Study On Secondary Metabolites Of Two Symbiotic Bacteria

Endophyte was of various species and lacked of research.During the long-term process of co-evolution with the host plant,endophyte could metabolize compounds with complex structure and outstanding biological activity.Microbes had the advantage of easily fermentation,short growth cycle and convenient production,which help to solve the problem of insufficient resources of medicinal animals and plants and possessed broad application prospect.Discovery of complicated structure and new carbon skeleton molecules and innate biological activity had made second metabolites from endophyte bearing great potential application in the field of biomedicine,which attracted much more research attention in the word gradually.Myrothecium roridum IFB-E008 was an endophyte isolated from medicinal plant Trachelospermum jasminoides by Institute of Functional Biomolecules,Nanjing University.Streptomyces rochei SR-1102 was an endophytic actinomycetes separated from the root of cucumber by Institute of Agricultural Science of Lixiahe area,Jiangsu.The chemical constituents of liquid cultures of these two endophytes were investigated in this dissertation.Fermentation broth of IFB-E008 and SR-1102 were extracted with ethyl acetate,respectively.The obtained crude extracts of these two endophytes were then fractionated by silica gel column chromatography and provided several different polarity fractions.Guided by 1H-NMR spectrum and TLC,further separation and purification was performed by silica gel column chromatography,gel filtration on Sephadex LH-20,reversed phase chromatography,HPLC and so on.Four compounds(1-4)had been obtained from M.roridum IFB-E008 and five compounds(5-9)had been provided by S.rochei SR-1102.Structures of the obtained compounds were elucidated by modern spectroscopic analysis(MS,1D and 2D NMR)and comparison with references.Aforementioned compounds were identified as 3'-iso-isororidin A(1),verrol(2),trichoverritone(3),N-acetyl-L-tryptophan(4),oxindole(5)and benzoic acid(6),in which compounds 1-3 were tricothecene compounds.Compound 1 had never been reported yet and was a new compound.Structures of 7-9 need to be further determined.In vitro cytotoxicity of compounds 2,4,5 and 6 to human gastric cancer cell lines SGC-7901 were evaluated by MTT method.The result showed that compound 2 inhibited the growth of SGC-7901 cells in a dose-dependent manner to some extent with an IC50 value of 59.79?g/mL,while IC50 value of the positive control cisplatin was 6.58?g/mL.Other compounds had no obvious activity to SGC-7901 cells.The discovery of new active natural product is the original and basic work for developing new drugs.Based on theory of endo-symbiosis,seeking for new or strong bioactive chemical constituents from second metabolites of endophyte was carried out in this dissertation,which hope to provide lead compounds for new drugs development.Tricothecene compounds obtained from liquid culture of endophytic IFB-E008 were characteristic second metabolites of M.roridum and reported to have several bioactivities.Further activity screening of obtained three tricothecene compounds would be performed in future research.