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2684 Short Reports

vnth the base peak at m/z 43 The ‘HNMR spectrum was DIrector, Herbarlo Barhosa Rodrlgues de ItaJal, Santa Catarma,
identical with the spectrum of neo-mosltol hexaacetate (2a) Braul, for the ldentdicatlon and collection of the plant mate&
(Calc for C,sH,,O,, C, 5000, H, 5 55, Found C, 5004, H, RM thanks CNPq, Brazil, for financial assistance
Acknowledgement-The authors thank Professors B Das,
Institut de Chlmle des Substances Naturelles, 91190 Glf-sur- 1 Angyal, S J and Matheson, N K (1955) J Am Chem Sot 77,
Yvette, France, and N S Bhacca, Louisiana State Umverslty, 4343
Baton Rouge, LouIslana, U S A, for mass and NMR spectra 2 Lshtenthaler, F W and Enug, P (1968) Carbohydr Res 7,
Thanks are also accorded to Professors S J Angyal, University of 121
New South Wales, Australia, for the samples of ( - )-L-bornesitol 3 Uneo, Y , Hasegawa, A and Tsuclnya, T (1973) Carbohydr
pentaacetate and neo-mositol hexaacetate and Roberto M Khen, Res 29,520

Phyfochemwry, Vol 23, No 1I, pp 2684-2685, 1984 0031-9422/84 $3 00 + 0 00

Pnnted m Great Bntam Q 1984 PergamoaPress Ltd



Botany Department, School of Life Science, Visva-Bharati Umverslty, Santmrketan-731235, India

(Reused received 26 Aprrl 1984)

Key Word Index-Dtoscorea composlta, Dloscoreaceae, chemoddferentlation, dlosgenm, tissue culture

Abstract-Dlosgemn was isolated from different parts of a three-year-old plant of Dloscorea compostta The amounts
(% on a dry wt basis) present were tubers, 3 6, vme Internodes and nodes with their leaves from first 20 nodes from the
tubers, 16, smnlarly from mtermechate 20 nodes, 0 039 and from upper 20 nodes, 0 03 The amounts ( y. on a dry wt
bask) from tissue culture of nodal explants were 30-day-old callus, 0 89,9O-day-old callus, 161, emergent shoots, 2 5,
regenerated roots, 0 08

INTRODUCTION and It has been reported that dlosgenm synthesis 1sgreater

in unorgamsed tissue culture than m orgamsed root
Dlosgenm 1s used as a starting material for the manufac-
culture of D deltouiea [S, 63 In the present study,
ture of steroid drugs, mcludmg cortlcosterolds and oral
however, it was found that dlosgenm synthesis IS greatest
contraceptives It 1s present m different amounts m the
in orgamsed shoot cultures of D composite, a finding
same plant growmg m different localities [l, 21, but little 1s which 1s reflected m the m SW production pattern of the
known regardmg Its levels m different parts of the plant
plant Perhaps, chemodlfferentlatlon of dlosgenm 1s m-
and Its synthesis m relation to organogenesls m culture
fluenced by organogenesls A similar response has heen
reported m the case of cardenohde blosynthesls m
RESULTS AND DISCUSSION Calotropts gzgantea [7]
In three-year-old plants harvested prtor to flowering EXPERIMENTAL
the upper young 20 nodes with leaves contained 0 030 %
dlosgenm on a dry weight basis, whereas the nodes (20) Plant materials (aenal part, tubers) were collected from the
adjacent to the tuberscontamed 16 %, the intermediate 20 University garden Three-year-old plants were divided into four
nodes contained 0 039 % and the tubers contained 3 6 % parts wz the internodes and leaves of the upper 20 nodes,
dlosgenm. The side shoots arising at the nodes were not sundarly of the 20 nodes adJacent to the tuber and likewise the 20
included m these analyses Young callus (30-day-old) mtermedlate nodes, and tubers Nodal explants of healthy
obtamed from MS1 medium contained 089% and the growing plants were cultured m RT (revised tobacco medium),
h@est content, 161”/, was observed m 90&y-old cul- supplemented with 2,4-D (2 mg/l) and Kn (0 5 me/l) for callus
ture The content was further enhanced to 2 52 % in the uutiation and organogenesls Diosgemn was also determined
emergent shoots derived from callus cultures However, from the mltiated callus (l-month-old), old callus @O-days-old),
regenerated roots contamed less dosgenm (0 08 %) the emergent shoots denved from callus, proliferated roots m
Studies with hssue cultures of D composrta have shown culture and regenerated plants m culture
that supplementation of the growth medium with 0 5 mg Extractaon procedure ofdlosgencn Dried finely powdered plant
benzyladenme stimulates dlosgenm biosynthesis [3, 41, matenal(0 5 kg m the case of tubers and aerial parts, 10 g m the
Short Reports 2685

case of callus tissues and regenerated shoots and roots for each HPLC analysis, the Indian Institute of Chenucal Biology,
phase of growth) was hydrolysed with 2 5 1 of HCI (5 %) under Jadavpur for IR spectra, the CSIR, New Delhi for financial
reflux for 6 hr, the volume of the mixture was mamtamed assistance, the CIMAP, Lucknow and ICAR, New Delhi for
constant by adding H,O from time to time After coohng, the plant matenal
nuxture was filtered and the residue washed thoroughly with
H,O (to free from acld)and then dned at 60” The dried mass was
then extracted m a Soxhlet apparatus wth n-hexane for 10 hr,
after which the extract was &t&d (to remove solvent) on a Cruzado, H J , Delpm, H and Roark, B A (1965) Tumalba
waterbath and then chromatographed over alummmm oxide 15,25
(neutral grade I, 20 g of Al,O, per g extract) The fractions were Coursey, D G (1967) m Yams, p 17 Longman, London
momtored on TLC for chosgemn A fraction of n-hexane- Heble, M R and Staba, E J (1980) Planta Med (suppl) 120
benzene (1 1) eluates showed the presence of only dlosgemn Datta, S K , Datta, K and Datta, P C (1982)m Tissue Culture
(compared by co-TLC wth reference diosgemn) The residue of Econonncally Important Plants (Rao, A N , ed ) pp 89-93
from these eluates were combined and crystalhzed from Me&O Singapore
Quantitation of dlosgemn from callus and regenerated shoots Kaul, B and Stabas, E J (1968) Lloydlo 31, 171
and roots m culture was performed by IR and HPLC analysis [8] Marshal, J G and Stabas, E J (1976) Phytochetmstry 15,53
Datta, S K and De, S (1983) Cell Chrom Res Bull 6, 10
Acknowledgements-We urlsh to express our smcere thanks to Mahato, S B , Sahu, N P and Roy, S K (1981)5 Chromatogr
Dr S K &mXJe, Orgamc Chenustry Lab, RRL, Jammu for 206,169

Phytochemrrrry,Vol 23, No 11, pp 2685-2686, 1984 0031-9422/84$3 00 +0 00

Printed m Great Bntarn Q 1984Pergamon Press Ltd



Istltuto di Chlrnica Orgamca e Blologlca, Umversld cl] Napoh, Via Mezzocannone 16, 80134 Napoh, Italy, l Istltuto Superlore dl
Samt& Vlale Regma Elena 299,00161 Roma, Italy

(Recerved 21 February 1984)

Key Word Index-Candrda bpolytrca, Ascomycetes, fung, sterols

Abstract-The sterols of Candlda lzpolytlcagrown on n-alkanes were isolated by reverse phase HPLC and found to be
mamly ergosterol, with small quantities of ergost-7-en-3b-01, ergosta-7,22-dlen-3fl-01, ergosta-7,24(28)-dlen-3B-ol and
ergosta-5,7,9( 11),22-tetraen-38-01

Recently much attention has been given to the yeasts acetate The sterols were identified on the basis of their
Candrda tropzah and Car&da hpolytrca, which, being mass, UV and ‘H NMR spectra The percent composltlon
capable of utdlvng ahphatlc hydrocarbons as the sole of the sterols m C lrpolytlca and the chromatographlc
carbon source, attracted commercial interest for produc- moblhty data are summarized m Table 1 A previous
tion of mlcroblal proteins which may be utlhzed as investigation of the sterol nuxture of C ltpolytrca grown
components m animal feeds [l] In contmumg our work on n-alkanes revealed the presence of ergosterol [4]
on sterols from fungi [2,3], we exammed the sterol
composltlon of C llpolytlca grown on nalkanes
The residue from the chloroform-methanol extract of
C llpolytlca upon saponrficatlon followed by column C bpolytrca was grown on n-alkanes by the mdustnal process
chromatography, gave the 4-demethylsterol mixture, of Italproteme HPLC was on a Waters Instrument equipped
whrch was acetylated The prehmmary GC of the stecyl with a dlfferentml refractometer and Whatman PartIs 5/25
acetates showed four small peaks beslde the major peak ODS-3 column (4 6 mm x 25 cm), ‘H NMR, 270 MHz, CDCI,,
identical with that of standard ergosteryl acetate The TMS as internal standard, UV. MeOH, GC, DB-1 fused slhca
sterol acetate rmxture, subjected to reverse phase HPLC, capdlary column (30 m x 0 25 mm) at 265”, MS, 70 eV
yielded ergosta-5,7,9( 11),22-tetraen-3/3-yl acetate, ergo- Extractton and separation of sterols C bpolytlca (58 g) was
steryl acetate, ergosta-7,24(28)-dlen-3/l-yl acetate, extracted x 3 at room temp with CHC13-MeOH (1 1) The
ergosta-7,22-dlen-3Byl acetate and ergost-7-en-3/?-yl solvents were evapd to Blve a vmcous od (8 4 g), wluch was