Study On The Chemical Constituents Of Three Endophytic Fungi From Anoectochilus Roxburghii

Purpose:Endophytic fungi are a range of microbes that live in the tissues or organs of healthy plants at a certain stage or whole stages.Long time of living with plants and interacting with each other makes it possible for endophytic fungi to influence secondary metabolites on structure and bioactivities.Anoectochilus roxburghii shows lots of potent pharmacologic activities including anti-inflammatory activity,liver protection,immunoregulation,nervine,hypoglycemic and blood lipid reduction,so this plant is known as"the king of medicine".However,little attention has been paid to the endophytic fungi of A.roxburghii until now.Thus,we concentrate on the research of endophytic fungi of A.roxburghii to search for biologically active secondary metabolites with novel structures.As a result,three endophytic fungi consist of Pestalotiopsis oxyanthi,Aspergillus versicolor and Aspergillus ostianus,which isolated from the leaves of A.roxburghii(Yongan City,Fujian Province,China),are selected to study on their secondary metabolites for the discovery of novel compounds with excellent bioactivity,and moreover,it will contribute to the breakthrough of leading compounds and the development of drugs.Methods:Study on three selected fungi in this dissertation mainly contains solid rice fermentation,separation of secondary metabolites and biological activities screening.The isolated compounds are obtained after suffering ODS,silica gel column chromatography,Sephadex LH-20 and HPLC.Their structures and absolute configurations are determined by comprehensive analysis of NMR,electronic circular dichroism(ECD)calculation quantum-chemical ¹³C NMR calculations,Rh₂(OCOCF₃)₄-induced ECD,selective oxidation reaction and X-ray crystallographic experiment.Part of isolated compounds are screened for their cytotoxicity,immunosuppressive activity and anti-inflammatory activity after the construction of experimental model.Result:Three selected endophytic fungi in this dissertation afford 109 compounds including 41 new ones in total.A total of 34 compounds are separated from P.oxyanthi,16compounds of which are new,including 12 azaphilones,one linear polyketone and three ergosterol-type steroids(Chapter one).A total of 44 compounds are obtained from A.versicolor.,including 13 new compounds which comprising two isochromane lactones,two benzolactones,one benzopyran,one benzofuran,one furancarboxylic derivate,one diketopiperazine,one ergosterol-type steroid,one pseurotin,two butyrolactones and two diphenolic derivatives(Chapter two).A total of 32 compounds are yielded from A.ostianus,12 new compounds among them contains nine hydroxyphenylacetic acid derivatives,one isochromane lactone,one terphenyl and one other compound(Chapter three).Part of isolated compounds are evaluated for their cytotoxic activity,anti-inflammatory activity and immunosuppressive activity,and some of them show corresponding activity.Chapter four reviews the research progress of secondary metabolites of the all genus Pestalotiopsis that have been reported from 2011 to 2020.Conclusion:The secondary metabolites of three endophytic fungi from A.roxburghii are different,and the activities of different compounds are also different.The secondary metabolites from these three fungi have been greatly enriched by the discovery of those new natural products.In bioactivities,the discovery of new ergosterol-type steroid with potent biological activity has laid a significant foundation for the subsequent research on the same type of analogues.Novelty:It is the first time that 12 azaphilones possess a methyl group at C-9 were reported;for the first time,the configuration at skeleton and branch chain of azaphilones are determined systematically and the culture conditions for single crystal are explored;it is the first time that ergosterol steroids with methyl group at C-23 are isolated from fungi as secondary metabolites;for the first time,the secondary metabolites of A.ostianus except marine sources are studied;in addition,nine hydroxyphenylacetic acids are firstly isolated from A.ostianus;the research progress of secondary metabolites of the genus Pestalotiopsis from 2011 to 2020 are firstly reviewed.