Isolation And Identification Of Endophytic Fungi From Gmelina Arborea And Secondary Metabolites Of The Strain WB151

Gmelina arborea is a semi-deciduous tree belonging to the family Verbenaceae.It is a national protective plant(Rank ?)and has high medicinal and economic value.Although the chemical composition of this plant has been studied,none endophytic fungi of plants in this genus Gmelina have been reported.In this paper,we studied the endophytic fungi from Gmelina arborea.A total of 66 strains were isolated from the roots,stems,and leaves.They were identified to 13 genera,i.e.,Phomopsis,Mucor,Rhizomucor,Fusarium,Cladosporium,Trichoderma,Chaetomium,Lasiodiplodia,Aspergillus,Pestalotiopsis,Colletotrichum,Talaromyces and Amorocoelophoma.Among them,Phomopsis,Mucor,and Fusarium are the dominant groups.By chemical screening and antimicrobial screening of the isolated strains,strain WB151 was selected for further taxonomic identification and chemical studies.It was identified as Amorocoelophoma sp.in the family Amorosiaceae by analyzing DNA sequence.The genus Amorocoelophoma was established in February,2019 and there is no reports on the study of secondary metabolites from this fungal group.The large-scale broth fermentation of the strain WB151 was carried out by using PDB medium.After separation and purification by various column chromatography,a total of 16 compounds were obtained.Their structures were identified by various spectroscopic analyses,including NMR,MS,and specific optical rotation.Three new products named as amorogenin A(1),amorogenin B(5),and amorogenin C(7).The absolute configuration of the new compound 1 was determined by X-ray diffraction analysis.The other 13 known compounds were identified to be massarigenin D(2),massarilactone E(3),massarilactone G(4),7-hydroxy-3,5-dimethyl-isochromen-1-one(6),enalin A(8),massariphenone(9),?-acetylorcinol(10),2,3-dihydroxy-4-methyl-benzaldehyde(11),3-hydroxy-4-methyl-benzaldehyde(12),3,5-dihydroxy-benzaldehyde(13),2,3-dihydroxy-6-methyl-benzoic acid(14),ergosterol(15),and cerebroside B(16).Compounds 11 and 14 are new natural products.Compound 12 was isolated from microorganisms for the first time.The results showed that the secondary metabolites of the strain WB151 mainly contained lactones,isocoumarins,and phenolic derivatives.The new compound 1 and the known ones 2-3 possess an unusual 2-oxaspiro[4,5]decane-1,7-dione skeleton.Compounds 5 and 6 are belonging to isocoumarins.Compound 7-14 are phenolic derivatives.The antimicrobial activity assay showed that compound 4 has strong inhibitory activities against Salmonella typhimurium and Candida albicans.It was preliminarily resumed that the ring-opening of pentalactone in the similar structures of compounds 1-4 might improve the antimicrobial activities of these compounds.The endophytic fungi of Gmelina arborea and their antimicrobial activities were studied for the first time in this paper.Three new compounds,two new natural products and one antimicrobial active compound were obtained from the strain Amorocoelophoma sp.WB151.The results could provide a preliminary research basis for discovering more bioactive substances from endophytic fungi of Gmelina arborea in the future.