Study On Chemical Constituents And Bioactivities Of Two Deep-sea-sediment Derived Fungi And Gelsemium Elegans Benth

Objectives: Natural products is an important part of the drug,its sources include animals,plants,minerals,and microorganisms,has a rich diversity of the structure and activity,is the important resource of drug development.This article studies the secondary metabolites from two Deep-Sea-Sediment derived Fungi 3A00409 and 3A00421 and chemical composition from root and stem parts of Gelsemium elegans Benth,a series of compounds have been isolated.Using chemical methods and the modern spectrum technology such as UV,NMR,MS,to identify the structure of compounds,and in vitro screen the antitumor activity of isolated compounds,to expect new structure,and better bioactive substances.Methods:(1)Methods of isolation of Stachybotry sp.3A00409 and Graphostroma sp.3A00421: At the early stage of the laboratory study,filtered the fungal Stachybotry sp.3A00409 and 3A00421,by solid fermentation in great quantities,repeatedly extracted with methanol,the filtrate was concentrated under reduced pressure,then harvested the total extraction.And the extraction was suspended in H2 O and then partitioned with Ac OEt.The Ac OEt layer was concentrated under reduced pressure,dissolved with methanol.By the means of Thin-Layer Chromatography,Silica gel colum chromatography,Sephadex LH-20 gel,semi-preparative HPLC,and etc.to isolate the Ac OEt layer of fungal Stachybotry sp.3A00409 and Graphostroma sp.3A00421.(2)Methods of isolation of Gelsemium elegans Benth: Air-dried roots and stems of Gelsemium elegans Benth were pulverized and extracted with Me OH(1 week,3×)at r.t..The filtrate was concentrated under reduced pressure,then the total extraction had harvested,then dissolved it in acidic solution,extracted the alkaloids from acidic solution and non-alkaloids from precipitation part.Then successively partitioned with Chloroform,Ac OEt and Bu OH.Each of the extraction layer concentrated under reduced pressure,harvested the layers of extraction.Adopted the means of Silica gel colum chromatography,Sephadex LH-20 gel,MCI gel column chromatography,and semi-preparative HPLC,and etc.to isolate the Chloroform or Ac OEt layer.(3)Activity test: Select several kinds of tumor cells,apply the method of MTT to screen anti-tumor activity for the isolated compounds.Results:(1)Used the above separation and isolation methods,we separated and identified 24 compounds(B1-B24)from Stachybotry sp.3A00409,including six diterpenoids,fourteen phenylspirodrimane meroterpenoids,four fatty acids.Among them,B6 was a new diterpene,B8,B9,B10,B11 and B12 were new meroterpenoids.(2)Used the above extraction and isolation methods,we separated and identified 4 compounds(C1-C4)from Graphostroma sp.3A00421,including two coumarins,one ?-pyranone and one steroid.(3)Used the above extraction and isolation methods,we got 15 compounds from the root and stem parts of Gelsemium elegans Benth,through chemical method and UV,IR,NMR,MS and other modern spectrum technology,identified 14 compounds structure,including nine alkaloids compounds(D1-D9),five non-alkaloids compounds(R1-R5).(4)During the antitumor activity in vitro screening test of secondary metabolites from fungi Stachybotry sp.3A00409,MTT method is used to detect the inhibition rate of monomer compounds on K562(human leukemia cells),Hela(human cervical cancer cells),HL60(human promyelocytic leukemia cells),the results showed that B1,B8,B9,B10,B11,B15,B16,B20 has a good inhibition to these three kinds of cells,and other compounds had no obvious effect to these cells.(5)During the antitumor activity in vitro screening test of compounds from Gelsemium elegans Benth,MTT method is used to detect the inhibition rate of compounds on NIH3T3(mouse embryonic fibroblast),K562(human leukemia cells),Hep G2(human hepatoma cells)and A549(humanlug cancer cells),the results showed that the alkaloids had a good inhibition to Hep G2 cells,and had no obvious effect to NIH3T3,K562,A549 cells.In cytotoxic activity test of the non-alkaloids,we found that R1,R2 has a good inhibition to K562,Hep G2,A549 cells,and R3,R6 has no effect to K562,Hep G2 and A549 cells.Conclusion: This article studies the secondary metabolites from two deep-sea-sediment fungi,28 compounds were isolated and purified,among them ther 6 were new compounds.14 compounds were isolated from Gelsemium elegans Benth,9 of them are alkaloids.Alkaloids compounds is the major chemical composition of Gelsemium elegans Benth.Most of them have good inhibitory effect on the Hep G2 cells.Deep-Sea-Sediment derived Fungi was a rich source of secondary metabolites and with structural diversity.What's more,some compounds have a good antitumor activity.Therefore,further research and development are necessary.