 g/mol and the melting temperature is 146°C.
Mixing in extruder. Mixture of LDPE with PHB: firstly, the preliminary mixing in a
ball mill was carried out. The ratios of the components (PHB/LDPE) were: 2/98, 4/96, 8/92,
16/84, and 32/68 wt. %. Blending was carried out in a single-auger extruder (ARP-20) with
the frequency of auger rotation 100 min –1 provided that the temperature of the melt was
not higher than 182°C. The plane-parallel slot was regulated in such a way so as to obtain
films 50–60 µm thick. A similar procedure was carried out when obtaining mixtures of
PHB/polyvinyl alcohol films (PVA, Russian brand 8/27) of structures 10/90, 20/80, 30/70,
and 50/50 wt. %. A similar procedure of extrusion was carried out for initial individual
polymers: PVA, LDPE and PHB.
Methods of analysis. The kinetic curves of water sorption / desorption and equilibri-
um sorption were measured by vacuum gravimetry using a quartz McBen’s microbalance
with the quartz spiral sensitivity of 0.67 mg/mm. The permeability of water vapors was reg-
istered in a traditional two-chamber cell described in [7]. The results of X-ray diffraction
at large angles were obtained with the help of a diffractometer with a linear-coordinate de-
tector constructed at the Joint Institute of Nuclear Research (Dubna, Russia). Since extru-
sion films are characterized by the orientation of both macromolecules and structural
elements in particular crystallites, such orientation was estimated by measuring their polar-
ization spectra. In investigations, the plane of the catalyst was oriented either along or trans-
versely to the direction of extrusion.
Spectral characteristics and round dichroism were measured on an IFS-48 Bricker IR
spectrometer.
2 Hydrophobization of poly-(3-hydroxybutyrate)
Differences in water diffusion behavior when comparing synthetic (gas and oil derivatives)
and natural (products of animals’ and plants’ vital functions) polymers are more clearly re-
vealed on the molecular and crystal level. For hydrophobic polymers (low density PE,
polypropylene, polysiloxane, synthetic rubbers, etc.), analysis of the chemical structure of
macromolecules shows the damage of the primary chemical structure and accumulation of
oxygen-containing functional groups in the polymer chain as a result of imperfect condi-
tions of polymer production. The appearance of groups “introduced” into the hydrophobic
matrix leads to an increase of equilibrium sorption of water and to a decrease of its effective
diffusion coefficient. The diffusion mobility is reduced as a result of water molecules’
a
8
1/Pw 109, cm2 h mmHg/g cm

6
0
0 5 10 15 30 35
PHB, %
2.6
b
2.4
2.2
1/R (1228 cm_1)
2.0
1.8
1.6
1.4
0 5 10 15 20 25 30 35
PHB, %
Figure 1 Comparison of inverse permeability (a) and dichroism (b) values as a function of the struc-
ture of the polymer mixture for films based on LDPE–PHB.
interaction with the damaged polar groups of the polymer. For hydrophobic polymers of
natural origin, the structure of which is formed on the cell and enzyme levels, the damage
of the primary structure is less typical and may occur only at later technological stages, e.g.,
in processing or mixing in extruders.
Under normal conditions, the solubility of water in saturated hydrocarbons is limitedly
low (several mils of percent) [8]. In hydrocarbons C7 –C16 the mole fraction of water (Xw)
is written as the following equation:
ln(Xw –1) = Vw (∆δ)2/RT, (2)
where Vw is the water molar volume, ∆δ is the difference between the solubilities of water
and hydrocarbon, R and T are the gas constant and temperature, respectively. The authors
of [8] by extrapolation of eq. (1) showed that at 25°C the values of Xw did not exceed 10–6
mol/l (~2·10–5 wt. %), so this value might be considered as an ideal solubility of water in
the hydrocarbon matrix.
In both synthetic (polyolefines) and natural hydrocarbon polymers the limit of the sol-
ubility of water is significantly higher and is significantly changed depending on the poly-
mer production method. Systematic investigations of water diffusion in films of low and
high density PE showed that the emergence of damaged fragments on the polymer chain
influenced water molecules’ sorption and diffusion [9]. Immobilization of water molecules
on such polar groups statistically distributed in the hydrocarbon nonpolar matrix leads to a
noticeable decrease of the effective diffusion coefficient of water.
Dw –1 = Dwf–1(1 + [Spf KL/KH]), (3)
where Dwf is the diffusion coefficient of a single water molecule (free diffusion), Spf is the
concentration of introduced (damaged) groups, KH and KL are the Henry and Langmuir con-
stants of sorption, respectively.
With the aim of obtaining biodegradable polymer materials able to destruct in soil but
to keep their transport and mechanical properties during long exploitation period we inves-
tigated new polymer compositional mixtures of poly-3-oxybutyrate and LDPE [10]. For
such mixtures, water permeability depends on PHB/PE, the ratio of components’ concen-
trations, see Fig. 1a where the value of inverse permeability is presented, i.e. water diffusion
resistance as a function of PHB concentration. Introduction of carbonyl groups of POB into
the nonpolar matrix of polyethylene (LDPE) enables regulation of transport parameters.
The dependence “water resistance vs. POB concentration” described by the extreme and
non-additive function is in good agreement with the segment orientation acquired during
the extrusion process during the melting of PE/POB mixtures. The degree of orientation is
determined by the method of Fourier spectroscopy under construction of the dichroism
function [11] and is presented in Fig. 1b.
The reason for the extremums to appear on the “dichroism–mixture structure” curves
may be the covering of two processes oppositely influencing the orientation of macromol-
ecules in crystal regions (and consequently the orientation of crystals). On the one hand,
with the increase of the content of rigid molecules of PHB in the melt the tendency of or-
dering all molecules while passing through the slot of the extruder is increased. On the other
hand, the viscosity of the mixture in this case is increased and consequently the time nec-
essary for structural elements’ (including crystals’) orientation is also increased. At low
concentrations of PHB (>8 wt. %) the orientation process prevails. And at higher PHB con-
centrations the tendency of disordering of the structural organization begins to be exhibited
more clearly. The decrease of the degree of crystallinity of both components observed in-
dependently by the DSC method confirms this fact.
The results of X-ray analysis for large and small angles showed the presence of crystal
axial a-texture realized in investigated films for both PE and PHB [12]. Crystallites belong-
ing to both polymers are oriented preliminary in such a way that axes a of elementary crystal
cells of both mixture components coincide with the direction of extrusion, and axes c coin-
ciding with macromolecules’ orientation in crystallites’ elementary cells are oriented trans-
versely to this direction. Such a position of crystallites is characteristic of all compositions
studied. That is why a combination of earlier obtained X-ray conceptions about the crystal
structure of the POB/PE mixture and the results of polarization IR Fourier spectroscopy
suggest that the cause of inversion of the dichroism function is probably the difference in
the angles of transition moment for methyl groups belonging to PE (=0°C) and POB
(=90°C), respectively.
3 Hydrophilization of poly-(3-hydroxybutyrate)
The results obtained by the DSC method for mixtures and initial samples of PHB and PVA
are presented in Table 1.
Table 1 Glass-transition temperature of mixture compositions with various contents of PHB.
PHB concentration, % 0 10 20 30 50 100
Tg, ºC 53.9 32.7 28.5 25.3 25.9 24.1
As we mentioned above, all samples were obtained by the extrusion method as de-
scribed in the experimental part. Analysis of the thermograms showed that there was a sta-
ble low-temperature transition in the interval 24–54°C reflecting, most probably, the
transition from the glassy to rubberlike state at the glass-transition temperature of ~37°C.
We should note that for initial polymer components the corresponding glass-transition
temperatures lie in the region of 24.1°C and 53.9°C for PHB and PVA, respectively. The
existence in the mixture compositions of only one phase transition of the second order
should testify to the interaction between two polymers [13] forming the mixture and to their
possible partial compatibility in inter-crystallite regions of mixture films. A combination in
polymer crystallite regions (their part is large and is equal to more than 70%) most likely
does not occur since on DSC thermograms two independent peaks belonging correspond-
ingly to PHB and PVA are observed.
The occurrence of independent crystallite phases of each of mixture-forming polymers
was observed by the method of wide angle X-ray scattering (WAXS). In the interval of PHB
concentration from 0 up to 30% each of components forms its own, slightly oriented crystal
phase that is confirmed by X-ray diffractograms of the mixtures; they represent a super-
position of diffractograms of individual polymers: PHB (degree of crystallinity ~70%) and
PVA (degree of crystallinity ~30%).
At all polymer ratios forming the given mixtures, the parameters of the elementary
orthorhombic crystal cell of PHB remain constant and have the following values, respec-
tively: a = 0.576 nm, b = 1.32 nm, and c = 0.596 nm [14]. X-ray reflexes for PVA show
that crystalline regions of polyvinyl alcohol are in gamma form well described earlier in
domestic literature by L.L. Razumovskaya et al. [15]. In particular, the reflex S = 2.21
nm –1 belongs to this modification of PVA structure.
The form of diffractograms obtained at various angles of X-ray irradiation incidence
in relation to sample plane shows that for all samples containing PVA the axial cylindrical
texture is observed where the axis of texture coincides with direction of extrusion through
the slot. In samples containing 10 and 20% of PHB, the axial texture is observed also for
poly-3-hydroxybutyrate. However, in contrast to PVA where macromolecules are oriented
along the direction of extrusion, in crystal regions of poly-3-hydroxybutyrate the axis a of
elementary crystal cell coincides with the direction of extrusion and, consequently, biopoly-
mer macromolecules are preliminarily arranged transversely to the axis of extrusion [16].
Diffractograms of mixture compositions containing 30 and especially 50% of PHB
show the absence of texture of the latter and so, these sample are preliminarily isotropic.
The transition from textured to isotropic crystal structures observing in the region of 30%
of PHB is probably related to the phase inversion of samples occurring in the mentioned
region of mixture structure. A sharp change of structural and physical characteristics of
polymer mixtures occurs namely in the region of phase inversion, see, e.g., [17].
4
PHB(110)
PHB(020) 1
3 PVA(110)
2
I, rel. units
PHB(111)
2
0
1.0 1.5 2.0 2.5 3.0 3.5
S, 1/nm
6
PHB(020)
PHB(110)
4
I, rel. units
PVA(110)
2 3
2
2
1
1, 3
0
1.0 1.5 2.0 2.5 3.0 3.5
S, 1/nm
Figure 2 WAXS diffraction of extrusion films prepared by mixing PHB and PVA (upper, 80:20%
and lower, 70:30%). The vector of X-ray irradiation is directed along the direction of extrusion (1),
transversely (2) and at an angle of 20º (3) to the direction of extrusion. Results and Discussion were
prepared by O.V. Shatalova and A.V. Krivandin (IBCS RAS).
Taking into account the results presented above, we studied the physicomechanical
characteristics of polymer mixtures with various contents of PHB. It is important to note
that on the curves “rupture stress–structure of mixture” and “elongation at rupture–
structure of mixture” the sharp changes of values of mechanical parameters are observed,
and in the same concentration interval (~ 30% PHB), where the above mentioned transition
occurs. Besides, the minimum of the extreme dependence of elasticity modulus on PHB
concentration in the mixture is situated in the same concentration interval. A decrease of
elasticity modulus in the region of low concentrations of PHB is caused by the reduction of
segment mobility of interaction macromolecules of PHB and PVA. A further increase of
elasticity modulus values reflects the increase of structural units’ content responsible for
material strength and namely of crystallites belonging to poly-3-hydroxybutyrate. The to-
tality of the results of mechanical tests for mixture films presented in Figure 3 confirms the
physical concept of structural inversion, which concerns both crystal and inter-crystal (con-
ditionally amorphous) regions of mixture compositions.
Transformation of amorphous polymer regions as a result of phase inversion should
be reflected on the level of diffusion processes and especially under diffusion and
100
250
Elongation at rupture, %
75 200
Rupture stress, MPa
150
50
100
50
25
0
0 20 40 60
PHB content, wt. %
Figure 3 Dependence of the tensile strain and rupture stress on the structure of a PHB–PVA poly-
mer mixture.
40
g/cm3 . molecular mass of mercury
35
30 T = 20°C
Water solubility,
25
20
15
10
5
0
0 20 40 60 80 100
PHB concentration, wt. %
Figure 4 Change of water solubility as a function of the structure of a mixture.
permeability of structurally-sensitive water molecules. It is well known that even so

small-size molecules as those of water are not capable of penetrating crystal regions of any
polymer and, consequently, information about diffusion flows, diffusion mobility and water
sorption reflects first of all the state of amorphous regions of the polymer matrix.
In mixture compositions, the content of water is determined by the ratio of the com-
ponents if polymers of various hydrophilicities are mixed. In our case, the mixture is formed
by moderately hydrophilic complex polyester (PHB) and typical hydrophilic polyalcohol
(PVA). The increase of the concentration of the latter in the mixture corresponds to a
monotonous growth of hydroxyl groups in the system. The equilibrium concentration of
water is increased analogously, since in accordance with the theory of group contributions
[18] there is a definite amount of water molecules interacting with the OH group per each
Water permeability . 108, g cm/cm2 hour mm

300 Pw 108
Hypothetically additive dependence
250
200
of mercury 150
100
50
0 20 40 60 80 100
Figure 5 Extreme dependence of water permeability through films based on PHB–PVA mixtures.
of these OH groups of polymer. The contribution of the ester group of PHB is significantly
lower and in this particular case we can neglect it. Actually, the equilibrium sorption of wa-
ter in investigated mixtures similarly increases monotonously with the rise of PVA concen-
tration or with the decrease of the PHB content (Figure 4). Characteristic inflections and
extreme points on the water sorption–mixture structure dependence are not observed and
consequently the equilibrium sorption is sensitive mainly to the primary chemical structure
of polymer, to be more exact, to the chemical structure of the polymer mixture. Thus, a mo-
notonous increase of water equilibrium sorption with the change of the mixture structure is
determined by the substitution of ester functional groups belonging to PHB and possessing
a low affinity to water molecules by more hydrophilic groups (hydroxyl) belonging to PVA.
In contrast to equilibrium sorption data the values of normalized water flows (perme-
ability) have a characteristic degraded maximum in the concentration region corresponding
approximately to 40% of PHB as shown in Figure 5. Obviously, in this case as a result of
rebuilding the structure the maximum accumulation of structural defects occurs due to
which the increase of water flow is observed.
In the interval of concentrations preceding this point (10–30% PHB) there is no
additivity of the permeability parameter and even a small addition of polyester (10%)
significantly reduces the values of water permeability first of all at the expense of segment
interaction between PHB and PVA macromolecules forming the mixture. This conclusion
is also confirmed by the shift of mixture glass-transition temperature. We want to note once
again that such interactions are realized in intercrystalline or better to say amorphous re-
gions and do not affect the crystalline regions of polymer. The descending line of the water
permeability–mixture structure dependence is approximately described by a linear function
and consequently obeys a property–structure additive dependence. So, in this region of
structure each of components independently leaks water flow and, thus, segment interaction
revealed in the concentration region below the point of structural transition is practically
absent.
When one analyzes the dependence of water permeability on structure, one should
remember that from the physical point of view permeability represents the product of two
other physical parameters: sorption capacity and diffusion mobility. We considered water
30 100
25
Diffusion coefficients 108, cm2/sec
Rupture stress, MPa

75
20
=σ = Dw
15
50
10
5 25
0
0 10 20 30 40 50
Figure 6 Similarity of the behavior of the diffusion coefficients and rupture stress for PHB–PVA
mixture compositions.
sorption above, and as the parameter characterizing the mobility of water molecules in mix-
ture films we chose the coefficient of water diffusion. In Figure 6 we show these coeffi-
cients in the form of a dependence on mixture structure. An analogous dependence for
rupture stress is presented in the same figure for clarity. Both the diffusion and mechanical
characteristics of mixture compositions are characterized by clear inflection points situated
in the region of structural transition at 30% of PHB that coincides with the results of X-ray
analysis. As in the case of mechanical tests in the region of low content of PHB the diffusion
behavior of the system resembles the diffusion in pure PVA (the region 0–20%) whereas
at concentrations exceeding 30% of PHB the diffusion coefficients of mixture compositions
sharply exceed the diffusion coefficients of water in PHB (Dw = 1–10-8 cm2/sec [19]).
In accordance with the results of water permeability measurements we assume that
high values of the diffusion coefficients are caused by structure defects more clearly re-
vealed as a result and after phase transition. Earlier, by WAXS, EPR, Fourier IR spectros-
copy and vacuum gravimetry methods we showed [20, 21] that crystalline transition from
anisotropic into isotropic state of PHB led to the change of water diffusion mobility. In the
case of the change of mixture structure we also observe the transition from the textured state
into a more disordered one, i.e. isotropic. Besides, an increase of defects of mixture com-
positions at a relatively high content of PHB does not make it possible to exclude the pos-
sibility of partial water leakage through the defect zones in the matrix analogously to the
case observed for PHB–LDPE mixtures [22].
Intensive studying of particularities of water transfer in mixture compositions on the
basis of poly-3-hydroxybutyrate represents a necessary stage in the investigation of such
fundamental processes as physical ageing at high dampness, corrosion stability of polymer
membranes; these results should also promote the development of novel ecologically and
biologically compatible materials for medicine and agriculture.
A complex of methods combining structural (DSC, IR Fourier polarization spectros-
copy), mechanical and diffusion experiments was used in this work. By varying the nature
of the second mixing component (its hydrophilicity) we may control the morphology of the
mixture and, consequently, the rate of diffusion processes. For example, mixing of PHB
with a representative of the class of hydrophobic polymers (LDPE) does not give compat-
ible compositions. Matrices of PHB–LDPE mixtures are partially crystalline, oriented in
relation to extrusion direction. The direct correlation between the diffusion resistance of
film to water flow and the degree of orientation of macromolecules is shown. On the con-
trary, hydrophilization of PHB by mixing with PVA gives the possibility of preparing novel
mixture films partially compatible in intercrystalline regions. The region of structural tran-
sition (at 30% of PHB) where mechanical and transport characteristics are sharply changed
was found.
References
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Concepts in Polymer Science, VSP Science Press: Utrecht–Tokyo (1988).
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(1999).
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676 (2000).
13. D.R. Paul and C.B. Bucknal, Polymer Blends, Wiley: New York, 1 (2000).
14. A.L. Iordanskii, A.A. Ol’khov, O.V. Shatalova, G.E. Zaikov, and U.J. Hanggi, Water Diffusion,
Crystalline Structure, and Mechanical Properties of Novel PHB–PVA Blends, ed by
A.L. Iordanskii, O.V. Startsev and G.E. Zaikov, Nova Science: New York, Ch. 9, 213 (2004).
15. L.L. Razumova, O.V. Shatalova, A.L. Iordanskii and G.E. Zaikov, Vysokomol. Soed., 38A, 1271
16. A.A. Ol’khov, S.V. Vlasov, A.L. Iordanskii, G.E. Zaikov and V.M. Lobo, J. Appl. Polymer, 90,
1471 (2003).
17. J.P. Runt, Crystalline Polymer Blends, in: Polymer Blends, ed. by D.R. Paul and C.B. Bucknall,
Wiley: New York, 1, 167 (2000).
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19. P.P. Kamaev, I.I. Aliev, A.L. Iordanskii and A.M. Wasserman, Polymer (UK), 42, 515 (2001).
20. P.P. Kamaev, A.L. Iordanskii, I.I. Aliev, A.M. Wasserman and U.J. Hänggi, Desalination, 126,
153 (1999).
21. A.L. Iordanskii and P.P. Kamaev, Polymer. Sci. Ser. B, 41 (1–2), 39 (1999).
22. A.L. Iordanskii, From Traditional to Novel Environmentally Friendly Polymers, in: Water
Transport in Synthetic Polymers, ed. by A.L. Iordanskii, O.V. Startsev, and G.E. Zaikov, Nova
Science: New York, Ch. 1, 1 (2004).
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12 A Novel Technique for

Measurement of
Electrospun Nanofiber
M. Ziabari, V. Mottaghitalab and A.K. Haghi*

University of Guilan, P.O. Box 3756, Rasht, Iran
*Haghi@Guilan.ac.ir
In electrospinning, fiber diameter is an important structural characteristic because

it directly affects the properties of the produced webs. In this chapter, we have
developed an image analysis based method called direct tracking for measuring
electrospun fiber diameter. In order to evaluate its accuracy, samples with known
characteristics have been generated using a simulation scheme known as
µ-randomness. To verify the applicability of the method, some real webs obtained
from electrospinning of PVA have been used. Due to the need of binary input
images, micrographs of the real webs obtained from scanning electron microscopy
were segmented using local thresholding. The method was compared with the
distance transform method. Results obtained by direct tracking were significantly
better than distance transform, indicating that the method could be used
successfully for measuring electrospun fiber diameter.
Keywords: electrospinning, fiber diameter, image analysis, direct tracking, distance trans-
form
Introduction
Conventional fiber spinning (like melt, dry and wet spinning) produce fibers with diameters
in the range of a micrometer. In recent years, electrospinning has gained much attention as
a useful method to prepare fibers in nanometer diameter range. These ultrafine fibers are
classified as nanofibers. The unique combination of high specific surface area, extremely
small pore size, flexibility and superior directional strength makes nanofibers a preferred
material form for many applications. Proposed uses of nanofibers include wound dressing,
drug delivery, tissue scaffolds, protective clothing, filtration, reinforcement and micro-
electronics.
362 M. Ziabari, V. Mottaghitalab and A.K. Haghi
Figure 1 Electrospinning setup.
α
d
O
Figure 2 Procedure for µ-randomness.
In the electrospinning process, a polymer solution held by its surface tension at the end
of a capillary tube is subjected to an electric field. Charge is induced on the liquid surface
by an electric field. Mutual charge repulsion causes a force directly opposite to the surface
tension. As the intensity of the electric field is increased, the hemispherical surface of the
solution at the tip of the capillary tube elongates to form a conical shape known as the Tay-
lor cone. When the electric field reaches a critical value at which the repulsive electric force
overcomes the surface tension force, a charged jet of the solution is ejected from the tip of
the cone. Since this jet is charged, its trajectory can be controlled by an electric field. As
the jet travels in the air, the solvent evaporates, leaving behind a charged polymer fiber
which lays itself randomly on a collecting metal screen. Thus, continuous fibers are laid to
form a nonwoven fabric. Figure 1 illustrates the electrospinning setup [1–6].
The properties of electrospun nonwoven webs depend on the nature of the component
fiber as well as its structural characteristics such as fiber orientation [7–12], fiber diameter
[13], pore size [15], uniformity and other structural features [16]. Analyzing the electrospun
webs yield results and information, which will help researchers in improving the quality and
predicting the overall performance of electrospun webs. Some of the reasons for character-
ization may be process control, process development and product or quality control. Fiber
diameter is the most important structural characteristics in electrospun nonwoven webs. De-
pending on the process and material variables, the diameter of fibers produced by electro-
spinning varies. Almost all researches who have done characterization, have reported the
effects of processing variables on electrospun fiber diameter. There is no standard technique
to measure the fiber diameter and analyze its distribution. This explains the need to study
the fiber diameter of electrospun webs. Recently, image analysis has been used to identify
fibers and measure structural characteristics in nonwovens. However, the accuracy and lim-
itations of these techniques have not been verified.
The objective of our research is to use image analysis for measuring electrospun fiber
diameter.
1 Methodology
1.1 Simulation of electrospun web

In order to reliably evaluate the accuracy of the developed methods, samples with known
characteristics are required. Since this end cannot be met with experiment, a simulation al-
gorithm has been employed for generating nonwovens with known characteristics. The use
of simulation is not a new idea. It was used by Abdel-Ghani and Davis [17] and Pourdey-
himi et al. [7] for simulation of nonwovens with both continuous and discontinuous fibers
by the use of idealized straight lines. The most important component of simulation is the
way in which lines or curves are generated. Abdel-Ghani and Davis [17] presented three
methods for generating a random network of lines:
1) Surface randomness known as S-randomness

2) Mean free path known as µ-randomness
3) Internal randomness known as I-randomness.
It is assumed that the lines are infinitely long (continuous filament) so that, at least in
the image plane, all lines intersect the boundaries.
The aim is to obtain unbiased arrays spatially homogeneous for infinitely long lines.
Lately it was discovered by Pourdeyhimi et al. [7] that the best way to simulate nonwovens
of continuous fibers is through the second method. Under this scheme, a line with a speci-
fied thickness is defined by the perpendicular distance d from a fixed reference point O
located in the center of the image and the angular position of the perpendicular α. Distance
d is limited to the diagonal of the image [7, 17]. Figure 2 demonstrates this procedure.
Several variables are allowed to be controlled during the simulation:
1) Web density: can be controlled using the line density which is the number of lines to
be generated in the image.
2) Angular density: useful for generating fibrous structures with specific orientation
distribution. The orientation may be sampled from either a normal or a uniform
random distribution.
3) Distance from the reference point: varies between 0 and the diagonal of the image,
restricted by the boundary of the image and is sampled from a uniform random
distribution.
4) Line thickness (fiber diameter): is sampled from a normal distribution. The mean
diameter and its standard deviation are needed.
5) Image size: can also be chosen as required.
1.2 Fiber diameter measurement

The electrospinning process produces very fine fibers, and this is one of the few methods
from which fibers of submicron size can be produced. So it becomes immensely important
Figure 3 Manual method.
to understand the behavior of fiber diameter and fiber diameter distribution in the electro-
spun web as impacted by the processing parameters. The first step in determining fiber di-
ameter is to produce a high quality image of the web at a suitable magnification using
electron microscopy techniques, called micrograph. The methods for measuring electro-
spun fiber diameter are described in the following sections.
1.2.1 Manual method

The conventional method of measuring the fiber diameter of electrospun webs is to analyze
the micrograph manually. The manual analysis usually consists of the following steps, de-
termining the length of a pixel of the image (setting the scale), identifying the edges of the
fibers in the image and counting the number of pixels between two edges of the fiber (the
measurements are made perpendicular to the direction of fiber axis), converting the number
of pixels to nm using the scale and recording the result. Typically 100 measurements are
carried out. Figure 3 illustrates this process.
However, this process is tedious and time-consuming especially for a large number of
samples. Furthermore, it cannot be used as an on-line method for quality control since an
operator is needed to perform the measurements. Thus, developing automated techniques
which eliminate the need for an operator and can be employed as on-line quality control is
of great importance.
1.2.2 Distance transform

The distance transform of a binary image is the distance from every pixel to the nearest non-
zero-valued pixel. The center of an object in the distance transform image will have the
highest value and lie exactly over the object’s skeleton. The skeleton of the object can be
obtained by the process of skeletonization or thinning. The algorithm removes pixels on the
boundaries of objects but does not allow objects to break apart. This reduces a thick object
to its corresponding object with one pixel width. Skeletonization or thinning often produces
short spurs which can be cleaned up automatically with a pruning procedure [18].
The algorithm for determining fiber diameter uses a binary input image and creates its
skeleton and distance transformed image. The skeleton acts as a guide for tracking the
a) b)
c)
Figure 4 a) A simple simulated image, b) its skeleton overlaid on its distance transform, c) a histo-
gram of fiber diameter distribution obtained by distance transform.
distance transformed image by recording the intensities to compute the diameter at all
points along the skeleton. This method was proposed by Pourdeyhimi et al. [13]. Figure 4
shows a simple simulated image, its skeleton overlaid on its distance transform and the his-
togram of fiber diameter obtained by this method.
1.2.3 Direct tracking

In order to measure the electrospun fiber diameter, we developed an image analysis-based
method called direct tracking. This method in which a binary image is used as an input de-
termines the fiber diameter on the basis of two scans; first a horizontal scan and then a ver-
tical scan. In the first scan, the algorithm searches for the first white pixel adjacent to a
black. Pixels are counted until reaching the first black. The second scan is then started from
the mid point of a horizontal scan and pixels are counted until the first black is encountered.
If the black pixel is not found, the direction changes. Having the number of horizontal and
vertical scans, the number of pixels in perpendicular direction which is the fiber diameter
could be measured from a geometrical relationship. The process is illustrated in Fig. 5.
In electrospun webs, fibers cross each other at intersection points and this brings about
some untrue measurements in these areas. To circumvent this problem, black regions are
labeled and it is identified which couple of regions consists of fiber. Then, in order to en-
hance the processing speed, the image is cropped to the size of selected regions. Afterwards,
fiber diameter is measured according to the explained algorithm. Finally, the data in pixels
are converted to nm and the histogram of fiber diameter distribution is plotted. Figure 6
shows a labeled simple simulated image and the histogram of fiber diameter obtained by
this method.
d
y
a) b)
Figure 5 a) Direct tracking, b) fiber diameter from the number of vertical and horizontal pixels.
a) b)
Figure 6 a) A simple simulated image which is labeled, b) histogram of fiber diameter distribution
obtained by direct tracking.
1.3 Real webs treatment

Both the distance transform and direct tracking algorithm for measuring fiber diameter re-
quire binary image as input. Hence, micrographs first have to be converted to black and
white. This is done by thresholding (known also as segmentation), which produces a binary
image from a grayscale (intensity) image. This is a critical step because segmentation af-
fects the result. In the simplest thresholding technique, called global thresholding, the image
is partitioned using a single constant threshold. One simple way to choose a threshold is by
trial and error. Then each pixel is labeled as an object or background depending on whether
the gray level of that pixel is greater or less than the value of the threshold, respectively.
The main problem here is that global thresholding can fail in the presence of nonuni-
form illumination or local gray level unevenness. An alternative to circumvent this problem
is to use local thresholding instead. In this approach, the original image is divided to sub-
images, and different thresholds are used for segmentation. Another variation of this ap-
proach which has been used in this study consists of estimating the background illumination
using morphological opening operation, subtracting the obtained background from the orig-
inal image and applying a global thresholding to produce the binary version of the image.
In order to automatically compute the appropriate threshold, Ostu’s method could be em-
ployed. This method chooses the threshold to minimize interaclass variance of black and
white pixels. Prior to segmentation, an intensity adjustment operation and a two dimension-
al median filter were applied in order to enhance the contrast of the image and remove noise
a) b)
c)
Figure 7 a) A real web, b) global thresholding, c) local thresholding.
[18–21]. As it is shown in Fig. 7, global thresholding resulted in some broken fiber seg-
ments. This problem was solved using local thresholding.
2 Experimental
Electrospun nonwoven webs used as real webs in image analysis obtained from electrospin-
ning of PVA with an average molecular weight of 72,000 g/mol, purchased from Merck, at
different processing parameters. The micrographs of the webs were obtained using a Philips
XL-30 environmental scanning electron microscope under magnification of 10,000× after
being gold coated. Figure 8 shows micrographs of electrospun webs used as real webs.

Two sets each composed of five simulated images generated by µ-randomness procedure
were used as samples with known characteristics to demonstrate the validity of the tech-
niques. The first set had random orientation with increasing constant diameters; the second
was also randomly oriented but with a varying diameter sampled from normal distributions
with a mean of 15 pixels and standard deviations ranging from 2 to 10 pixels.
Tables 1 and 2 show the structural features of these simulated images, which are shown
in Figs. 9 and 10. Moreover, the applicability of the techniques was tested using five real
webs obtained from electrospinning of PVA.
Mean and standard deviation of the simulated images in the first and second set are
shown in Tables 3 and 4, respectively. Table 5 shows the results for real webs in terms of
pixel and nm. Figures 11 and 12 show histograms of fiber diameter distribution for simu-
lated images in the first and second set, respectively. Histograms for real webs are given in
Fig. 13.
In the first set, for simulated images with the line thickness of 5 and 10 pixels, the
distance transform presents mean and standard deviation of fiber diameter closer to the
R1 R2
R3 R4
R5
Figure 8 Micrographs of the electrospun webs.
simulation. For the line thickness of 15, the standard deviation of the diameter obtained
from the direct tracking method is closer to that of the simulation. However, in this case the
distance transform measured the average diameter more accurately. For the simulated webs
with a line thickness of more than 15 in the first set, the direct tracking method resulted in
a better estimation of the mean and standard deviation of fiber diameter. This is due to the
fact that as the lines get thicker, there is a higher possibility of branching during the skele-
tonization (or thinning), and these branches remain even after pruning. Although these
branches are small, their orientation is typically normal to the fiber axis, thus causing a wid-
ening of the distribution.
Table 1 Structural characteristics of first set images.
Image No Angular range Line density Line thickness
C1 0–360 30 5
C2 0–360 30 10
C3 0–360 30 15
C4 0–360 30 20
C5 0–360 30 25
C1 C2
C3 C4
C5
Figure 9 Simulated images with constant diameter.
Table 2 Structural characteristics of second set images.
Image No Angular range Line density Line thickness
Mean Standard
V1 0–360 30 15 2
V2 0–360 30 15 4
V3 0–360 30 15 6
V4 0–360 30 15 8
V5 0–360 30 15 10
Furthermore, the distance transform method fails in measuring the diameter of fiber in
intersections. Intersections cause to overestimate fiber diameter. Since in the direct tracking
method an image is divided into parts where single fibers exist, the effect of intersections
which causes an inaccurate measurement of fiber diameter is eliminated. Therefore, there
will be a better estimate of fiber diameter.
V1 V2
V3 V4
V5
Figure 10 Simulated images with varying diameter.
In the second set, irrespective of the line thickness in the simulation, for all simulated
webs direct tracking resulted in a better measurement of mean and standard deviation of
fiber diameter. For real webs, mean and standard deviation of fiber diameter for direct track-
ing were closer to those of the manual method, which concurs with the trends observed for
the simulated images.
Table 3 Mean and standard deviation for series 1.
C1 C2 C3 C4 C5
Simulation mean 5 10 15 20 25
standard 0 0 0 0 0
Distance transform mean 5.486 10.450 16.573 23.016 30.063
standard 1.089 2.300 5.137 6.913 10.205
Direct tracking mean 5.625 11.313 17.589 22.864 29.469
standard 1.113 2.370 4.492 5.655 7.241
C1 C2
C3 C4
C5
Figure 11 Histograms for simulated images with constant diameter.
Table 4 Mean and standard deviation for series 2.
V1 V2 V3 V4 V5
Simulation mean 15.247 15.350 15.243 15.367 16.628

standard 1.998 4.466 5.766 8.129 9.799
Distance transform mean 16.517 16.593 17.135 17.865 19.394
standard 5.350 6.165 7.597 9.553 11.961
Direct tracking mean 16.075 15.803 16.252 16.770 18.756
standard 2.606 5.007 6.129 9.319 10.251
V1 V2
V3 V4
V3 V4
V5
Figure 12 Histograms for simulated images with varying diameter.
Table 5 Mean and standard deviation for real webs.
R1 R2 R3 R4 R5
Manual mean pixel 24.358 24.633 18.583 18.827 17.437

nm 318.67 322.27 243.11 246.31 228.12
standard pixel 3.193 3.179 2.163 1.984 2.230
nm 41.77 41.59 28.30 25.96 29.18
Distance transform mean pixel 27.250 27.870 20.028 23.079 20.345

nm 356.49 364.61 262.01 301.94 266.17
standard pixel 8.125 7.462 4.906 7.005 6.207
nm 106.30 97.62 64.18 91.64 81.21
Direct tracking mean pixel 27.195 27.606 20.638 21.913 20.145

nm 355.78 361.15 269.99 286.68 263.55
standard pixel 4.123 5.409 4.148 4.214 3.800
nm 53.94 70.77 54.27 55.14 49.72
R1 R2
R3 R4
R5
Figure 13 Histograms for real webs.
4 Conclusion
Fiber diameter is the most important structural characteristics in electrospun nonwoven
webs. The typical way of measuring electrospun fiber diameter is through the manual meth-
od which is a tedious, time-consuming and an operator-based method and cannot be used
as an automated technique for quality control. We investigated the use of image analysis
for determining the fiber diameter and developed an automated method called direct track-
ing. Since this is a new technique, its accuracy needs to be evaluated using samples with
known characteristics. To that end, the µ-randomness procedure was used in order to sim-
ulate electrospun nonwoven webs. Based on this scheme, two sets of simulated images,
each containing 5 webs, were generated. The first set had random orientation with increas-
ing constant diameter. For the second set, the diameter values were sampled from normal
distributions with a mean of 15 and standard deviation ranging from 2 to 10 pixels. We
compared our method with the distance transform method. For all the simulated webs with
varying diameter and for those with constant diameter more than 15, the direct tracking
method resulted in the mean and standard deviation closer to the simulation. However, for
the simulated webs with smaller constant diameter, the distance transform measured the
mean and standard deviation of fiber diameter more accurately. The results suggest that the
direct tracking method is an accurate, direct measurement technique, because it extracts the
fiber diameter for the samples by tracking fixed segment of the fiber and eliminates the ef-
fect of intersections.
We have demonstrated the general applicability of the method using real webs. Five
real electrospun nonwoven webs obtained by electrospinning of PVA were used. Since the
methods needed binary images as input, the images first had to be segmented. A local
thresholding method together with Ostu’s method was employed in order to automatically
compute the appropriate threshold. The results obtained for real webs confirm the trends
suggested by simulated images. The results show that the use of image analysis in order to
determine the fiber diameter in electrospun nonwoven webs has been successful.
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13 Image Analysis of Pore Size

Distribution in Electrospun
Nanofiber Webs: New
Trends and Developments
M. Ziabari, V. Mottaghitalab and A.K. Haghi*

University of Guilan, P. O. Box 3756, Rasht, Iran
*Haghi@Guilan.ac.ir
Nanofibers produced by the electrospinning method are widely used for drug
delivery, as tissue scaffolding materials and filtration purposes where specific pore
characteristics are required. For continued growth in these areas, it is critical that
nanofibers be properly designed for these applications to prevent failure. Most of
the current methods only provide an indirect way of determining the pore
structure parameters and contain inherent disadvantages. In this study, we
developed a novel image analysis method for measuring the pore characteristics of
electrospun nanofiber webs. Five electrospun webs with different pore
characteristics were analyzed by this method. The method is direct and rapid and
presents valuable and comprehensive information regarding the pore structure
parameters of webs. Two sets of simulated images were generated to study the
effects of web density, fiber diameter and its variations on pore characteristics.
The results indicated that web density and fiber diameter significantly influence
the pore characteristics whereas the effect of fiber diameter variations was
insignificant.
Keywords: image analysis, pore size, nanofibers, electrospinning
Introduction
Fibers with a diameter of around 100 nm are generally classified as nanofibers. What makes
nanofibers of great interest is their extremely small size. Nanofibers compared to conven-
tional fibers, with higher surface area to volume ratios and smaller pore size, offer an op-
portunity for use in a wide variety of applications. To date, the most successful method of
Figure 1 Electrospinning setup.
producing nanofibers is through the process of electrospinning. The electrospinning process

uses high voltage to create an electric field between a droplet of polymer solution at the tip
of a needle and a collector plate. When the electrostatic force overcomes the surface tension
of the drop, a charged, continuous jet of polymer solution is ejected. As the solution moves
away from the needle and toward the collector, the solvent evaporates and the jet rapidly
thins and dries. On the surface of the collector, a nonwoven web of randomly oriented solid
nanofibers is deposited [1 – 5]. Figure 1 illustrates the electrospinning setup.
Material properties such as melting temperature and glass transition temperature as
well as structural characteristics of nanofiber webs such as fiber diameter distribution, pore
size distribution and fiber orientation distribution determine the physical and mechanical
properties of the webs. The surface of electrospun fibers is important when considering
end-use applications. For example, the ability to introduce porous surface features of a
known size is required if nanoparticles need to be deposited on the surface of the fiber, if
drug molecules are to be incorporated for controlled release, as tissue scaffolding materials
and for acting as a cradle for enzymes [6]. Besides, filtration performance of nanofibers is
strongly related to their pore structure parameters, i.e., percent open area (POA) and
pore-opening size distribution (PSD). Hence, the control of the pore of electrospun webs is
of prime importance for nanofibers produced for these purposes. There is no literature avail-
able about the pore size and its distribution of electrospun fibers and in this work, the pore
size and its distribution was measured using an image analysis technique.
Current methods for determining PSD are mostly indirect and contain inherent disad-
vantages. Recent technological advancements in image analysis offer great potential for a
more accurate and direct way of determining the PSD of electrospun webs. Overall, the im-
age analysis method provides a unique and accurate method that can measure pore opening
sizes in electrospun nanofiber webs.
1 Methodology
The porosity, εV, is defined as the percentage of the volume of the voids, Vv, to the total
volume (voids plus constituent material), Vt, and is given by
Vv
εV = ×100 .
Vt
Image Analysis of Pore Size Distribution in Electrospun Nanofiber Webs 377
Figure 2 Equivalent opening size, Oi, based on (a) equivalent area, (b) equivalent size.
Similarly, the Percent Open Area (POA), εA, that is defined as the percentage of the
open area, Ao, to the total area At, is given by
Ao
εA = × 100 .
At
Usually porosity is determined for materials with a three-dimensional structure, e.g.,

relatively thick nonwoven fabrics. Nevertheless, for two-dimensional textiles such as wo-
ven fabrics and relatively thin nonwovens it is often assumed that porosity and POA are
equal [7].
The size of an individual opening can be defined as the surface area of the opening,
although it is mostly indicated with a diameter called Equivalent Opening Size (EOS). EOS
is not a single value and may differ for each opening. The commonly used term in this case
is the diameter, Oi, corresponding to the equivalent circular area, Ai, of the opening.
Oi = (4 Ai / π )1/ 2 .
This diameter is greater than the side dimension of a square opening. A spherical par-
ticle with that diameter will never pass the opening (Fig. 2a) and may therefore not be con-
sidered as an equivalent dimension or equivalent diameter. This will only be possible if the
diameter corresponds to the side of the square area (Fig. 2b). However, not all openings are
squares, yet the equivalent square area of openings is used to determine their equivalent di-
mension because this simplified assumption results in one single opening size from the open
area. It is the diameter of a spherical particle that can pass the equivalent square opening,
hence the equivalent opening or pore size, Oi, results from
Oi = ( Ai )1/ 2 .
From the EOSs, Pore Size Distribution (PSD) and an equivalent diameter for which a
certain percentage of the opening have a smaller diameter (Ox, pore opening size that x per-
cent of pores are smaller than that size) may be measured.
The PSD curves can be used to determine the uniformity coefficient, Cu, of the inves-
tigated materials. The uniformity coefficient is a measure for the uniformity of the openings
and is given by
Cu = O60 O10 .
The ratio equals unity for uniform openings and increases with decreasing uniformity
of the openings [7].
Pore characteristic is one of the main tools for evaluating the performance of any non-
woven fabric and for electrospun webs as well. Understanding the link between processing
parameters and pore structure parameters will allow for better control over the properties
of electrospun fibers. Therefore there is a need for the design of nanofibers to meet specific
application needs. Various techniques may be used to evaluate pore characteristics of po-
rous materials including sieving techniques (dry, wet and hydrodynamic sieving), mercury
porosimetry and flow porosimetry (bubble point method) [8, 9]. As one goes about selecting
a suitable technique for characterization, the associated virtues and pitfalls of each tech-
nique should be examined. The most attractive option is a single technique which is non-
destructive, yet capable of providing a comprehensive set of data [10].
1.1 Sieving methods

In dry sieving, glass bead fractions (from finer to coarser) are sieved through the porous
material. In theory, most of the glass beads from the first glass bead fraction should pass.
As larger and larger glass bead fractions are sieved, more and more glass beads should be-
come trapped within and on top of the material. The number of pores of a certain size should
be reflected by the percentage of glass beads passing through the porous material during
each glass bead fraction sieved; however, electrostatic effects between glass beads and be-
tween glass beads and the material can affect the results. Glass beads may stick to fibers
making the pores effectively smaller and they may also agglomerate to form one large glass
bead that is too large to pass through the any of the pores. Glass beads may also break from
hitting each other and the sides of the container, resulting in smaller particles that can pass
through smaller openings.
In hydrodynamic sieving, a glass bead mixture is sieved through a porous material un-
der alternating water flow conditions. The use of glass bead mixtures leads to results that
reflect the original glass bead mixture used. Therefore, this method is only useful for eval-
uating the large pore openings such as O95. Another problem occurs when particles of many
sizes interact, which likely results in particle blocking and bridge formation. This is espe-
cially a problem in hydrodynamic sieving because the larger glass bead particles will settle
first when water is drained during the test. When this occurs, fine glass beads which are
smaller than the pores are prevented from passing through by the coarser particles.
In wet sieving, a glass bead mixture is sieved through a porous material aided by a wa-
ter spray. The same basic mechanisms that occur when using the hydrodynamic sieving
method also take place when using the wet sieving method. Bridge formation is not as pro-
nounced in the wet sieving method as in the hydrodynamic sieving method; however, par-
ticle blocking and glass bead agglomeration are more pronounced [8, 9].
The sieving tests are very time-consuming. Generally two hours are required to per-
form a test. The sieving tests are far from providing a complete PSD curve because the ac-
curacy of the tests for pore sizes smaller than 90 µm is questionable [12].
1.2 Mercury porosimetry

Mercury porosimetry is a well known method which is often used to study porous materials.
This technique is based on the fact that mercury as a non-wetting liquid does not intrude
into pore spaces except under applying sufficient pressure. Therefore, a relationship can be
found between the size of pores and the pressure applied.

In this method, a porous material is completely surrounded by mercury and pressure
is applied to force the mercury into pores. As mercury pressure increases the large pores
are filled with mercury first. Pore sizes are calculated as the mercury pressure increases. At
higher pressures, mercury intrudes into the fine pores and when the pressure reaches a max-
imum, total open pore volume and porosity are calculated.
The mercury porosimetry thus gives a PSD based on total pore volume and gives no
information regarding the number of pores of a porous material. Pore sizes ranging from
0.0018 to 400 µm can be studied using mercury porosimetry. Pore sizes smaller than 0.0018
µm are not intruded with mercury and this is a source of error for porosity and PSD calcu-
lations. Furthermore, mercury porosimetry does not account for closed pores as mercury
does not intrude into them. Due to applying high pressures, sample collapse and compres-
sion is possible, hence it is not suitable for fragile compressible materials such as nanofiber
sheets. Other concerns would include the fact that it is assumed that the pores are cylindri-
cal, which is not the case in reality. After the mercury intrusion test, sample decontamina-
tion at specialized facilities is required as highly toxic mercury is trapped within the pores.
Therefore this dangerous and destructive test can only be performed in well-equipped lab-
oratories [6, 8, 9].
1.3 Flow porosimetry (bubble point method)

The flow porosimetry is based on the principle that a porous material will only allow a fluid
to pass when the pressure applied exceeds the capillary attraction of the fluid in largest pore.
In this test, the specimen is saturated with a liquid and continuous air flow is used to remove
liquid from the pores. At a critical pressure, the first bubble will come through the largest
pore in the wetted specimen. As the pressure increases, the pores are emptied of liquid in
order from largest to smallest and the flow rate is measured. PSD, number of pores and po-
rosity can be derived once the flow rate and the applied pressure are known. Flow porosim-
etry is capable of measuring pore sizes within the range of 0.013–500 µm.
As the air only passes through the through pores, the characteristics of these pores are
measured while those of closed and blind pores are omitted. Many times, 100% total flow
is not achieved. This is due to porewick evaporation from the pores when the flow rate is
too high. Extreme care is required to ensure the air flow does not disrupt the pore structure
of the specimen. The flow porosimetry method is also based on the assumption that the
pores are cylindrical, which is not the case in reality. Finding a liquid with low surface ten-
sion, which could cover all the pores, has no interaction with material and does not cause
swelling in material is not easy all the times and sometimes is impossible [6, 8, 9].
1.4 Image analysis

Because of its convenience to detect individual pores in a nonwoven image, it seemed to be
advantageous to use image analysis techniques for pore measurement. Image analysis was
used to measure pore characteristics of woven [11] and nonwoven geotextiles [12]. In the
former, successive erosion operations with increasing size of structuring element was used
to count the pore openings larger than a given structuring element. The main purpose of the
erosion was to simulate the conditions in the sieving methods. In this method, the voids con-
nected to border of the image which are not complete pores are considered in measurement.
Performing opening and then closing operations proceeding pore measurement cause the
pore sizes and shapes deviate from the real ones. The method is suitable for measuring pore
sizes of woven geotextiles with fairly uniform pore sizes and shapes and is not appropriate
for electrospun nanofiber webs of different pore sizes.
In the later case, cross sectional image of nonwoven geotextile was used to calculate
the pore structure parameters. A slicing algorithm based on a series of morphological op-
erations for determining the mean fiber thickness and the optimal position of the uniform
slicing grid was developed. After recognition of the fibers and pores in the slice, the pore
opening size distribution of the cross sectional image may be determined. The method is
useful for measuring pore characteristics of relatively thick nonwovens and cannot be ap-
plied to electrospun nanofiber webs due to extremely small size.
Therefore, there is a need for developing an algorithm suitable for measuring the pore
structure parameters in electrospun webs. In response to this need, we have developed a new
image analysis based method and presented in the following.
In this method, a binary image of the web is used as an input. First of all, voids con-
nected to the image border are identified and cleared using morphological reconstruction
[13], where mask image is the input image and marker image is zero everywhere except
along the border. Total area which is the number of pixels in the image is measured. Then
the pores are labeled and each considered as an object. Here the number of pores may be
obtained. In the next step, the number of pixels of each object as the area of that object is
measured. Having the area of pores, the porosity and EOS regarding to each pore may be
calculated. The data in pixels may then be converted to nm. Finally PSD curve is plotted
and O50, O95 and Cu are determined.
1.4.1 Real webs

In order to measure pore characteristics of electrospun nanofibers using image analysis, im-
ages of the webs are required. These images called micrographs usually are obtained by a
scanning electron microscope (SEM), transmission electron microscope (TEM) or atomic
force microscope (AFM). The images must be of high-quality and taken under appropriate
magnifications.
The image analysis method for measuring pore characteristics requires the initial seg-
mentation of the micrographs in order to produce binary images. This is a critical step be-
cause the segmentation affects the results dramatically. The typical way of producing a
binary image from a grayscale image is by global thresholding [13], where a single constant
threshold is applied to segment the image. All pixels up to and equal to the threshold belong
to object and the remaining belong to the background. One simple way to choose the thresh-
old is picking different thresholds until one is found that produces a good result as judged
by the observer. Global thresholding is very sensitive to any inhomogeneities in the
gray-level distributions of object and background pixels. In order to eliminate the effect of
inhomogeneities, the local thresholding scheme [13] could be used. In this approach, the
image is divided into subimages where the inhomogeneities are negligible. Then optimal
thresholds are found for each subimage. A common practice in this case, which is used in
this study, is to preprocess the image to compensate for the illumination problems and then
apply a global thresholding to the preprocessed image. It can be shown that this process is
equivalent to segment the image with locally varying thresholds. In order to automatically
select the appropriate thresholds, Otsu’s method [14] is employed. This method chooses the
threshold to minimize interaclass variance of the black and white pixels. As it is shown in
Fig. 3 global thresholding resulted in some broken fiber segments. This problem was solved
a) b)
c)
Figure 3 a) A real web, b) Global thresholding, c) Local thresholding.
using local thresholding. Note that, since the process is extremely sensitive to noise con-
tained in the image, before the segmentation, a procedure to clean the noise and enhance
the contrast of the image is necessary.
1.4.2 Simulated webs

In is known that the pore characteristics of nonwoven webs are influenced by web proper-
ties and so are those of electrospun webs. There are no reliable models available for pre-
dicting these characteristics as a function of web properties [15]. In order to explore the
effects of some parameters on pore characteristics of electrospun nanofibers, simulated
webs are generated. These webs are images simulated by straight lines. There are three
widely used methods for generating random network of lines. These are called S-random-
ness, µ-randomness (suitable for generating a web of continuous filaments) and I-random-
ness (suitable for generating a web of staple fibers). These methods have been described in
detail by Abdel-Ghani et al. [16] ] and Pourdeyhimi et al. [17]. In this study, we used µ-ran-
domness procedure for generating simulated images. Under this scheme, a line with a spec-
ified thickness is defined by the perpendicular distance d from a fixed reference point O
located in the center of the image and the angular position of the perpendicular α. Distance
d is limited to the diagonal of the image. Figure 4 demonstrates this procedure.
α
d
O
Figure 4 Procedure for µ-randomness.

No.1 No.2
No.3 No.4
No.5
Figure 5 Micrographs of the electrospun webs.
One of the most important features of simulation is that it allows several structural
characteristics to be taken into consideration with the simulation parameters. These param-
eters are: web density (controlled as line density), angular density (sampled from a normal
or random distribution), distance from the reference point (sampled from a random distri-
bution), line thickness (sampled from a normal distribution) and image size.
2 Experimental
Nanofiber webs were obtained from electrospinning of PVA with average molecular weight
of 72000 g/mol (Merck) at different processing parameters for attaining different pore
characteristics. Table 1 summarizes the electrospinning parameters used for preparing the
webs. The micrographs of the webs were obtained using a Philips XL-30 environmental
scanning electron microscope under magnification of 10000× after being gold coated. Fig-
ure 5 shows micrographs of the electrospun webs.
Table 1 Electrospinning parameters used for preparing nanofiber webs.
No Concentration Spinning distance Voltage Flow rate

(%) (Cm) (kV) (ml/h)
1 8 15 20 0.4
2 12 20 15 0.2
3 8 15 20 0.2
4 8 10 15 0.3
5 10 10 15 0.2
Figure 6 PSD curves of electrospun webs.

Due to previously mentioned reasons, sieving methods and mercury porosimetry are not ap-
plicable for measuring pore structure parameters in nano-scale. The only method which
seems to be practical is flow porosimetry. However, since in this study, the nanofibers were
made of PVA, finding an appropriate liquid for the test to be performed is almost impossible
because of solubility of PVA in both organic and inorganic liquids.
As an alternative, image analysis was employed to measure pore structure parameters
in electrospun nanofiber webs. PSD curves of the webs, determined using the image anal-
ysis method, are shown in Fig. 6. Pore characteristics of the webs (O50, O95, Cu, number of
pores, porosity) measured by this method are presented in Table 2. It is seen that decreasing
Table 2 Pore characteristics of electrospun webs.
No O50 O95 Cu Pore Porosity

No
pixel nm pixel nm
1 39.28 513.9 94.56 1237.1 8.43 31 48.64

2 27.87 364.7 87.66 1146.8 5.92 38 34.57
3 26.94 352.5 64.01 837.4 3.73 64 26.71
4 22.09 289.0 60.75 794.8 3.68 73 24.45
5 19.26 252.0 44.03 576.1 2.73 69 15.74
the porosity, O50 and O95 decrease. Cu also decreases with respect to porosity, that is to say
increasing the uniformity of the pores. The number of pores has an increasing trend with
decreasing porosity.
The image analysis method presents valuable and comprehensive information regard-
ing to pore structure parameters in nanofiber webs. This information may be exploited in
preparing the webs with needed pore characteristics to use in filtration, biomedical appli-
cations, nanoparticle deposition and other purposes. The advantages of the method are list-
ed below:
1) The method is capable of measuring pore structure parameters in any nanofiber webs
with any pore features and it is applicable even when other methods may not be
employed.
2) It is so fast. It takes less than a second for an image to be analyzed (with a 3 GHz
processor).
3) The method is direct and so simple. Pore characteristics are measured from the area
of the pores which is defined as the number of pixels of the pores.
4) There is no systematic error in measurement (such as assuming pores to be cylindrical
in mercury and flow porosimetry and the errors associated with the sieving methods
which were mentioned). Once the segmentation is successful, the pore sizes will be
measured accurately. The quality of images affects the segmentation procedure.
High-quality images reduce the possibility of poor segmentation and enhance the
accuracy of the results.
5) It gives a complete PSD curve.
6) There is no cost involved in the method and minimal technical equipments are needed
(SEM for obtaining the micrographs of the samples and a computer for analysis).
7) It has the capability of being used as an on-line quality control technique for large
scale production.
8) The results obtained by image analysis are reproducible.
9) It is not a destructive method. A very small amount of sample is required for
measurement.
In an attempt to establish the effects of some structural properties on pore characteris-

tics of electrospun nanofibers, two sets of simulated images with varying properties were
generated. The simulated images reveal the degree to which fiber diameter and density af-
fect the pore structure parameters. The first set contained images with the same density
varying in fiber diameter and images with the same fiber diameter varying in density. Each
image had a constant diameter. The second set contained images with the same density and
mean fiber diameter while the standard deviation of fiber diameter varied. The details are
given in Tables 3 and 4. Typical images are shown in Figs. 7 and 8.
Table 3 Structural characteristics of first set images.
No Angular range Line density Line thickness
1 0–360 20 5
2 0–360 30 5
3 0–360 40 5
4 0–360 20 10
5 0–360 30 10
6 0–360 40 10
7 0–360 20 20
8 0–360 30 20
9 0–360 40 20
Table 4 Structural characteristics of second set images.
No Angular range Line density Line thickness
Mean Standard
1 0–360 30 15 0
2 0–360 30 15 4
3 0–360 30 15 8
4 0–360 30 15 10
No.1 No.2
No.3 No.4
No.5 No.6
No.7 No.8
No.9
Figure 7 Simulated images of the first set.

No.1 No.2
No.3 No.4
Figure 8 Simulated images of the second set.
Pore structure parameters of simulated webs were measured using the image analysis
method. Table 5 summarizes the pore characteristics of the simulated images in the first set.
For webs with the same density, increasing the fiber diameter resulted in a decrease in O95,
number of pores and porosity. No particular trends were observed for O50 and Cu. Figures
9 and 10 show the PSD curves of the simulated images in the first set. As the web density
increases, the effects of fiber diameter are less pronounced since the PSD curves of the webs
become closer to each other.For webs with the same fiber diameter, the density increase re-
sulted in a decrease in O50, O95, Cu and porosity whereas the number of pores increased
with the density.
Table 5 Pore characteristics of the first set of simulated images.
No O50 O95 Cu Pore No Porosity
1 27.18 100.13 38.38 84 79.91

2 15.52 67.31 22.20 182 71.78
3 13.78 52.32 18.71 308 69.89
4 36.65 94.31 43.71 67 66.10
5 17.89 61.64 22.67 144 53.67
6 12.41 51.60 16.70 245 47.87
7 24.49 86.90 33.11 58 41.05
8 16.31 56.07 21.66 108 32.53
9 13.11 45.38 17.75 126 22.01
Table 6 Pore characteristics of the second set of simulated images.
No O50 O95 Cu Pore No Porosity
1 14.18 53.56 18.79 133 35.73

2 13.38 61.66 20.15 136 41.89
3 18.14 59.35 22.07 121 41.03
4 15.59 62.71 20.20 112 37.77
a)
b)
c)
Figure 9 PSD curves of the first set of simulated images; effect of density, images with the diameter
of a) 5, b) 10, c) 20 pixels.
Table 6 summarizes the pore characteristics of the simulated images in the second set
set. No significant effects for variation of fiber diameter on pore characteristics were
observed. Suggesting that average fiber diameter is determining factor not variation of
a)
b)
c)
Figure 10 PSD curves of the first set of simulated images; effect of fiber diameter, images with the
density of a) 20, b) 30, c) 40 lines.
diameter. Figure 11 shows the PSD curves of the simulated images in the second set.
4 Conclusion
The evaluation of electrospun nanofiber pore structure parameters is necessary as it facili-
tates the improvement of the design process and its eventual applications. Various
Figure 11 PSD curves of the second set of simulated images, the effect of fiber diameter variation.
techniques have been developed to assess pore characteristics in porous materials. Howev-
er, most of these methods are indirect, have inherent problems and are not applicable for
measuring pore structure parameters of electrospun webs. In this investigation, we have
successfully developed an image analysis-based method as a response to this need. The
method is simple, comprehensive and fast and directly measures the pore structure param-
eters.
The effects of web density, fiber diameter and its variation on pore characteristics of
the webs were also explored using some simulated images. As the fiber diameter increased,
O95, the number of pores and porosity decreased. No particular trends were observed for
O50 and Cu. Increasing the density resulted in a decrease in O50, O95, Cu and porosity
whereas the number of pores increased with density. The effects of variation of the fiber
diameter on pore characteristics were insignificant.
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AUTHORS:
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14 Electrospun Biodegradable
and Biocompatible Natural
Nanofibers: A Detailed
Review
A.K. Haghi and R.K. Haghi

University of Guilan, P.O. Box 3756, Rasht, Iran
email: haghi@guilan.ac.ir
This chapter presents a detailed review on processing of biodegradable and

biocompatible natural nanofibers.
1 Introduction
An emerging technology of manufacturing of thin natural fibers is based on the principle
of the electrospinning process. In conventional fiber spinning, the mechanical force is ap-
plied to the end of a jet, whereas in the electrospinning process the electric body forces act
on an element of charged fluid. Electrospinning has emerged as a specialized processing
technique for the formation of submicron fibers (typically between 100 nm and 1 µm in di-
ameter), with high specific surface areas. Due to their high specific surface area, high po-
rosity and small pore size, the unique fibers have been suggested for a wide range of
applications.
Electrospinning of natural fibers offers unique capabilities for producing novel natural
nanofibers and fabrics with controllable pore structure. Current research effort has focused
on understanding the electrospinning of natural fibers in which the influence of different
governing parameters are discussed.
Electrospinning [1–10] is an economical and simple method used in the preparation of
polymer fibers. The fibers prepared by this method typically have diameters much smaller
than is possible to attain using standard mechanical fiber-spinning technologies [11].
Electrospinning of polymer solutions has gained much attention in the last few years as a
392 A.K. Haghi and R.K. Haghi
Figure 1 Schematic of the electrospinning setup.
cheap and straightforward method to produce nanofibers [12 – 16]. Electrospinning differs
from the traditional wet/dry fiber spinning in a number of ways, of which the most striking
differences are the origin of the pulling force and the final fiber diameters. The mechanical
pulling forces in the traditional industrial fiber spinning processes lead to fibers in the mi-
crometer range and are contrasted in electrospinning by electrical pulling forces that enable
the production of nanofibers [14]. Depending on the solution properties, the throughput of
single-jet electrospinning systems ranges around 10 ml/min. This low fluid throughput may
limit the industrial use of electrospinning. A stable cone-jet mode followed by the onset of
the characteristic bending instability, which eventually leads to a great reduction in the jet
diameter, necessitate the low flow rate [6, 17]. When the diameters of polymer fiber mate-
rials are shrunk from micrometers (e.g. 10 – 100 µm) to submicrons or nanometers, there ap-
pear several amazing characteristics such as a very large surface area to volume ratio (this
ratio for a nanofiber can be as large as 103 times of that of a microfiber), flexibility in sur-
face functionalities, and superior mechanical performance (e.g. stiffness and tensile
strength) compared with any other known form of material. These outstanding properties
make the polymer nanofibers optimal candidates for many important applications [18].
These include filter media [19–21], composite materials [18, 22], biomedical applications
(tissue engineering scaffolds [23–27], bandages [28], drug release systems [29–30]), pro-
tective clothing for the military [31–34], optoelectronic devices and semiconductor mate-
rials [35–38], biosensors/chemosensors [39–42].
1.1 Electrospinning setup

A schematic diagram to interpret electrospinning of polymer nanofibers is shown in Fig. 1.
There are basically three components to fulfill the process: a high voltage supplier, a cap-
illary tube with a pipette or needle of small diameter, and a metal collecting screen. In the
electrospinning process, a high voltage is used to create an electrically charged jet of poly-
mer solution or melt out of the pipette. Before reaching the collecting screen, the solution
jet evaporates or solidifies, and is collected as an interconnected web of small fibers
[43–45]. One electrode is placed into the spinning solution/melt or needle and the other is
attached to the collector. In most cases, the collector is simply grounded. The electric field
is applied to the end of the capillary tube that contains the solution fluid held by its surface
tension. This induces a charge on the surface of the liquid. Mutual charge repulsion and the
Electrospun Biodegradable and Biocompatible Natural Nanofibers 393
contraction of the surface charges to the counter electrode cause a force directly opposite
to the surface tension [46]. As the intensity of the electric field is increased, the hemispher-
ical surface of the fluid at the tip of the capillary tube elongates to form a conical shape
known as the Taylor cone [47]. Further increasing the electric field, a critical value is at-
tained with which the repulsive electrostatic force overcomes the surface tension and the
charged jet of the fluid is ejected from the tip of the Taylor cone [18, 48]. The jet exhibits
bending instabilities due to repulsive forces between the charges carried with the jet. The
jet extends through spiraling loops, as the loops increase in diameter the jet grows longer
and thinner until it solidifies or collects on the target [49].
2 Effect of systematic parameters on electrospun nanofibers

It has been found that the morphology such as fiber diameter and uniformity of electrospun
polymer fibers are dependent on many processing parameters [50]. These parameters can
be divided into 3 groups: a) solution properties, b) processing conditions, c) ambient con-
ditions. Each of the parameters was found to affect the morphology of electrospun fibers.
2.1 Solution properties
Solution parameters such as viscosity of solution, polymer concentration, molecular weight

of polymer, electrical conductivity, elasticity and surface tension, which attribute to poly-
mer and its solution characteristics, have an important effect on the morphology [18].
2.1.1 Viscosity
The viscosity range of different polymer solutions which are spinnable is different. One of
the most significant parameters influencing the fiber diameter is the solution viscosity. A
higher viscosity results in a large fiber diameter. Figure 2 shows the representative images
of beads and beaded fibers for solutions with a range of viscosity. Beads and beaded fibers
are less likely to be formed for more viscous solutions. The diameter of beads becomes big-
ger and the average distance between beads on fibers longer as viscosity increases. Mean-
while, the shape of beads gradually changes from spherical to spindle-like [51].
2.1.2 Solution concentration

The change of the diameter of electrospun fibers as a function of electrical conductivity of
the solutions is shown in Fig. 3. The diameter dropped significantly when the conductivity
increased. Beads were also observed due to the low conductivity of the solution, which re-
sulted in an insufficient elongation of a jet by electrical force to produce uniform fibers.
As the solution concentration increase, a mixture of beads and fibers is obtained [52].
Higher solution concentration may result in fewer beads. The shape of the beads changed
from spherical to spindle-like when the solution concentration varied from low to high lev-
els [18, 53]. The fiber distribution became gradually broader with increasing concentration,
which is consistent with the results obtained by Ryu et al. and Kidoaki et al. [54]. The fiber
diameter increased with increasing solution concentration because the higher viscosity
resisted the extension of the jet [50, 55]. However, it is impossible to electrospin if the
Figure 2 The morphology of beaded fibers versus solution viscosity [51].
20
Figure 3 Solution conductivity effects on the diameter of the electrospun P(LLA-CL) (70/30
wt. %) fibers [50].
solution concentration or the corresponding viscosity is too high due to the difficulty in liq-
uid jet formation [56–58].
Figure 4 shows the morphology of fibers obtained from solutions of 6, 8, 10 and 12
wt. % concentration at applied voltage 10 kV with a constant tip-to-collector distance of
10 cm. Spindle-like beads formed for fibers obtained from solution of 6 wt. %.
a b c d
Figure 4 The morphology of nanofibers at concentrations from 6% to 12% with a constant tip-to-
collector distance of 10 cm at applied voltage 10 kV. (The values below the images and the brackets
show the average fiber diameter (nm) and the standard deviation of fiber diameter. The scale bars for
fibers from solution from 6 to 10 wt.% and 12 wt.% are 2 mm and 5 mm, respectively.). above: Dis-
tribution of fiber diameters obtained at 10 kV with a tip-to-collector distance of 10 cm. The concen-
trations for (a), (b), (c) and (d) are 6, 8, 10 and 12 wt. %, respectively [55].
2.1.3 Molecular weight
Molecular weight also has a significant effect on the rheological and electrical properties
such as viscosity, surface tension, conductivity and dielectric strength. It has been reported
that too low molecular weight polymers tend to form beads rather than fibers and high mo-
lecular weight polymers yield fibers with a larger average diameter [59].
Note that in Figure 5, as the molecular weight increases, several changes can be ob-
served in the electrospun structure of PVA. At a molecular weight of 9000–13,000 g/mol,
Figure 5 Photographs showing the typical structure in the electrospun polymer for various molec-
ular weights. The samples were obtained at the center of the deposition area. (a) 9000–10,000 g/mol;
(b) 13,000–23,000 g/mol; and (c) 31,000–50,000 g/mol (solution concentration: 25 wt. %) [58].
Figure 6 Solution viscosity and surface tension as a function of polymer solution concentration
[60].
the fibrous structure is not completely stabilized and a bead-on-string structure is obtained,
indicating the resistance of the jet to extensional flow.
2.1.4 Surface tension
Surface tension seems more likely to be a function of solvent compositions, but is negligibly
dependent on the solution concentration. Different solvents may contribute different sur-
face tensions. However, not necessarily a lower surface tension of a solvent will always be
more suitable for electrospinning. Generally, surface tension determines the upper and low-
er boundaries of electrospinning window if all other variables are held constant. Formation
of droplets, beads and fibers can be driven by the surface tension of the solution and lower
surface tension of the spinning solution helps electrospinning to occur at a lower electric
field [57].
2.1.5 Number of entanglements
The above variables are not independent of each other. Solution viscosity is a function of
both concentration and polymer molecular weight. These two parameters also affect the
number of entanglements [52].
Figure 6 shows the viscosity and surface tension of solution dissolved in formic acid;
they were 400–5550 cps and 49–68 mN/m, respectively. These values increased with in-
creasing of concentration. Specifically, the viscosity increased from 1400 to 5100 cps for
solution concentration from 25 to 30 wt. % [60].
2.1.6 Solution conductivity

Jun et al. [56] have shown that increasing the solution conductivity by addition of a salt can
significantly aid fiber formation (Fig. 7). Addition of a salt has a positive effect on the elec-
trospinning number (Vq/γR2). Specifically, the electrical energy (Vq) increases. On the oth-
er hand, the change in the surface free energy due to a change in surface tension is not
Figure 7 Effect of NaCl amount in the PVA solution on fiber morphology (DH = 98%, voltage =
5 kV, tip–target distance = 10 cm; flow rate = 0.2 ml/h). NaCl amount based on H2O: (a) 0.05%, (b)
0.10%, (c) 0.15%, (d) 0.2%. Original magnification 10,000 [55].
Figure 8 Electric conductivity as a function of polymer solution concentration [60].
expected to be significant. As the charges carried by the jet increased, higher elongation
forces were imposed to the jet under the electrical field, resulting in smaller beads and thin-
ner fiber diameters (Fig. 8).
A comparison of the diameter of the electrospun fibers with the electrical conductivity
of the solutions is shown in Fig. 9. There was a significant drop in the diameter of the elec-
trospun polymer fibers when the electrical conductivity of the solution increased. Beads
were also observed due to low conductivity of the solution, which results in insufficient
elongation of a jet by electrical force to produce uniform fiber.
Electrospun polymer nanofibers with the smallest fiber diameter were obtained with
the highest electrical conductivity. This suggests that the drop in the size of the fibers was
due to the increased electrical conductivity [50].
2.1.7 Effect of salt addition

Figure 10 shows the SEM images of PAA nanofibers fabricated by electrospinning into so-
lutions with different ionic strengths. Magnified PAA nanofibers are also shown. At a con-
centration of 1 M NaCl, no PAA nanofibers were synthesized. Bead structures were
Figure 9 Solution conductivity effects on the diameter of the electrospun P(LLA-CL)

(70/30 wt. %) fibers [50].
Figure 10 SEM images of PAA nanofibers with different concentrations of NaCl [61].
observed from all prepared PAA nanofibers. With increasing the concentration of NaCl, the
relative viscosity slightly decreased. Variations in viscosity may be caused by the chain
conformation change of PAA in solution upon adding NaCl. With increasing the ionic
strength of the solution, the chain conformation of PAA changes from the extended linear
conformation to the coil conformation because PAA is a polyelectrolyte. Although relative
viscosity decreases slightly with increasing the amount of NaCl [61].
The effect of NaCl addition on the morphology of electrospun PVA fibers was shown
in Fig. 11. Even a little sodium chloride added in the solution made its conductivity to in-
crease sharply. Net charge density carried by the jet in the electrospinning process can be
Figure 11 Effect of NaCl amount in the PVA solution on fiber morphology (DH = 98%, voltage =
5 kV, tip–target distance = 10 cm; flow rate = 0.2 ml/h). NaCl amount based on H2O: (a) 0.05%, (b)
0.10%, (c) 0.15%, (d) 0.2%. Original magnification 10,000 [56].
affected by the conductivity of the solution. PVA fiber diameters were gradually decreased
from 214±19 nm to 159±21 nm with increasing content of NaCl from 0.05% to 0.2%, for
the higher net charge density increased the electrical force exerted on the jet and led to de-
creased fiber diameter. When 0.2% NaCl was added in PVA solution, many small particles,
perhaps sodium chloride crystals, were observed on the SEM micrograph (Fig. 11d).
2.1.8 Solvent
The effects of solvents and their properties on electrospinnability of as-prepared polysty-
rene (PS) solutions and the morphological appearance of as-spun PS fibers were investigat-
ed qualitatively by means of a scanning electron microscope (SEM) in [62]. The eighteen
solvents used were benzene, t-butyl acetate, carbon tetrachloride, chlorobenzene, chloro-
form, cyclohexane, decahydronaphthalene (decalin), 1,2-dichloroethane, dimethylforma-
mide (DMF), 1,4-dioxane, ethylacetate, ethylbenzene, hexane, methylethylketone (MEK),
nitrobenzene, tetrahydrofuran (THF), 1,2,3,4-tetrahydronaphthalene (tetralin) and toluene.
DMF was the best solvent to dissolve PS pellets within six hours, while others were found
to dissolve PS pellets within one to three days, with the exception of hexane which was not
able to dissolve PS pellets after seven days. Only PS solutions in 1,2-dichloroethane, DMF,
ethylacetate, MEK, and THF could produce fibers with high enough productivity, while PS
solutions in benzene, cyclohexane, decalin, ethylbenzene, nitrobenzene, and tetralin were
not spinnable [62].
2.2 Processing condition
2.2.1 Applied voltage

In the case of electrospinning, the electric current due to the ionic conduction of charge in
the polymer solution is usually assumed small enough to be negligible. The only mecha-
nism of charge transport is the flow of polymer from the tip to the target. Thus, an increase
in the electrospinning current generally reflects an increase in the mass flow rate from the
Figure 12 Effect of voltage on morphology and fiber diameter distribution from a 7.4% PVA/water
solution (DH = 98%, tip–target distance = 15 cm, flow rate = 0.2 ml/h). Voltage: (a) 5 kV, (b) 8 kV,
(c) 10 kV, (d) 13 kV. Original magnification 10,000 [56].
capillary tip to the grounded target when all other variables (conductivity, dielectric con-
stant and flow rate of solution to the capillary tip) are held constant [16].
With the increase of the electrical potential the resulting nanofibers became rougher.
It was already reported that a diameter of electrospun fibers was not significantly affected
by an applied voltage. This voltage effect was particularly diminished when the polymer
concentration was low. According to earlier works, higher voltage was reported to induce
not only a larger diameter but also a smaller diameter. Applied voltage may affect some fac-
tors such as the mass of polymer fed out from a tip of needle, elongation level of a jet by
an electrical force, morphology of a jet (a single jet or multiple jets), etc. A balance among
these factors may determine a final diameter of electrospun fibers. It is also noted that bead-
Figure 13 Main effect plots of applied voltage on average fiber diameter [55].
Figure 14 a) SEM micrographs of P(LLA-CL) fibers electrospun from a polymer concentration of

5 wt. % at different applied voltage: (a) 9 kV, (b) 12 kV and (c) 15 kV. b) Relation between fiber
diameter and applied voltage in the electrospinning with 5 wt. % P(LLA-CL) solution [64].
ed fibers have been found to be electrospun with too high a level of applied voltage. Al-
though voltage effects show different tendencies, the voltage did not show a significant role
in controlling the fiber morphology [59].
A series of experiments were carried out when the applied voltage was varied from 5
to 13 kV and the tip to target distance was held at 15 cm. The results are shown in Fig. 12.
There was a slightly increase in average fiber diameter with increasing applied electric field.
A considerable amount of thin fibers with diameters below 150 nm were found when the
applied voltage was above 10 kV. A narrow distribution of fiber diameters was observed at
a lower voltage of 5 kV, while a broad distribution in the fiber diameter was obtained at
higher applied voltages of 10 – 13 kV. Increasing the applied voltage, i.e., increasing the
electric field strength will increase the electrostatic repulsive force on the fluid jet which
favors the thinner fiber formation. On the other hand, the solution will be removed from the
capillary tip more quickly as jet is ejected from the Taylor cone. This results in the increase
of the fiber diameter. Corona discharge was observed at voltages above 13 kV, making elec-
trospinning impossible [56]. Also, an increase in voltage from 9.2 kV to 25 kV did not in-
duce a significant change in the mean diameter of the fibers [54]. The diameter of
electrospun PAN fibers did not change significantly over the range of applied voltage for
the various solution concentrations in the experimental region [56].
SEM micrographs of nanofibers electrospun at different electrospinning voltage from
a constant polymer concentration of 5 wt. % are shown in Fig. 14a. A fiber diameter tended
to decrease with increasing electrospinning voltage, although the influence was not as great
as that of polymer concentration. Figure 14b shows fiber diameter as a function of electro-
spinning voltage. The increase of the electrospinning voltage causes an increase of the elec-
trostatic stress on the jet, which may be analogous to an increase of the draw rate in
conventional fiber spinning [64].
2.2.2 Feed rate

The morphological structure can be slightly changed by changing the solution flow rate as
shown in Fig. 15. At the flow rate of 0.3 ml/h, a few big beads were observed on the fibers.
The flow rate could affect the electrospinning process. When the flow rate exceeded a crit-
ical value, the delivery rate of the solution jet to the capillary tip exceeded the rate at which
the solution was removed from the tip by the electric forces. This shift in the mass balance
resulted in a sustained but unstable jet and fibers with big beads were formed [56].
Figure 16 shows that the diameter of the electrospun HM-PLLA fibers was not signif-
icantly changed with the varied volume feed rate. The influence due to the volume feed rate
also diminished when the polymer concentration was low.
The solution’s electrical conductivity was found to be the dominant parameter to
Figure 15 Effect of flow rate of 7% PVA water solution on fiber morphology (DH = 98%, voltage
= 8 kV, tip–target distance = 15 cm). Flow rate: (a) 0.1 ml/h; (b) 0.2 ml/h; (c) 0.3 ml/h. Original mag-
nification 10,000 [56].
Figure 16 Volume feed rate effects on the diameter of the PLLA (Mw: 300 K) fibers electrospun
from solutions with different polymer concentration [50].
control the morphology of electrospun polymer fibers [9]. In the case of low-molecu-
lar-weight liquid, when a high electrical force is applied, formation of droplets can occur.
A theory proposed by Rayleigh explains this phenomenon. As evaporation of a droplet
takes places, the droplet decreases in size. Therefore, the charge density of its surface is in-
creased. This increase in charge density due to Coulomb repulsion overcomes the surface
tension of the droplet and causes the droplet to split into smaller droplets. However, in the
case of a polymer solution (high molecular weight liquid) the emerging jet does not break
up into droplets, but is stabilized and forms a string of beads connected by a fiber. As the
concentration is increased, a string of connected beads is seen, and with a further increase
there is reduced bead formation until only smooth fibers are formed. And sometimes spin-
dle-like beads can form due to the extension caused by the electrostatic stress. The changing
of fiber morphology can probably be attributed to a competition between surface tension
and viscosity. As concentration was increased, the viscosity of the polymer solution in-
creased. The surface tension attempted to reduce the surface area per unit mass, thereby
caused the formation of beads/spheres. Viscoelastic forces resisted the formation of beads
and allowed for the formation of smooth fibers. Therefore formation of beads at lower poly-
mer solution concentration (low viscosity) occurred where surface tension had a greater in-
fluence than the viscoelastic force. However, bead formation was reduced and finally
eliminated at higher polymer solution concentration, where viscoelastic forces had a greater
influence in comparison with surface tension. But when the concentration was too high,
high viscosity and rapid evaporation of solvent made the extension of jet more difficult,
thicker and nonuniform fibers were formed [55].
2.2.3 Distance of needle tip to collector

Tip–target distance had no significant effect on the electrospun fiber morphology of fully
hydrolyzed PVA, as shown in Fig. 17. Micrographs are indistinguishable for electrospin-
Figure 17 Effect of tip–target distance on fiber morphology from a 7.4% PVA / water solution (DH
= 98%, voltage = 5 kV, flow rate = 0.2 ml/h). Tip–target distance: (a) 8 cm, (b) 10 cm, (c) 12 cm, (d)
15 cm. Original magnification 10,000 [56].
Figure 18 Processing map obtained based on the systematic parameter study: (a) jet elongation / an
electrical force (affected by electrical conductivity of solvents, applied voltage), (b) mass of polymer
(affected by polymer concentration, applied voltage, volume feed rate) [50].
ning at 8 –15 cm of the tip–target distance. It was assumed that solution jets were elongated
and solidified quickly after they flowed out of the needle tip because of the high conduc-
tivity of fully hydrolyzed PVA used [56].
Based on the processing parameter studies, all the parameters’ effects on the morphol-
ogy of the electrospun nanofibers were summarized in a processing map (Fig. 18). A suit-
able level of processing parameters must be optimized to electrospin polymers into
nanofibers with a desired morphology and the parameters’ levels are dependent on the prop-
erties of polymers and solvents used in each electrospinning process. Understanding of how
each of processing parameter affects the morphology of the electrospun nanofibers is es-
sential. All the parameters have been divided into two groups; i.e. one with parameters
which affect the mass of polymer fed out from a tip of needle, and the other with parameters
which affect an electrical force during electrospinning. Polymer concentration, applied
voltage and volume feed rate were considered to affect the mass of polymer. Polymer con-
centration and feed rate directly reflect the mass of polymer. Increased polymer concentra-
tion and feed rate tend to bring more mass of polymer into the polymer jet. It is noteworthy
that the minimum polymer concentration to electrospin uniform fibers was determined by
the molecular weight of polymer. High molecular weight of polymer provides a sufficient
level of solution viscosity to produce a uniform jet during electrospinning even when poly-
mer concentration is relatively low. Applied voltage reflects the force to pull a solution out
from the needle hence higher applied voltage causes more solution coming out. On the other
hand, it was considered that solution electrical conductivity and applied voltage affect a
charge density thus an electrical force, which acts to elongate a jet during electrospinning.
Hence, higher solution electrical conductivity and applied voltage increase the jet elonga-
tion. Therefore, it is summarized that electrospun fibers with a smaller diameter can be pro-
duced with lower polymer concentration, feed rate and applied voltage when the effect of
the mass of polymer dominates to determine the final diameter of electrospun fibers, while
a smaller diameter of fibers can be electrospun with higher solution electrical conductivity
and applied voltage when the effect of the jet elongation is dominant. For both cases,
non-uniform / beaded fibers were found if the parameters were either too high or too low.
In fact, applied voltage affects both the polymer mass and jet elongation; however, the ef-
fect is not as dominant as the other parameters for controlling the morphology of electro-
spun fibers. It must be noted that polymer concentration, molecular weight and solution
electrical conductivity play a prime role in determining the morphology of electrospun fi-
bers. Polymer fibers with a smaller diameter can be electrospun with higher electrical con-
ductivity of solution and lower polymer concentration which can be further decreased by
higher molecular weight of polymer [50].
3 Theory and modeling

Though easily realizable in the laboratory, electrospinning is a complex phenomenon to an-
alyze because of the coupling between the electric field and the deformation of the fluid,
the latter in turn determined by the rheology of the material. Typically, electrospinning has
two stages. In the first stage, the polymer jet issues from a nozzle and thins steadily and
smoothly downstream. In the second stage, the thin thread becomes unstable to a non-
axisymmetric instability and spirals violently in large loops. The enormously increased con-
tour length produces a very large stretch ratio and a nanoscale diameter [9]. For the steady
stretching in the first stage, Spivak and Dzenis [65] published a simple model that assumes
the electric field to be uniform and constant, unaffected by the charges carried by the jet.
Reneker et al. modeled the viscoelasticity of the jet by a linear Maxwell equation [9].
Hohman et al. [67, 68] developed a slender-body theory for electrospinning that couples jet
stretching, charge transport and electric field. The model encounters difficulties, however,
with the boundary condition at the nozzle.
For the second stage, the bending instability has been carefully documented by two
groups (Reneker et al. [69, 70] and Shin et al. [71]); each has proposed a theory for the
Figure 19 Momentum balance on a short section of the jet [9].
instability. Reneker et al. modeled the polymer jet by a linear Maxwell equation.
Like-charge repulsion generates a bending force that destabilizes the jet. Hohman et al. [67]
built an electrohydrodynamic instability theory, and predicted that under favorable condi-
tions, a nonaxisymmetric instability prevails over the familiar Rayleigh instability and a
varicose instability due to electric charges. In theoretical work to date, the rheology of the
polymer jet has been represented by a Newtonian viscosity [67, 68], a power-law viscosity
[66] and the linear Maxwell equation [9, 69, 71].
The jet is governed by four steady-state equations representing the conservation of
mass and electric charges, the linear momentum balance and Coulomb’s law for the field
E [9]. Mass conservation requires that
π R2υ = Q, (1)
where Q is a constant volume flow rate. Charge conservation may be expressed by
πR2KE + 2 π Rυσ = I, (2)
where E is the z component of the electric field, K is the conductivity of the liquid, and I is
the constant total current in the jet. The momentum equation is formulated by considering
the forces on a short segment of the jet (Fig. 19):
d d γ
(π R 2 ρυ 2 ) = π R 2 ρ g + [π R (− P + τ zz )] + ⋅ 2π RR ′ + 2π R (tte − tne R ′) , (3)
dz dz R
where τzz is the axial viscous normal stress, p is the pressure, γ is the surface tension, and
t et and t en are the tangential and normal tractions on the surface of the jet due to electricity.
The prime indicates the derivative with respect to z, and R´ is the slope of the jet surface.
The ambient pressure has been set to zero. The electrostatic tractions are determined by the
surface charge density and the electric field:
ε σ 2 ε′−ε 2
tne = ( En2 − Et2 ) ≈ − E , (4)
2 2ε 2
tte = σ Et ≈ σ E , (5)
where ε and ε are the dielectric constants of the jet and the ambient air, respectively, En
Figure 20 A vast classification chart for fibers [72].
and Et are the normal and tangential components of the electric field at the surface, and ||*||
indicates the jump of a quantity across the surface of the jet. We have used the jump con-
ditions for En and Et: ||ε En|| = ε E – εEn = σ, ||Et|| = E t – Et = 0, and assumed that εEn <<
(see Ganan–Calvo [13]) and Et ≈ E. The overbar indicates quantities in the surrounding air.
The pressure p(z) is determined by the radial momentum balance, and applying the normal
force balance at the jet surface leads to
γ
− p + τ rr = tne − , (6)
R
Inserting Eqs. (4)–(6) into Eq. (3) yields:
3 d γ R′ σσ ′ 2σ E
ρυυ ′ = ρ g + (η R 2υ ′) + + + (ε − ε ) EE ′ + . (7)
R 2 dz
R 2 ε ′ R
4 Natural fibers
The fibers in modern textile manufacture can be classified into two groups: natural and
man-made fibers. A vast classification of fibers is illustrated in Fig. 20. Natural fibers are
those provided by nature in a ready-made fibrous form. Natural fibers can be subdivided
into three main classes, according to the nature of their source:
• vegetable fibers
• animal fibers
• mineral fibers.
Vegetable fibers include cotton – the most important of all textile fibers – together with
flax, hemp, jute and other fibers, which are produced by plants. They are based on cellulose,
the material used by nature as a structural material in the plant world.
Figure 21 Silk micrograph (ITF) [73].
Animal fibers include wool and other hair-like fibers, and fibers such as silk, produced
as filaments by cocoon-spinning creatures. These animal fibers are based on proteins, the
complex substances from which much of the animal body is made [72].
5 Electrospinning of silk fibers
5.1 Introduction
The silk fiber protein is synthesized by the silk gland cells and stored in the lumen of silk
gland. Subsequently, it is converted into silk fibers. Each strand of silk fiber is a double
structure with two fibroin filaments covered by sericin (Fig. 21).
In fibroin, alanine and glycine together account for 70% of the total composition,
whereas in sericin they make up about 15%. The chief component of sericin is another ami-
no acid, serine (30% of the total).
Silk is the only natural fibre which exists as a continuous filament. Each Bombyx mori
cocoon can yield up to 1600 meters of filament. These can be easily joined together using
the adhesive qualities of sericin to form a theoretically endless filament.
The silk fiber’s triangular cross-section gives it excellent light reflection capability.
The silk fiber is smooth, unlike those of wool, cotton and others. This is one of the rea-
sons why silk fabrics are so lustrous and soft.
Silk can absorb moisture up to 30% of its weight without creating a damp feeling.
When moisture is absorbed, it generates ‘wetting-heat’ which helps to explain why silk is
comfortable to wear next to the skin.
Silk has a tenacity of approximately 4.8 grams per denier, slightly less than that of ny-
lon.
Silk has poor resistance to ultraviolet light and for this reason is only recommended
for those curtains that are lined or not exposed to direct sunlight [73].
5.2 Crystal structure of silk (fibroin) at various stages of electrospinning

The physiological properties of SF matrices strongly depend on its molecular conformation
and surface texture. SF exhibits at least three crystalline forms: silk I, silk II, and alpha-
helix. All three conformations can be formed from the appropriate preparation conditions
Figure 22 ATR-IR spectra of as-prepared and methanol-treated SF matrices: (a) as-spun SF-N; (b)
methanol-treated SF-N; (c) as-cast SF-F; (d) SF-M [75].
and each is interchangeable under certain conditions. Crystallization of SF involves the con-
formational transition that can be easily induced by simple physical (thermal), mechanical
or chemical treatments. The most common method to convert the distorted conformation
(random coil or silk I) of SF into the more stable β-sheet (silk II) conformation is to treat
the SF film with an organic solvent. It is well known that organic solvents, particularly
methanol, are highly effective in crystallizing SF from a distorted conformation to the
α-sheet.
Infrared spectroscopy (IR) has been often applied to study the molecular conformation
of silk fibroin fibers or films. The sensitive absorption bands on the IR spectrum are located
in the spectral regions of ~1625 cm–1 (amide I), ~1528 cm–1 (amide II), ~1230 cm–1 (amide
III) and ~700 cm–1 (amide V). To characterize the structure of SF matrices, the ATR-IR
spectra in the amide I and II regions are examined. The IR spectra of SF matrices are shown
in Fig. 22(a)–(c). The as-spun SF-N matrix was characterized by absorption bands at 1651
cm–1 (amide I) and 1528 cm–1 (amide II), attributed to the random coil conformation, as
shown in Fig. 22a. The methanol-treated SF-N matrix showed strong β-sheet absorptions
at 1622 and 1514 cm–1 within a methanol-treating time of 10 min, indicating that the ran-
dom coil conformation of the SF nanofibers rapidly converted into β-sheet structure
(Fig. 22(b)) [75].
On the contrary, both as-prepared SF-F and SF-M matrices showed strong β-sheet ab-
sorptions at 1622 and 1514 cm–1 without methanol treatment, as shown in Fig. 22(c) and
(d). The raw SF had highly ordered intermolecular hydrogen bonds (β-sheet), and the SF
film took a mainly β-sheet conformation, when cast from the formic acid solution.
Solid-state 13C NMR has been shown to be a more effective analytical tool for dem-
onstrating the formation of β-sheets in polypeptides and proteins, because the isotropic 13C
NMR chemical shifts of carbon atoms in proteins are sensitive to the β-sheet’s secondary
structure. It is well established that SF conformations are dependent upon the species of
silkworms and conditions of sample preparation. In particular, it has been reported that fi-
broin from Bombyx mori adopts two dimorphic structures, silk I and silk II. The silk II form
is identified by the 13C chemical shifts of glycine (Gly), serine (Ser), and alanine (Ala) that
are indicative of β-sheets, while the silk I form produces chemical shifts that are associated
with a loose helix or distorted β-turn. However, when compared with silk II, the less stable
silk I shows a relatively unresolved structure, and the conformation of the soluble form of
Figure 22 13C CP/MAS NMR spectra (a) and the expanded 13C NMR spectra of the methyl alanine
region (b) of SF matrices. The dotted arrow indicates the chemical shift of alanine in the β-sheet con-
formation [75].
SF rapidly undergoes a transition to the insoluble silk II conformation. In 13C CP/MAS

NMR structural analyses of B. mori silk fibroins, the two crystalline forms of silk fibroins,
silk I and silk II (β-sheet), have been distinguished by the conformation-dependent 13C
chemical shifts of the respective amino acid residues. Figure 22 shows 13C CP/MAS NMR
spectra (a) and the expanded 13C NMR spectra of the methyl alanine region (b) of SF ma-
trices. The dotted arrow indicates the chemical shift of alanine in the β-sheet conformation.
Solid-state 13C NMR spectra of the as-prepared SF-N and SFF matrices, together with that
of the SF-M. Assignments were made according to the literature. The chemical shifts of the
Ala residue in Fig. 22a were 17.6 ppm for AlaCβ, 49.9 ppm for AlaCα, and 173.5 ppm for
Ala C=O carbon. The observed 13C NMR chemical shifts of the peaks suggest that the Ala
residue of the SF-N matrix takes a mainly non β-sheet conformation (random coil and silk
I). The vertical dashed line and arrow in Fig. 22 show the chemical shift of Ala in the β-sheet
conformation. A shoulder was detected at ~20.4 ppm and assigned to Ala Cβ in a β-sheet
conformation, indicating that as-spun SF-N take some β-sheet conformations. The β-sheet
structure of as-spun SF nanofibers could be formed partially by the elongational forces dur-
ing the electrospinning process. However, the characteristic resonances of SF-F matrix, es-
pecially the Ala Cβ at 20.4 ppm and Ala Cα at 48.9 ppm are similar to that of the SF-M
matrix [75].
Shimizu has reported that there are two types of crystal formation during solidification
of liquid fibroin (by losing water) inside the silk gland. One of them corresponds to the crys-
tal form of fibrous fibroin. The fibroin molecular chain which constitutes the spatial crystal
form is considered to be the intermediate form during the course of shifting the molecular
chain of liquid silk to the fibrous fibroin molecule chain, that is, β-chain. It is therefore
named the α-type of fibroin. The condition for the appearance of a type thereafter include
solidification when there is no stress from liquid silk and the temperature being less than
40°C. It becomes β-type when the temperature is above 50°C. Both α- and β-types of none
stretched are insoluble in water and the α-type is softer than the β-type [76].
• β-structure of silk
β-structure is the structure where the polypeptide chain is elongated. The structure can
be the type where all the molecular chains run in the same direction and form a parallel
pleated sheet or the type where the molecular chains run in the alternate direction and form
an antiparallel chain pleated sheet. In the case of a β-structure, there are three important fea-
tures, namely the period that is repeating period of the polypeptide chain, the spacing of the
molecular chain in the sheet and the distance between the sheets [76].
5.3 Spinning dope preparation for electrospinning
5.3.1 Degumming
The natural gum sericin is normally left on the silk during reeling, throwing and weaving.
It acts as a size, which protects the fiber from mechanical injury. The gum is removed from
finished yarns or fabrics, usually by boiling with soap and water [73]. The fibers were heat-
ed in an aqueous Na2CO3 (0.02 M) or 0.5% (w/w) NaHCO3 and rinsed with water to extract
sericin [75, 77].
5.3.2 Dissolving of fibroin

The extracted fibers (degummed silk) were then dissolved in 50% CaCl2 [77] (100°C, then
cooled) or in 9.3 M LiBr solution at 60°C yielding a solution. The solution is poured into
regenerated cellulose dialysis tubing to carry out dialysis against 1000 ml deionized water
(for 48 h at 23°C). Another way reported by Min et al. [75, 76]: degummed silk (SF) is dis-
solved in ternary CaCl2/CH3CH2OH/H2O (1:2:8 in molar ratio) at 70°C for 6 h and then
dialyzed with cellulose tubular membrane. After these stages the regenerated silk fibroin
sponge is obtained by lyophilization. The silk sponge solution is electrospun in formic acid
(98–100%).
5.4 Electrospinning of silk fibroin

Electrospinning generally produces non-woven matrices with randomly arranged, ultrathin
fibers that have nanometer scale diameters. Figure 23 shows a SEM micrograph (magnifi-
cation 500×) of the woven SF microfiber and as-spun SF nanofiber matrix. From the image
analysis, the SF nanofibers have an average diameter of 80 nm and their diameters range
from 30 to 120 nm, while the diameter of the SF microfiber is 11 nm. The average diameter
of SF nanofibers is about two orders of magnitude smaller than a SF microfiber [75].
Figure 23 The morphology of fibers at electric field of 3 kV/cm at concentrations from 5 to 19.5%
with a constant spinning distance of 7 cm. The figure also shows the average, standard deviation,
maximum and minimum values of the fiber diameter [77].
5.4.1 Effect of silk polymer concentration on fiber diameter

Silk concentration plays a major role in fiber diameter. No fibers were formed at less than
5% silk concentration for any electric field and spinning distances. Figures 24 and 25 show
the morphology of fibers obtained at electric fields of 3 and 4 kV/cm, respectively, at
silk/formic acid concentrations of 5, 8, 10, 12, 15, and 19.5% with a constant tip-to-collec-
Figure 24 SEM image of woven SF microfibers and nonwoven SF nanofibers [75].
tion plate distance of 7 cm. At a 8% concentration, less than a 30 nm diameter fibers were
formed with beads (drops of polymer over the woven mesh); they were not uniform and
were branched off (Fig. 25). At a 10% concentration with 5 cm spinning distance and 2, 3
and 4 kV/cm electric fields, drops were formed instead of fibers. Continuous fibers were
Figure 25 The morphology of fibers at electric field of 4 kV/cm at concentrations from 5 to 19.5%
with a constant spinning distance of 7 cm [77].
obtained above 12% regardless of electric field and distance (Figs. 24 and 25). At 19.5%,
the average fiber diameter was much larger than that of fibers spun at lower concentrations.
In the short distance as well as low concentration (10%), the solution reaches the collection
plate before the solvent fully evaporates. This explains the formation of droplets and beads
at the low concentration and distance. Fewer beads were observed in electrospun fibers at
higher concentration. Increase in the regenerated silk concentration in the formic acid in-
creases the solution viscosity. At low concentrations beads are formed instead of fibers and
at high concentrations the formation of continuous fibers are prohibited because of the in-
ability to maintain the flow of the solution at the tip of the needle resulting in the formation
of larger fibers. Continuous nanofibers were obtained above 12% regardless of electric field
and distance and at higher concentration of 19.5% the average fiber diameter was larger
than at lower concentrations [77].
The SEM micrographs of nanofibers electrospun from SF solutions with different con-
centrations or viscosities ranged from 3% to 15% by weight are shown in Fig. 26. At a con-
centration below 9% by weight, beads or beaded fibers were generated by electrospinning.
Continuous nanofibers can be obtained at a concentration above 12% by weight, and this
Figure 26 SEM micrographs of electrospun SF nanofibers with concentration of (a) 3%, (b) 6%, (c)
9%, (d) 12%, or (e) 15% by weight [75].
concentration appears to correspond to the onset of significant chain entanglements in the

viscosity data shown in Fig. 26. Therefore, it can be concluded that extensive chain entan-
glements are necessary to produce continuous fibers by electrospinning [75].
5.4.2 Effect of voltage and spinning distance on morphology and diameter

Figure 27 shows the relationship between the mean fiber diameter and electric field with
concentration of 15% at spinning distances of 5, 7 and 10 cm. The mean fiber diameter ob-
tained at 2 kV/cm is larger than other electric fields. The effect of two factors, concentration
and electric field, on fiber diameter was investigated by two-way analysis of variance. The
interaction effect between two factors is also obtained from this analysis [77].
Figure 27 The relationship between mean fiber diameter and electric field with concentration of
15% at spinning distances of 5, 7 and 10 cm (please refer to text for explanation) [77].
Figure 28 The relationship between the fiber diameter and the concentration at three electric fields
(2, 3, 4 kV/cm) [77].
Figure 28 shows that concentration apparently has a greater effect on the fiber diameter
than electric field. The multiple regression analysis was carried out to evaluate the contri-
bution of concentration and electric field on the fiber diameter [77].
Sukigara et al. [78] used RSM analysis (Response Surface Methodology) of the ex-
perimental results to develop a processing window which will produce nanoscale regener-
ated silk fibers by the electrospinning process. RSM is used in situations where several
variables influence a feature (called the response) of the system. The steps in the procedure
are described briefly as follows.
1. Identification of variables ζ1; ζ2; ζ3… for response η.

2. Calculation of corresponding coded variables x1; x2; x3… by using the following
equation:
ζ i − [ζ Ai + ζ Bi ] / 2
xi = ,
[ζ Ai + ζ Bi ] / 2
where ζAi and ζBi refer to the high and low levels of the variables ζi; respectively.
3. Determination of the empirical model by multiple regression analysis to generate
theoretical responses (y): The second-order model is widely used in RSM. The general
equation for response η of the second-order model is given by:
k k k
η = βο + ∑ βi xi + ∑ βii xi2 + ∑ ∑ βij xi x j ,
i =1 i =1 i< j =2
Figure 29 RSM procedure to optimize the electrospinning condition of regenerated silk [78].
where k is the number of factors, xi are the coded variables and β are coefficients.
4. Calculation of the coefficients β to fit the experimental data as closely as possible.
The relationship between the response and the variables is visualized by a response
surface or contour plot to see the relative influence of the parameters, to find an optimum
parameter combination, and to predict experimental results for other parameters. For two
variables, when k = 2, the empirical model from the general equation (2) becomes
y = βο x1 + β1 x1 + β 2 x2 + β11 x12 + β 22 x22 + β12 x1 x2 + ε .
The RSM procedure to optimize the process parameters for the electrospinning silk is
shown in Fig. 29.
Sukigara et al. [78] designed a factorial experiment by using two factors (electric field
and concentration). For a quadratic model, experiments must be performed for at least three
levels of each factor. These levels are best chosen equally spaced. The two factors (silk con-
centration and electric field) and three levels resulted in nine possible combinations of fac-
tor settings. A schematic of the experimental design is shown in Fig. 30 (A) and (B).
5.5 Characterization
The conformational changes of the secondary structure of silk fibroin, which occur during
the electrospinning process, were analyzed by Raman spectroscopy. The spectra for pris-
tine, degummed and electrospun fibers are shown in Fig. 31 (A). Pristine and degummed
silk fibroins display characteristic conformational bands in the range 1650–1667 and
Figure 30 Experimental design (A) spinning distance 7 cm and (B) spinning distance 5 cm. The
values at the coordinate points show the mean fiber diameter (nm) of 100 measurements and coded
values are shown in brackets (electric field and concentration). NF: no fiber formation [78].
1241–1279 cm–1, which correspond to amide I and complex amide III, respectively. In this
study, the amide I (random coil) pristine band was observed at 1665 cm–1 and the amide III
(β-sheet) pristine band at 1231 cm–1. These well-defined bands were chosen because they
give a clear indication of changes in the secondary structures from random to β-sheet. The
degummed silk also shows absorption bands at these wavelengths. No significant spectral
changes were observed indicating that the fibroin conformation is unchanged during the de-
gumming process. The Raman spectra of the electrospun fiber are essentially the same as
that of the pristine and degummed fibers although minor bands and some differences in
peak intensities appear. This shows that the electrospinning process preserves the natural
conformation of fibroin. Figure 31 (B) shows that the amide I (1665 cm–1, random) to amide
III (1228 cm–1, β-sheet) ratio of the electrospun fiber is less than that of the pristine fiber.
This means that the electrospun fiber has a higher β-sheet content than the pristine fiber.
6 Electrospinning of cellulose and cellulose acetate

Cellulose is the most abundant naturally occurring polysaccharide formed out of glu-
cose-based repeat units, connected by 1,4-beta-glucosidic linkages. Cellulose and its deriv-
atives are widely used as tough versatile materials. Cellulose nitrate, cellulose acetate (CA)
pristine
degummed
electrospun
Figure 31 (A) Raman spectroscopy of pristine, degummed and electrospun nonwoven silk mat. (B)
Secondary structural compositions of silk fibroin showing the fraction of amide I to amide III confor-
mations [79].
and cellulose xanthate (rayon) can be easily molded or drawn into fibers for textile appli-
cations, for designing composite materials (safety glass), as thermoplastics etc. [80].
6.1 Electrospinning of CTA solution

CTA was dissolved in MC or solvent mixtures of MC/EtOH (90 / 10, 85 / 15, and 80 / 20,
v/v). The needle (ID = 0.495 mm) was connected to a high voltage supply (Chungpa EMT,
CPS-40K03), which can generate positive DC voltages up to 40 kV. The distance between
the needle tip and the collecting target was 10 cm. Positive voltages applied to polymer so-
lutions were 15 kV. CA solutions were delivered via a syringe (20 mL) with a mass flow
rate of 1 mL/h. The optimum concentration of CTA solution for fiber formation was 5
wt. %. All electrospinnings were carried out at room temperature [81].
Figure 32 shows SEM photographs of ultrafine CTA fibers electrospun from a 5 wt. %
CTA solution in MC. CTA fibers showed a mixed pattern of flat ribbons and round shapes
(Fig. 32(a)). These flat ribbons were wrinkled or twisted in an irregular way. At a higher
magnification (×20,000), it was found that ultrafine CTA fibers had isolated circular pores
with a narrow size distribution in the range of 50–100 nm (Fig. 32(b)). The porous structure
was induced by phase separation resulting from the rapid evaporation of MC during the
electrospinning process. The polymer-rich phase formed the fiber matrix and solvent-rich
phase gave rise to pores. Figure 32c shows a SEM image for the fractured cross-section of
Figure 32 SEM images of electrospun CTA fibers: (a) magnification ×1000, (b) magnification
×20,000, and (c) fractured cross-section of porous CTA fibers [81].
a porous CTA fiber. Isolated pores were also observed inside the fiber, which were larger
than those in the fiber surface. In liquid–liquid demixing of polymer solution, the resulting
polymer morphology is dependent on the phase separation mechanism. The structure
formed by the spinodal decomposition (SD) mechanism shows interconnected pores,
whereas the nucleation and growth (NG) mechanism mainly results in isolated spherical
pores [81].
Figure 33 shows SEM photographs of ultrafine CTA fibers electrospun from different
ratios of MC/ EtOH (90 / 10, 85 / 15, and 80 / 20 v/v). All ultrafine CTA fibers were elec-
trospun with the same electrospinning condition for a CTA solution in MC. It was found
that addition of EtOH changed not only the pore structure but also the fiber diameter. In the
case of MC/EtOH (90 / 10), the resulting CTA fibers had interconnected pores in the range
of 200–500 nm, indicating that the phase separation proceeded according to the SD mech-
anism. However, non-porous corrugated fibers were obtained from other mixed solvents
(Figs. 33(b) and (c)) [81].
Figure 33 SEM images of ultrafine CTA fibers electrospun using different volume ratios of
MC/EtOH: (a) 90 / 10, (b) 85 / 15, and (c) 80 / 20 [81].
7 Concluding remarks
Nanotechnology has become in recent years a topic of great interest to scientists and engi-
neers, and is now established as the prioritized research area in many countries. The reduc-
tion of size to the nanometer range brings an array of new possibilities in terms of material
properties, in particular with respect to achievable surface to volume ratios. Electrospinning
of natural fibers is a novel process for producing superfine fibers by forcing a solution
through a spinnerette with an electric field. A comprehensive review on this technique has
been made in this paper. Based on this review, many challenges exist in the electrospinning
process of natural fibers, and a number of fundamental questions remain open. The electro-
spinning technique provides an inexpensive and easy way to produce natural nanofibers on
low basis weight, small fiber diameter and pore size. It is hoped that this chapter will pave
the way toward a better understanding of the application of electrospinning of natural fibers.
Nevertheless, among the biodegradable and biocompatible polymers, SF was exten-
sively studied as one of the candidate materials for biomedical applications, because it has
several distinctive biological properties including good biocompatibility, biodegradability,
and minimal inflammatory reaction. One of the promising applications of SF in biomedical
engineering are scaffolding materials for tissue engineering. It was reported that SF matri-
ces could be useful for the culture of fibroblasts and osteoblasts as well as stem cells, and
could enhance the adhesion, growth, and differentiation of the cells in a manner similar to
that of collagen matrices. Nonwoven matrices of electrospun nanofibers could be prepared
from a regenerated SF solution, and the matrices were effective in cell attachment and
spreading of normal human keratinocytes and fibroblasts [82].
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Nomenclature
R = radius of the jet
R ' = slope of the jet surface
Q = constant volume flow rate
I = constant total current in the jet
σ = surface charge density
υ = velocity
E = z component of the electric field
K = conductivity of the liquid
γ = surface tension
Et = tangential components of the electric field at the surface
En = normal components of the electric field at the surface
g = gravity
τ zz = axial viscous normal stress
P = pressure
t ne = normal traction on the surface of the jet due to electricity
t te = tangential traction on the surface of the jet due to electricity.
ε = dielectric constants of the jet
ε– = dielectric constants of the ambient air
η = viscosity

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Chemical Processes

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Biophysical and -Chemical Aspects of Porphyrins,

ISBN 978 90 04 16210 5

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1.3.1 Spectral criterion 199

1 Partitioning and diffusion 292

Chemistry is a miracle, interest, delight,

two-electron oxidation and subsequent partial reduction of metalporphyrin was determined

aspects of electrospinning biodegradable and biocompatible natural nanofibers.

1 Synthetic and Natural

M.A. Grin and A.F. Mironov

This chapter considers natural bacteriochlorins, their properties, isolation and

Penetration into tissues, mm

Porphyrins Chlorins Bacteriochlorins

Scheme 1 Dependence of the penetration of light into tissues on the wavelength.

In the reduction of one bond (dihydroporphyrins–chlorins) or both bonds (tetrahydro-

Scheme 2 Reduction of porphyrins and their metal complexes with diimide.

A: p-MeC6H4SO2NHNH2, K2CO3/Py, t [N2H2]

Scheme 3 Reduction of 5,10,15,20-tetrakis(m-hydroxyphenyl)porphyrin with diimide.

Scheme 4 Interaction of TPP with diazoacetic-acid methyl ester.

Another method of bacteriochlorin synthesis is based on the treatment of porphyrins

Scheme 5 Interaction of octaethylporphyrin and its Zn complex with OsO4.

Pinacoline regrouping of vicinal tetrahydroxybacteriochlorins formed leads to an isomeric

A: OsO4/Py, H2S; B: H2SO4

Scheme 6 Oxidation of methyl ester of mesopyropheophorbide a 14 and trimethyl ester of meso-

Scheme 7 Oxidation of 3-formyl-3-devinylpurpurin 18.

A: OsO4/Py, H2S; B: H2SO4

Scheme 8 Effect of electron-acceptor substituents in porphyrins on the regioselectivity of OsO4

R. Pandey et al. studied the effect of electron-acceptor substituents in porphyrins and

A: OsO4/Py, H2S; B: H2SO4

Scheme 10 A Diels–Alder reaction with divinylporphyrins.

divinylporphyrins with various dienophiles, e.g., with dimethyl ester of acetylenedicarbox-

2 Natural Bacteriochlorins and Their Chemical Modifications

Scheme 11 Interaction of 8-vinylpurpurine 18 with dienophiles.

uents. Several modifications of bacteriochlorophylls are known. Thus, bacteriochlorophylls

Scheme 12 Interaction of 8-vinyl-8-deethylmesochlorin p6 with dienophiles.

Scheme 13 Intramolecular cyclization of bis(vinylformyl)porphyrins.

Bacteriochlorophyll ɚ: R1=COCH3 Bacteriochlorophyll c: R1=R3=R5=CH3

Scheme 14 Major types of natural bacteriochlorophylls.

The second group includes derivatives containing no exocyclic fragment:

Ⱥ: isopropanol; B: 1. O2/KOH, 2. HCl

Scheme 15 Production of bacteriopurpurine.

A: 0.1% HCl; B: 5% H2SO4/MeOH; C: O2 of the air; D: KOH/CH3OH; E: CH2N2; F: KOH/CH3OH, CH2N2

Scheme 16 Chemical transformations of Bchl a in acidic and alkaline media.

of chlorophyll chemistry, Nobelist R. Willstätter [25]. He introduced the term “allomeriza-

the anhydride cycle

Scheme 17 Possible chemical modifications of bacteriopurpurine.

A: C6H13NH2; method 1 – B: DCA; ɋ: DBU; method 2 – from 67 to 69: 1. CH2N2, 2. KOH/CH3OH

Scheme 18 Synthesis of cyclic imides in the chlorophyll a series.

A: C6H13NH2; method 1 – B: DCA; ɋ: DBU; method 2 – from 70 to 72: 1. CH2N2, 2. KOH/CH3OH

Scheme 19 Synthesis of cyclic imides in the bacteriochlorophyll a series.

Scheme 20 A possible mechanism of formation of 12-formyl- and 12-hydroxymethyl cycloimide

Scheme 21 Transformations of 12-hydroxymethylcycloimide of bacteriochlorin 76 in an acidic

Scheme 22 Synthesis of fluorine-containing cyclic imides of chlorin p6.

A: NH2OH.HCl, Py; B: NaOH/CH3OH; C: Ac2O; D: CH2N2, (C2H5)2O

Scheme 23 Interaction of bacteriopurpurine with hydroxylamine.

Scheme 24 Syn- and anti-isomers of oxime 86b.

Scheme 25 Synthesis and modifications of 3-vinyl-3-deacetylbacteriopurpurine 90.

Scheme 26 Synthesis of bacteriopyropheophorbide derivatives with various substituents in

A: NH2SO3H/NaClO2, THP, 2-methyl-2-butene; B: t-BuNH2*BH3, CH2Cl2;

Scheme 27 Chemical transformations of 3-formyl-3-deacetylbacteriopyropheophorbide.