Secondary Metabolites From Endophytic Fungi Harbored In The Medicinal Plant Mahonia Fortunei By Using OSMAC Strategy

Endophytic fungi colonize the living tissues of healthy plants without causing any negative effects and are associated with specific ecological environment.Endophytic fungi are found in almost all plants and co-exist with host plants mutually.During long-term evolution,the diversity of plants led to diverse endophytic fungi and their strong biosynthetic capabilities.Endophytic fungi can produce characteristic and diverse secondary metabolites,which already have been a vital source for active lead compounds.However,under laboratory conditions,most endophytic fungal biosynthetic gene clusters(BGCs)express little or not,resulting in many active products that cannot be found.The One Strain MAny Compounds(OSMAC)strategy mainly change the culture environment of the strain and activate the silent BGCs,which is an effective method to discover secondary metabolites.The project isolated and purified endophytic fungi from the medicinal plant Mahonia fortunei.Under the guidance of OSMAC strategy,the endophytic fungi were fermented in different conditions.After the component analyses and activity evaluations of the crude fermentation extracts,three endophytic fungi were selected for systematic chemical composition studies.Firstly,the three endophytic fungi were subjected to large-scale fermentation under the conditions selected by the OSMAC strategy.The fermentation products were extracted with ethyl acetate and separated by column chromatography techniques,such as normal phase silica gel column and Sephadex LH-20 column.They were then purified by high-pressure preparative chromatography or high-performance liquid chromatography to obtain pure compounds.The structures of compounds were identified by NMR,IR,UV and MS.Finally,the bioactivities of some compounds were evaluated.The chemical investigation on Pleospolares sp.F46 was carried out.After the endophytic fungus Pleospolares sp.F46 was fermented in PDA solid media,and the products were extracted.A total of nine compounds were isolated and identified,including two new heptaketides:pleospolarins H and I(1 and 2)and seven known compounds:(-)-balticol A(3),(-)-balticol B(4),6-deoxybostrycoidin(5),(3R,4R)-3,4,8-trihydroxy-6-methoxy-3,4-dihydro-1(2H)-naphthalenone(6),(+)-cis-and(-)-trans-3,4-di-hydro-2,4,8-trihydroxynaphthalen-1(2H)-ones(7 and 8)and(4R)-4,8-dihydroxy-atetralone(9).The endophytic fungus Pleospolares sp.F46 was statically fermented in Fungus 2~#liquid media,and its products were extracted.A total of six known compounds were isolated and identified:5?,8?-epidioxy-ergosta-6,22-diene-3?-ol(10),5-hydroxy-3-methyl-4-methoxyfuranone(11),N-(2-hydroxyphenyl)-acetamide(12),dihydro-3,4,8-trihydroxy-1(2H)-naphthalenone(13),cyclo(L-Leu-L-Pro)and(14),cyclo(Leu-Pro-Ile-Pro)(15).The Pleospolares sp.F46 was further fermented in the Fungus 2~#liquid media under shaking,and its products were extracted after fermentation.A total of one compounds were separated and identified:linoleic acid(16).The products of endophytic fungi FY10 and FY18 cultured with rice media were studied.Ethyl acetate was used to extract its products.Currently,two known compounds from the endophytic fungus FY18 were isolated and identified:2,2'-oxybis(1,4-di-tert-butylbenzene)(17)and fupyrone B(18).In summary,the subject mainly explores the secondary metabolites of endophytic fungi from the medicinal plants M.fortunei by using OSMAC strategy.The endophytic fungi Pleosporales sp.F46,FY18 and FY10 were fermented to extract their secondary metabolites.A total of 18 compounds were isolated and identified,including two new heptaketides and 16 known compounds.By using MTT method to evaluate cytotoxic activities of compounds 1-9 with A549,CT-26,MCF-7 and MDA-MB-231 cell lines,no significant cytotoxicities were observed.The results obtained in this subject suggest that the OSMAC strategy is an effective strategy to induce the expression of silent biosynthetic pathways,and it can be used to explore the biosynthetic potential of endophytic fungi to obtain novel compounds with diverse structures and activities.