Study On The Active Metabolites Of Endophytic Fungi CWJ2 In Acanthopanax Senticosus

Acanthopanax'senticosus(Rupr.et Maxim)Harms is a medicinal plant which mainly distributes in Northeastern area of China,due to the widely utilization in clinical and pharmaceutical industry,the wild resource of the plant is limited.Endophytes are microorganisms which grow within their plant hosts without causing apparent disease symptoms.Many studies confirmed that endophytes are capable of producing secondary metabolites that are also biosynthesized by their host plants,and many endophytes are also prolific producers of novel compounds with outstanding potentials for generating novel therapeutic leads.As a result,it has great scientific research value and development potentiality for researching the endophytic fungi resources in medicinal plants which will bring significant ecological and economic benefit.In this work,an endophytic fungi named CWJ2 which isolated from the root of Acanthopanax senticosus was chosen as the research object.Gas chromatography-mass spectrometry(GC-MS)was used to determine its volatile of fermental liquid and the active secondary metabolites of CWJ2 were seperation and identified by modern analytical technology,then the DPPH radical scavenging assay was used to preliminarily detect the antioxidant activities of its secondary metabolites.Meanwhile,the species relationship of strain CWJ2 was determined.In conclusions:1.Volatile compounds of fermental liquid of endophytic fungi CWJ2 was extracted by ethyl acetate and gas chromatography-mass spectrometry(GC-MS)was used to determine its chemical components,and normalization was employed to determine its relative contents.Results showed that 50 compounds given by GC-MS was identified,the main kinds components contained s-Triazaborane;1,2-Benzenedicarboxylic acid butyl octyl ester;2-Cyclohexen-l-one and 2-ethyl-1,4-dimethyl-Benzene.2.The endophytic fungi CWJ2 were fermented in large scale,then the bioacitivity compounds in the fermentation liquid were isolated by liquid-liquid extraction,and compounds in mycelium were isolated by Ultrasonic extraction.Silica gel column chromatography,Polyamide TLC and other methods were used to seperate the motabolites of CWJ2.Three compounds were obtained,they are Crystal 1,Crystal 2 and Crystal 3.3.The structures of three compounds were analyzed by means of HPLC,1H-NMR,13C-NMR and HPLC/MS.Crystal 1 was pseurotin A,but the structure of Crystal 2 and Crystal 3 can not be determined for little amount.4.The antioxidant activities of three compounds were preliminarily evaluated by using DPPH radical scavenging assay.The results showed that three samples have antioxidant activity,and the activity of antioxidant were enhanced with the adding of concentration between 12.5 and 200?g/mL.In 200?g/mL,the DPPH radical scavenging rate of the three samples were 47.4%,95.6%and 94.8%,respectively.5.The endophytic fungi CWJ2 was identified as Aspergillus versicolor by morphological characteristics and 18S rDNA sequence analysis.The active metabolites of endophytic fungi CWJ2 were isolated and purified for the first time.The fungus appears to produce the important bioactive ingredient,pseurotin A.The current research indicates that a pseurotin A-producing endophytic fungus CWJ2 isolated from Acanthopanax senticosus was got.