Studies On Precursors During Biotransforming Process Of Increasing Diosgenin Contents Of Discorea Nipponica By Endophytic Fermentation

Dioscorea nipponica is the drying rhizome of perennial vine Dioscorea nipponica Makino which belongs to Dioscoreaceae and Dioscorea and contains abundant diosgenin.Diosgenin is one of the most important precursors to synthetize many kinds of steroid hormones and contraceptives.But with the development of the pharmaceutical industry and the increasing demand of domestic and international market for diosgenin,the resources of Dioscorea nipponica have become more failure.Now,the endophytic fungi C39 was isolated from the rhizome and leaf of D.nipponica Makino and then fermented it with herbs,which significantly improve the content of the diosgenin.To study more about the mechanism of biotransformation by C39 and find the precursors of biotransformation,based on the liquid fermentation,the steroidal saponins from Dioscorea nipponica were studied.First of all,based on the different polarity yam of chemical composition,parts of different polarity components parts(petroleum ether parts,ethyl acetate parts,n-butanol parts and water parts)were prepared and they were liquid fermented with C39.According to the different changes of diosgenin content,it is found that the precursors exist in n-butyl alcohol parts of the herbs extract in the biotransformation.Second,the liquid fermentation of n-butanol part with C39 was did and the TLC and UPLC results were analyzed.Combined with the TLC and HPLC results of the steroidal saponins from liquid fermentation product,We found that furostanol saponins was gradually reduce,but spirostanol saponins is gradually increasing.So a conclusion was drew that furostanol saponins transformed into spirostanol saponins in the process of biotransformation.Therefore,the furostanol saponins as a target of the separation was determined.Third,the technologies of macroporous adsorption resin and slica column chromatography.etc was applied to separate the possible substrate.These possible substrate was transformed by C39.The TLC and UPLC results of the biotransformation by C39 which increased the content of diosgenin in Dioscorea nipponica found and authenticated two kinds of furostanol saponins.Finally,combined with the analysis of nuclear magnetic resonance spectroscopy(13 C NMR,1H-NMR and HMBC spectrum.et al),mass s pec-trometry and chemical analysis,one of the compounds,compound 2,w-as determine the structure of 26-oxygen-beta-D-glucopyranosyl-3,26-diol-25(R or S)-Delta 5,20(22)-diene-furosta-3-oxygen-alpha-L-rhamno pyranosyl(1?2)-[alpha-L-rhamnopyranosyl(1?4)]-beta-D-glucopyranoside.Th-at is 26-O-?-D-glucopyranosyl-3,26-diol-25(R or S)-? 5,20(22)-dien e-furosta-3-O-?-L-rhamnopyranosyl(1?2)-[?-L-rhamnopyranosyl(1?4)]-?-D-glucopyranoside.The chemical structure of compound 1 remains to be f urther isolated and identificated.