| Object:In this paper,in order to get more 3?-O-{?-L-rhamnopyranosyl-(1?3)-[?-D-xy lopyranosyl-(1?2)]-?-D- glucopyranosyl-(1?4)-[?-D-glucopyranosyl-(1?2)]-?-Larabinopyranosyl}-cyclamiretin A(refered Ag3) structural analogs,Ag3 which is a kind of triterpenoid saponins with anti-cancer activity in Ardisia gigantifolia stapf was used as the substrate and reacted by the microorganisms and the commercial enzymes.And researched into the anti-cancer activity of biotransformation products.Methods:(1)Ag3 was used as the substrate and reacted by the bacteria which were isolated, purified and identified from the stem and the rhizosphere soil of Ardisia gigantifolia stapf.The biotransformation results were detected by TLC to find the strain with biotransformation capacity,and expand the biotransformation system to get conversion products which were isolated and purified by a silica gel column;(2)Four commercial enzymes were screened for biotransformation activity on Ag3.The transformation conditions of the high active enzyme was optimized,and designed preparative experiment.The structural features of transformed products were determined by MS and NMR after purification;(3) Cytotoxicity against MCF-7?CNE?HepG-2?Hela?Bel7402and A549 of all products were investigated by CCK.The most efficient product was researched for the impact on apoptosis and cell cycle of the tumor cell by Annexin V-FITC/PI and PI.Results:It was isolated, purified and identified to obtain 39 different types of bacteria,there were 3 bacteria which had transformation activity on Ag3.They were confirmed Sphingomonas yabuuchiae GTC 868T(AB071955) from the stem ? Bacillus licheniformis ATCC 14580T(AE017333)from soil and Bacillus asahii MA001T(AB109209)from soil.Ag3 was reacted with preparative scale conversion by Sphingomonas yabuuchiae GTC 868T(AB071955).The transformed products were separated by column chromatography,and getted a compound identified as3 ?-O-{ ?-L-rhamnopyranosyl-(1?3)-[ ?-D-xylopyranosyl-(1?2)]- ?-D-glucopyranosyl-(1?4)-?-L- arabinopyranosyl}- cyclamiretin A.Through biotransformation activity screening of the four enzymes,it was be found that Ag3 could be converted into four products by Pectinex Ultra AFP.They were determinedas 3?-O-{?-D-glucopyranosyl-( 1 ? 4)-[?-D-glucopyranosyl-( 1 ? 2) ]-?-Larabinopyranoside}- cyclamiretin A ? 3?-O-{?-D-glucopyranosyl-( 1 ? 2)-?-Larabinopyranoside}-cyclamiretin A ? 3-O-?-L-arabinopyranosyl cyclamiretin A ?cyclamiretin A.5 products were evaluated for cytotoxicity against 6 kinds of tumor cells.The results showed that the inhibition on HepG-2 of 3?-O-{?-D-glucopyranosyl-( 1 ? 4)-[?-D-glucopyranosyl-( 1 ? 2) ]-?-L- arabinopyranoside}- cyclamiretin A was more significant than substrate Ag3 and positive drug DDP.The compound was researched for the impact on apoptosis and cell cycle phase of HepG-2,it was found that the administration of the apoptosis of cells HepG-2 of 7 ? mol ? L-1 group reached 21%.that verified the pro-apoptotic effects of this compound. Periodic data showed that the cycle arrest of HepG-2 occurred in S phase.Conclusion:Biotransformation method can directionally change the substrate to get a series of derivative,and there are some high pharmacologically active compounds among these analogs.In this study,it verified the anti-tumor activity of triterpenoid saponins in Ardisia gigantifolia stapf through the preliminary research of the conversion products anti-tumor activity. And speculated that the anti-tumor activity of the class composion was related with the number and type of the sugar chain.There preliminarily drawed that the activity of saponin with five sugars>three sugars>four sugars>two sugars>one sugars and a sapogenin, rhamnose and the end of glucose could respectively reduce and enhance the anti-tumor activity.Whereas,the specific structure-activity relationship is need to be furtherly investigated. |
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