REVISTA BOLIVIANA DE QUÍMICA The Bolivian Journal Of Chemistry VOLUMEN 19, No.

1 - 2002

ANTIMALARIAL ACTIVITY OF ORMOSANINE AND HOMOORMOSANINE FROM

BOWDICHIA VIRGILOIDES

José Antonio Bravo;a* Catherine Lavaud;b Geneviève Bourdy;c Eric Deharo; c Alberto Giménez;d Michel
Sauvain c
a
Laboratorio de Química de Productos Naturales, Instituto de Investigaciones Químicas-IRD, Universidad Mayor de San Andrés, CP 303, La Paz
Bolivia; bLaboratoire de Pharmacognosie UMR 6013 CNRS Bâtiment 18, BP 1039, 51097 Reims, Cedes 2, France; cInstitut de Recherche pour le
Développement (ex-ORSTOM), 213 rue Lafayette, 75480 Paris cedex 10, France; dInstituto de Investigaciones Fármaco Bioquímicas,
Universidad Mayor de San Andrés, CP 20606, La Paz, Bolivia
*Corresponding author: jbravo@red-sol.net

Key Word Index: Bowdichia virgiloides; Leguminosae; stem-bark; ormosia alkaloids; ormosanine;
homoormosanine; Plasmodium falciparum

RESUMEN American tropical forests from Venezuela to Brazil.

De la farmacopea Tacana, la corteza de tronco de
Bowdichia virgiloides H.B.K., Leguminosae, ha sido
estudiada mediante una separación extractiva y
cromatográfica bioguiada, por sus usos contra los
síntomas del paludismo. Dos alcaloides del tipo
ormosia, ormosanine y homoormosanine demostraron
una buena actividad in vitro contra cepas cloroquino
sensibles y cloroquino resistentes de Plasmodium
falciparum. Ambos compuestos en su forma sinérgica
no presentaron citotoxicidad de acuerdo a pruebas sobre
células KB. Las identificaciones estructurales fueron
establecidas mediante técnicas de espectrometría de
masas y de RMN además de un importante aporte
bibliográfico.
ABSTRACT
From the Tacana Pharmacopoeia, the stem bark of
Bowdichia virgiloides H.B.K., Leguminosae, has been
studied due to properties against malaria symptoms
applying bioguided extractive and chromatographic
techniques. Two ormosia-like type alkaloids,
ormosanine and homoormosanine showed a good in
vitro activity against chloroquine sensitive and
chloroquine resistant strains of Plasmodium falciparum.
Acceptable low cytotoxicity was evidenced from KB
cells tests. Structural identifications were carried out by
application of mass and NMR techniques besides
bibliographic research.
INTRODUCTION
Bowdichia virgiloides H. B. K.;1 is a small tree of
about 4 m high used for its pharmacological
properties in traditional medicine. The genus
Bowdichia comports four species distributed in south
In Bolivia the species B. virgiloides has been reported
in the departments of Beni and Santa Cruz.
Ethomedical uses of Bowdichia species,2, 3 include
malaria high fever relief, rheumatism as well as
dismenorrhoea and hemorrhage treatment with pain
relief.4 The Tacana people, an ethnic group of
northern western Bolivia, use stem bark of B.
virgiloides in decoction for dysentery as well as for
malaria high fever relief, also the crushed stem bark
is applied as cataplasms against wounds of
Leishmaniasis cutaneous form.5 Some biological
activities for Bowdichia species extracts have been
reported, mainly dermatitis provoked by topical
application of the ether extract of B. nitida,6
probably due to the presence of 2,6-dimethoxy-p-
benzoquinone causing allergy on Guinea’s pig skin.7
This compound is also thought to be the responsible
of the resistance of Bowdichia woods to termites. B.
virgiloides has been previously tested against some
bacteria and fungi8 namely Eschericha coli, Bacillus
subtilis, Staphylococcus aureus and Streptococcus
faecalis, Neurospora crassa, Mycobacterium
smegmatis and Candida albicans, showing no
activity. Two species, B. nitida and B. virgiloides
have been surveyed from a chemical stand point.
Flavonoids9 bowdichione, centisteine, 3’-
hydroxyformononetin and homopterocarpin besides
2,6-dimethoxy-p-benzoquinone7, 9 have been
described in B. nitida. The existence of triterpenes
like betulinic acid, betulinol10 and lupeol11 and of
alkaloids homoormosanine, homo-18-epiormosanine,
piptanthine, homopiptanthine, podopetaline11 have
been established in B. virgiloides. The
pharmacological properties described above talk
about certain specificity in active principles of B.
virgiloides, fact that incited us looking for
antimalarial activities in the plant.

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REVISTA BOLIVIANA DE QUÍMICA The Bolivian Journal Of Chemistry VOLUMEN 19, No.1 - 2002

RESULTS AND DISCUSSION Antimalarial tests results with extracts, fractions and
pure compounds from Bowdichia virgiloides
The in vitro assays against P. falciparum of the Fraction % inhib. ¶ % inhib. †
[µg/ml]
hydroethanolic (3:7) extract of stem bark of B. EtOH 1 100 100
virgiloides exhibited an IC50 = 1.0 µg/ml. Dried and TA 1 100 100
powdered stem bark were extracted by a classical acid- 5 (VLC)* 1 100 100
base method12 to determine the total alkaloids content IC50 = 0.5 IC50 = 0.5
(µg/ml) (µg/ml)
(TA). The TA extract showed and antiplasmodial 6 (VLC)* 1 100 100
activity of CI50 < 1,0 µg/ml demonstrating that the IC50 = 0.5 IC50 = 0.5
initial activity of the hydroethanolic extract (µg/ml) (µg/ml)
corresponded to the ormosia-like type alkaloids 7 (VLC)* 1 100 100
IC50 = 0.5 IC50 = 0.5
previously reported in the species.11 The TA extract was
(µg/ml) (µg/ml)
submitted for separation to a VLC system on silica gel 33-48 (LC)§ 20 45 20
employing binary mixtures of CH2Cl2-MeOH of 2 18 9
increasing eluting power. From the sixteen fractions 0.2 0 7
obtained, activity concentrated on fractions 5, 6 and 7 ( IC50 > 20 IC50 > 20
(µg/ml) (µg/ml)
100% of inhibition of parasitaemia at 1.0 µg/ml). 1 (LC) § 20 50 43
Fractions 5, 6 and 7 revealed by TLC to contain the 2 27 0
same three compounds (comparable Rf’s) and being 0.2 0 27
gathered were treated in an open column LC system on IC50 > 20 IC50 > 20
silica gel. The elution system to try to separate them was (µg/ml) (µg/ml)
2 (LC) § 20 89 89
AcOEt-MeOH in mixtures of increasing polarity. From 2 23 32
262 total fractions, three alkaloids were isolated, TLC 0.2 0 0
analysis put in evidence that fractions 13-48 contained IC50 = 5 (µg/ml) IC50 = 5 (µg/ml)
the less polar compound, fractions 55-238 a more polar *VLC: Vacuum Liquid Chromat. Silica 60H
§LC: Liquid Chromat. In open column Silica 60
second alkaloid and fractions 248-262 the most polar of ¶: Chloroquine sensitive strain, radio isotopic method
†: Chloroquine resistant strain, radio isotopic method
the three of them. Spectroscopic analysis and literature A next step in this pathway should conduct us to the
data permitted to confirm the purity of compounds and identification of the third less polar alkaloid of the
to assign structural identification to alkaloids the more ternary mixture, to determine all the components of
polar in the tripartite mixture, namely homoormosanine such a synergic composition. An HPLC analysis would
1, (fr. 55-238), and ormosanine 2, (fr. 248-262). The provide us with the percentage composition of the
structure of the third alkaloid is unconfirmed by now. synergic mixture. Anyway, the good activity found for
The activity manifested by fractions and extracts in this
the synergic mixture, IC50 = 0.5 (µg/ml), still remains
biologically guided pathway is resumed in the table
lower than that observed for the reference molecule,
below.
chloroquine. The three alkaloids and the synergic
H
mixture, have been separately evaluated for their
H
toxicity levels through the test over human KB
N
H
N carcinoma cells and Hela. Only no toxicity was found
H H
H for 5-7 VLC frs. according to KB: EC50 = 25 µg/ml
H
H
and Hela: EC50 = 25 µg/ml. From a chromatographic
N N NH HN analysis stand point and following literature data,11 we
Compound 1 Compound 2 can speculate that the third and less polar alkaloid (frs.
Neither compound 1 nor compound contained in 13-48) in the synergic mixture should be either
fractions 13-48, manifested antiplasmodial activity like homopiptanthine or homo-18-epiormosanine, because
that showed by compound 2. Regarding the quest of the they are among the three less polar alkaloids in B.
antimalarial activity manifested by B. virgiloides, we virgiloides. The other two alkaloids in B. virgiloides,
can conclude that the most active fractions have been pipthantine and (-) podopetaline, are more polar than
obtained from the TA active extract. Only pure ormosanine and should be eventually gathered in VLC
ormosanine (2), possesses an important antimalarial fractions 8-16. An increasing polarity order for the six
activity, however this activity appears weaker (IC50 5 described alkaloids in B. virgiloides, has been
µg/ml) than the activity observed in assays with the ,
reported:2, 3 10, 11 homo-18-epiormosanine (Rf 0.73),
most active fractions from the TA extract, namely VLC homopiptanthine (Rf 0.69), homoormosanine ( Rf
frs. 5, 6 and 7 (100% of inhibition of parasitaemia at 0.60), ormosanine (Rf 0.39), piptantine (Rf 0.39).
µg/ml). These fractions represent a mixture of three According to TLC analysis made by us under the same
alkaloids (1, 2 and frs. 13-48 from LC). experimental conditions than those reported in the

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 REVISTA BOLIVIANA DE QUÍMICA The Bolivian Journal Of Chemistry VOLUMEN 19, No.1 - 2002

literature over the TA extract, the six alkaloids have

been identified based on a comparison of experimental H H H

and reported Rf values. Considering these results we N N N

H H H
can deduce that the synergic mixture is composed by H H

homopiptanthine (frs. 13-48, for which a single H H H

revealed spot appears on TLC with a purity stated by NH HN NH HN NH HN

the 20 discrete peaks on the 13CNMR spectrum), ormosanine piptanthine (-)-podopetaline

compound 1, homoormosanine and compound 2,
ormosanine. The structural identification confirmation for 1 and 2,
came out from NMR and EIMS analysis. Alkaloids 1
H H H
and 2, were identified after comparing 13CNMR
N N
experimental signals with literature as
N
H H H homoormosanine 3 and ormosanine13, respectively (see
H H H
18 table below).
H H H
N N N N N N

homo-18-epiormosanine homopiptanthine homoormosanine

13
CNMR data for compounds 1, 2 and literature data for homoormosanine and ormosanine

δ δ δ δ
Atom Homoormosanine* Compound 1† Ormosanine¶ Compound 2§ ∆‡
2 55.5 55.5 55.2 53.4 +1.8
3 25.6 25.3 26.8 25.1 +1.7
4 25.0 24.6 25.3 24.6 +0.7
5 32.1 31.9 32.0 30.0 +2.0
6 64.9 65.3 63.8 62.2 +1.6
7 34.5 34.9 35.4 33.8 +1.6
8 30.0 29.5 30.7 28.6 +2.1
9 36.5 36.4 39.3 38.3 +1.0
10 63.3 62.4 56.1 54.2 +1.9
11 66.7 66.2 66.1 65.0 +1.1
13 53.2 53.1 47.0 45.4 +1.6
14 25.5 24.9 26.6 24.6 +2.0
15 - 33.5 33.2 31.6 +1.6
16 31.6 31.7 34.1 33.4 +0.7
17 37.9 37.5 38.0 37.1 +0.9
18 68.2 67.8 66.6 65.6 +1.0
19 19.8 19.7 26.8 24.6 +2.2
20 16.5 16.4 24.0 21.4 +2.6
21 25.0 24.7 25.6 23.9 +1.7
22 53.4 53.2 48.1 46.1 +2.0
24 69.8 69.7
*In CDCl3 at 22.5 MHz, δ in ppm from TMS; † In CDCl3 at 75 MHz, δ in ppm from CHCl3 (77 ppm); ¶ In CDCl3
at 50 MHz, δ in ppm from TMS; § In CDCl3-CD3OD in an equimolar proportion in deuterium, at 75 MHz, δ in
ppm from CHCl3 (77 ppm);‡ Positiv value of the difference between δ values of ormosanine and 2 due to the
mixture CDCl3-CD3OD used with 2 instead of pure CDCl3 reported in literature data for ormosanine

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REVISTA BOLIVIANA DE QUÍMICA The Bolivian Journal Of Chemistry VOLUMEN 19, No.1 - 2002

4 3
4 3
2
2
4
4
5 3 5
H H 6 3 5
N1 H
5 H 6
6 H N1
2 10 6 H
7 14 2 10
N 15 16 7
1 17 N 14 15
17 8 1 16
H 7 8 17
H 10 N 17 H 7
13 9 10 N
9 19 12 H 13 9
18 11 8 9 19 12
N 18 11 8
15 16 11 20 24 23 H 23N
H 15 16 11 20 H H
H 18 HH
22 H 18
N N 22
14 21 NH HN
12 23 14 21
13 24 22 19 12 23
13 22 19
21 21
20 20

Compound 1 : homoormosanine Compound 2 : ormosanine

EIMS spectral data obtained for 1 and 2 permits to General- NMR spectra were run in a AC 300
corroborate the structural assignments according to BRUKER spectrometer. Mass spectra acquired under
literature data for homoormosanine3 and ormosanine.14 electronic impact at 70 eV, Optical activity
This survey of Bowdichia virgiloides conducted us to the measurements were measured on a PERKIN ELMER
qualitative characterization of a three-alkaloid-sinergic- 241. VLC Silica gel : 60H MERCK (5-40 µm). LC
mixture, namely homopiptanthine, homoormosanine and open column silicagel :MERCK 60, 0.063-0.200 mm
ormosanine with an IC50 = 0.5 µg/ml over Plasmodium (70-230 mesh ASTM). TLC plates: WHATMAN (250
falciparum in vitro. The mixture represents the and 500 µm, PK 6F, gel 60A).
antiplasmodial fraction coming up from the TA extract. Plant material - B. virgiloides H.B.K., Leguminosae
Also the antiplasmodial in vitro activity indexes for each was collected during ethnobotanical field work in the
of the three alkaloids have been established. A Tacana ethnic group territories, Apolo province of the
quantitative analysis of the synergic mixture should be La Paz department at 600 meters over sea level. A
carried out in another research in order to define the voucher specimen is deposited at the National
proportional composition of it. The fact of the Herbarium of the San Andrés Major University of La
disappearance of the methylene group at C-24 forming an Paz Bolivia under the code GB 1734.
open space for a double pole for interactions between N- Extraction and isolation of compounds – Obtention of
12 and N-23 and any substrate in compound 2, could be the TA extract: Dried and powdered stem-bark (280.1
the reason for the activity loss in compound 1 which g) was moisturized in a NH4OH (NH3-H2O, 1:1, 10 ml)
possesses in contrast such a 24-methylene group. solution. Alkaloids salt pass to their CH2Cl2 soluble
free base form. Alkalinized stem bark is extracted
EXPERIMENTAL during 24 hr. under CH2Cl2 in a Soxhlet apparatus. The
organic layer is extracted with a series of acidic
aqueous fractions (5% v/v de HCl, 100 ml each).
Alkaloids pass from their free base form to water
soluble salts. The operation is repeated until the
transfer of alkaloids from the organic layer is
exhausted. The aqueous layer is alkalinized under
NH4OH (NH3-H2O, 1:1, 10 ml) until pH 9 to get free
alkaloids. The aqueous layer is extracted under various
fractions of CH2Cl2, 100 ml each, until the transfer of
alkaloids from the aqueous layer is exhausted. The
organic layer is dried with anhydrous sodium sulfate.
Solvent evaporation in vacuo affords the TA extract
(4.64g, 1.7 % of stem bark). Alkaloids isolation: 2 g of
TA extract were chromatograped in a VLC system
(silica gel 60 H until 5 cm high in a fritted base funnel
of 7 cm of internal diameter) eluted with mixtures of
CH2Cl2-MeOH of increasing polarity, to afford 16
fractions: [100:0 (200 ml), 4.3 mg; 95:5 (200 ml), 1
mg; 90:10 (200 ml)X3, 346.9 mg, 596 mg et 84.8 mg;
87:13 (200 ml)X2, 30.4 mg and 29.6 mg; 85:5 (200
ml)X2, 14.7 mg and 16.2 mg; 83:17 (200 ml)X2, 14.6
Bowdichia virgiloides H.B.K. (Leguminosae). mg and 23.7 mg; 80:20 (200 ml)X3, 43.4 mg, 51.2 mg
Illustration by C. Maldonado, LPB. and 64.4 mg; 0:100 (200 ml)X2, 100.5 mg and 280.0

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 REVISTA BOLIVIANA DE QUÍMICA The Bolivian Journal Of Chemistry
mg]. Fractions 5, 6 and 7 [90:10 (200 ml), 84.8 mg,
87:13 (200 ml)X2, 30.4 mg and 29.6 mg] that
manifested the best activity indexes, and after
corroborating the same chemical composition in
alkaloids through TLC analysis (SiO2, CH2Cl2 and
NH3 vapor, revealing agent: Dragendorff reagent),
were gathered (166 mg) and then separated in a LC
open column on silica gel 60 (6.6 g) using AcOEt-
MeOH mixtures. 262 fractions were obtained: [98:2
(2.5 l); 97:3 (0.08 l); 96:4 (0.15 l); 95:5 (0.12 l); 94:6
(0.09 l); 93:7 (0.01 l); 92:8 (0.14 l); 90:10 (0.12 l);
85:15 (0.12 l); 80:20 (0.32 l), 0:100 (0.05 l)]. Fractions
13-48 afforded one pure alkaloid according to TLC
and NMR analysis, AcOEt-MeOH (98:2). Fractions
55-238 afforded compound 1 AcOEt-MeOH (98:2,
97:3, 96:4, 95:5, 94:6, 93:7, 92:8, 90:10, 85:15).
Fractions 248-262 afforded compound 2 AcOEt-
MeOH (80:20, 0:100).
Compound 1. Homoormosanine. [α]D -5.7° (CHCl3-
MeOH, 1.0:0.2 c 0.30); EIMS 70 eV: m/z (rel. int.): 329
[M]+ (100), 247 (25), 246 (80), 233 (23), 232 (18), 231
(62), 217 (13), 205 (16), 164 (12), 163 (25), 162 (22),
150 (27), 149 (25), 148 (26), 136 (17), 134 (19), 123
(12), 122 (13), 110 (18); 1HNMR (300 MHz): 3.06 (1H,
H-1), 2.12 (1H, H-1’), 1.86 (1H, H-3), 1.61 (1H, H-3’),
1.76 (1H, H-5), 1.01 (1H, H-5’), 1.93 (1H, H-6), 1.68
(1H, H-7), 1.81 (1H, H-8), 0.94 (1H, H-8’), 2.91 (1H, H-
10), 2.70 (1H, H-10’), 2.80 (1H, H-11), 2.78 (1H, H-13),
1.95 (1H, H-13’), 1.56 (1H, H-15), 1.58 (1H, H-16),
1.41 (1H, H-17), 1.10 (1H, H-17’), 1.49 (1H, H-18),
1.10 (1H, H-19), 0.85 (1H, H-19’), 1.80 (1H, H-20),
1.18 (1H, H-20’), 2.93 (1H, H-22), 3.40 (1H, H-24),
3.24 (1H, H-24’); 13CNMR (75.0 MHz): 55.5 (t, C-2),
25.3 (t, C-3), 24.6 (t, C-4), 31.9 (t, C-5), 65.3 (d, C-6),
34.9 (d, C-7), 29.5 (t, C-8), 36.4 (s, C-9), 62.4 (t, C-10),
66.2 (s, C-11), 53.1 (t, C-13), 24.9 (t, C-14), 33.5 (t, C-
15), 31.7 (d, C-16), 37.5 (t, C-17), 67.8 (d, C-18), 19.7
(t, C-19), 16.4 (t, C-20), 24.7 (t, C-21), 53.2 (t, C-22),
69.7 (t, C-24).
Compound 2. Ormosanine. [α]D -9.1° (CHCl3-MeOH,
1.0:0.2 c 0.28); EIMS 70 eV: m/z (rel. int.): 317 [M]+
(65), 259 (7), 246 (15), 234 (43), 233 (34), 232 (34),
231 (19), 219 (100), 205 (7), 191 (13), 176 (8), 162
16
VOLUMEN 19, No.1 - 2002
(14), 151 (44), 150 (20), 149 (91), 134 (27) 122 (16),
110 (22); 13CNMR (75.0 MHz): 53.4 (t, C-2), 25.1 (t,
C-3), 24.6 (t, C-4), 30.0 (t, C-5), 62.2 (d, C-6), 33.8 (d,
C-7), 28.6 (t, C-8), 38.3 (s, C-9), 54.2 (t, C-10), 65.0
(s, C-11), 45.4 (t, C-13), 24.6 (t, C-14), 31.6 (t, C-15),
33.4 (d, C-16), 37.1 (t, C-17), 65.6 (d, C-18), 24.6 (t,
C-19), 21.4 (t, C-20), 23.9 (t, C-21), 46.1 (t, C-22),
69.7 (t, C-24).
Antimalarial in vitro assays. The culture of
chloroquine resistant (Indo) and chloroquine sensitive
(F32-Tanzania) Plasmodium falciparum strains is
realized according to Trager and Jensen15 in a RPMI
1640 medium enriched by glucose and completed by a
10% of human serum at 37ºC. 50 µl of DMSO are
added to samples of vegetal extracts solved in RPMI
1640 medium using an ultrasound box. Final
concentration of DMSO never exceeds 0.1%. 150 µl of
culture medium containing the dilute extract and a
human hematies (“O” + group, 5% of hematocrite)
with 1% of parasitaemia, all placed in 96-well-plates
for micro titration. All test were executes by triplicate.
After a 24-hours-incubation period at 37°C in an
incubator containing a candle, the medium is renewed
and incubation continues during still 48 more hours. At
the third day a sample of blood is extracted from each
well to measure the parasitaemia. Each assay is
correlated to a solvent untreated witness and
chloroquine positive witness. The parasitaemia of each
receptacle as well as the inhibition percentage for each
extract concentration are calculated regarding the
untreated witness. The values of the IC50 were
determined graphically taking into account the
concentration versus the percentage.
ACKNOWLEDGMENTS
The authors wish to thank the National Herbarium of
Bolivia. The ethnic group Tacana. The FONAMA for
financial support. Miss Lic. Yvon Rojas for
antimalarial ant cytotoxic assays.
REVISTA BOLIVIANA DE QUÍMICA The Bolivian Journal Of Chemistry VOLUMEN 19, No.1 - 2002

REFERENCES

1
BRANDAO, M. G. L., GRANDI, T. S. M., ROCHA, E. M. M., SAWYER, D. R. KRETTLI, A. U.
J. of Ethnopharmacology, 1992, 36, 175.
2
ACHENBACH, H., BAUEREIβ, P., TORRENEGRA, R.
Arch. Pharm. Weinheim Ger., 1988, 321, 675.
3
TORRENEGRA, R. G., ESCARRIA, S. R., BAUEREIβ, P., ACHENBACH, H.
Planta Medica, 1985, 3, 276.
4
ELISABETSKY, E., POSEY, D. A.
J. of Ethnopharmacology, 1989, 26, 299.
5
BOURDY, G.
Personal communication
IRD-UMSA, Mission La Paz, Bolivia.
6
GONÇALO, S.
Contact Dermatitis, 1992, 26, 205.
7
SCHULTZ, K. H., GARBE, I., HAUSEN, B. M., SIMATUPANG, M. H.
Arch. Dermatol. Res., 1979, 264, 275.
8
CHIAPPETA, A. D. A., FRANCISCO DE MELO, J., MACIEL, G. M.
Rev. Inst. Antibiot. Univ. Fed. Pernambuco Recife, 1983, 21, 43.
9
BROWN, P. M., THOMSON, R. H., HAUSEN, B. M.
Justus Liebigs Ann. Chem., 1974, 1295
10
MARINHO, L. C., CARNEIRO DA CUNHA, M. T. M., THOMAS, G., BARBOSA, FILHO, J. M.
Fitoterapia, 1994, LXV, 475.
11
TORRENEGRA, R., BAUEREIβ, P., ACHENBACH, H.
Phytochemistry, 1989, 28, 2219.
12
BRAVO, J. A., SAUVAIN, M., BALDERRAMA, L., MORETTI, C., RICHOMME. P. BRUNETON, J.
Rev. Bol. de Quim., 1996, 13, 19
13
BHACCA, N. S., BALANDRIN, M. F., KINGHORN, A. D., FRENKIEL, T. A., FREEMAN, R.,
MORRIS, G. A.
J. Am. Chem. Soc. 1983, 105, 2538
14
KINGHORN, D. A., BALANDRIN, M. F., LIN, L-J
Phytochemistry, 1982, 21, 2269. (14)
15
TRAGER W, JENSEN J B
Human malaria in continous culture.
Science, 1976, 193, 673-675.

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