Study On The Metabolites Of Four Mangrove Endophytic Fungi

Marine mangrove endophytic fungi is a kind of special microorganism.The environment with high salt,high temperature,and low oxygen make these microorganisms produce their own unique metabolic pathways.Among them,Aspergillus fungi as one of the main dominant strains,which can produce a large number of novel and active metabolites.It has a positive role for the development of medicine.In this paper,the chemical constituents of the secondary metabolites of four endophytic fungi Fusarium sp.R5,Aspergillus clavatus R7,Trichoderma sp.09,Fusarium sp.L5 from marine mangrove plants were studied.The fermentation broth were extracted by ethyl acetate and n-butanol,respectively,and totally 31 compounds were isolated and purified by using normal phase silica gel column chromatography,gel column chromatography,thin layer chromatography and preparation liquid chromatography techniques.The structures were identified mainly by analysis of the nuclear magnetic resonance,high resolution mass,infrared and ultraviolet spectra,as well as by comparison of spectral data of with literature precedents.The results were as follows.Totally 10 compounds were isolated from endophytic fungi Fusarium sp.R5: 5-hydroxymethyl-2-furaldehyde(1),(3R,4R)-cis-4-hydroxymellein(2),(3R,4R)-cis-4,7 dihydroxymellein(3),flazine(4),?-sitosterol(5),3?-cholest-5-en-3-ol(6),butanedioic acid(7),maleic acid(8),ergosta-4,6,8(14),22-tetraen-3-one(9),clavatol(10);13 compounds were isolated from endophytic fungi Aspergillus clavatus R7: 24-hydroxyergosta-4,6,8(14),22-tetraen-3-one(11),4-(?,?-dimethyl allyl oxygen radicals)-9,10-dimethyl-2-hydroxy acetophenone(12),4,4?-dimethoxy-5,5?-dimethyl-7,7?-oxydicoumarin(13),7-hydroxy-4-methoxy-5-methylcoumarin(14),7-((?,?-dimethyl allyl oxygen radicals)-4-methoxy-5-methylcoumarin(15),kotanin(16),orlandin(17),clavatustides A(18),clavatustides B(19),thymine(20),xanthine(21),allantoin(22),L-arabinitol(23);4 compounds were isolated from endophytic fungus Trichoderma Sp.09 : dichlorodiaportinolide(24),dichlorodiaportin(25),diaportinol(26),diaporthin(27).4 compounds were isolated from endophytic fungi Fusarium sp.L5: 2-hydroxy-3-phenyl-1-cyclopentene-2-ketone(28),(S)-5-hydroxy-2,6-dimethyl-4H-furo[3,4-g]benzopyran-4,8(6H)-dione(29),kojic acid(30),lumichrome(31).Compounds 11,12,13,15,24,and 29 were new compounds which have not been reported in the literature,and a new natural product 28 was found.Among the known compounds,compounds 14,20,21,and 22 were isolated from Aspergillus sp.for the first time,and compound 2,3,4,10,and 31 was isolated from the Fusarium sp.for the first time.The antibacterial activity,antioxidant activity and cytotoxic activity of some selected isolated compounds were studied.Disc-agar diffusion assay of antibacterial activity experiment showed that at the concentration of 250 ?g/mL,compound 4 presented high antibacterial activity against Colletotrichum musae(Berk.&M.A.Curtis)Arx.(C.musae)and moderate antibacterial activity against Fusarium oxysporum Schlecht.f.sp.lycopersici(Sacc.)W.C.Snyder et H.N.Hansen(F.oxysporum)and Fusarium graminearum Schw.(F.graminearum).Broth dilution antifungal experiment indicated that compounds 11 possessed high antibacterial activity(MIC? 50 ?g/m L)against Fusarium oxysporum,Colletotrichum musae and Penicillium italicum Wehme(P.italicm),and compound 10,17 had moderate antibacterial activity against Fusarium oxysporum;Antibacterial experiment showed that compounds 9,10 had high antimicrobial activity against escherichia coli(MIC value = 20 ?g/m L,35 ?g/mL,respectively).DPPH radical scavenging assay showed that compounds 9 and 11 had weak antioxidant activity.MTT cytotoxicity assay showed that compounds 9 and 11 had moderate inhibitory activity on the growth of hepatoma cells(HepG2),ovarian cancer cells(OVCAR-3)and breast cancer cells(MDA-MB-231),and compound 12 had moderate inhibitory activity on the growth of hepatoma cells(HepG2).The cytotoxic activities of the compounds 9,11,12,13,14,15,17 for these three cancer cells had not been reported before.In this study,the metabolites of four mangrove endophytic fungi were analyzed and some compounds showed significant inhibitory activity to plant pathogens and human bacteria,which promoted the research and development of marine mangrove microbial metabolites.