Study On Endophytic Bacteria Of Astragalus And Its Active Metabolites

Astragalus membranaceus(Fisch.)Bge.,a famous traditional herb,whichis native to northChina.It contains a variety of pharmacological compounds and has antioxidant activity,diuretic activity,improving immunological activity and many other effects,which has been demanded greatly,so the wild Astragalus membranaceus resources is in short supply.In this research,endophytes were isolatedfrom Astragalus membranaceus and its active metabolites have been studied to find the similar or new active metabolites as their hosts.The researchmay alleviate the shortage of medicinal plant resources and create a method for producing the active ingredients ofAstragalus membranaceus.In this paper,healthy Astragalus membranaceus,which was collected from Da Hinggan Mountains was chosen as objects,andendophytes was isolatedfrom Astragalus membranaceus by surface disinfection method.All the endophytes were screened by antimicrobialand antioxidant tests,meanwhile,thefermented extractsof endophytes was analyzed by using HPLC.Finally,the target strains were identified.Resultswere as follows:66 endophytes were seperated from Astragalus membranaceus,there are 30 endophytic fungiand 36 endophytic bacteria,and 66 strains of endophytes and their secondary metabolites were screened through antimicrobial activities against 11 kinds of pathogens,such asEscherichia coli,Staphlococcus aureus,Candida albicansand so on.There are 56 strains have different degrees of inhibition activity,accounting for 84% in the total 66 endophytes.The rate of restrain Escherichia coli,Bacillus subtilis,Staphylococcus aureus,Bacillus pumilus,Enterococcus faecium,Enterococcus faecalis,Listeria monocytogenes,Acinetobacter baummanii,Klebsiella pneumoniae,Pseudomonas aeruginosa,Candida albicansisrespectively46.9 %,54.5%,30.3%,25.8%,57.6%,51.5%,25.8%,57.6 %,46.9%,42.4%,22.7%.In this work,antioxidantactivities of 66 endophytes sampleswerestudiedbyDPPHmethod,andDPPHfreeradicalscavengingcapabilities were 9 strains endophytes had antioxidant activity,among them,PS13,NS8,NS14,and NR14 have the same antioxidant capacitybelow the level of vitamin C's antioxidant and the endophytic fungi PS9 has nearly the same effect of clearing DPPH as Vitamin C.Compared with host plantsmples,the metabolites of endophytes were analysis with HPLC,results showed that the metabolits of PS13,NS8,NS12 and NS14 may produce the similar active compounds as Astragalus membranaceus,and then,the fermentation broth of PS13,NS8,NS12 and NS14 may have formononetin.30 fungi were divided into 3 Orders,4 families,12 genuses by morphological identification,and the Aspergillus strains was advantage fungi,which account for 26.7%.The target strain PS13 was identied as Alternaria alternate by morphology observationand 18 S rDNA sequence analysis,NS8 was determined as Bacillus amyloliquefaciens by physiological experiments,biochemical experiments and 16 S rDNAsequence analysis.As a conclusion,there are a lot of endophytes in healthy plant of Astragalus membranaceus,some of them can producethe similar or new active metabolites as their hosts.These provided a new way to exploit endophytes and develop new drugs.Meanwhile,thefeasibilityofmicrobialfermentationfor Astragalus membranaceus active metabolites have the great significance to protect the ecological environment.