Isolation And Identification Of Endophytic Fungi From Medical Plants Corydalis Yanhusuo,Preliminary Exploration On Endophyte Fungi With Functional Enzyme Activity And Phosphate Dissolving Ability

Corydalis?Corydalis yanhusuo?was also known as:Yan Hu,Yuan Hu,Poppy etc.Corydalis was one kind of perennial herb with spherical tubers;petal purple andcapsule cylindrical,tapering at both ends.Summer flowered.It had considerable effects on tadditional Chinese medicine and was important traditional medical resource.Endophytic fungi were those colonizing inside of various tissues and organs from healthy plants or all stages of their life cycles without leading the infected host plants to show obvious external symptoms,and their endogenous could be tested by histology method or isolatation from strict surface sterilized plant tissue,otherwise directly amplified microorganisms from plant tissue.Endophytic fungi belonged to special ecoenvironment microorganisms,the majority of which had characteristics about obvious patience of high salt,alkali,acid,cold.And the metabolites of endophytic fungi were not only same or similar with the host plants,,but also important source of novel compound,with potential development value of agricultural production,natural medicine property and industrial fermentation.The essence of the study was to isolate and identify endophyte fungi from wild Corydalis yanhusuo,to inquiry endophyte fungi with the ability of dissolving inorganic insoluble phosphorus,functional enzyme and antibacterial activity,furthermore to explore changes on the ability of dissolving inorganic insoluble phosphorus about 4 endophyte fungi isolates in Pikovaskaia liquid medium.1.Isolation and identification of Endophytic Fungi from medical plants Corydalis yanhusuoTo select fresh tubers of wild Corydalis yanhusuo from Chenggu Shaanxi as experiental material,endophytic fungi were isolated and purified by the application of a composite potato dextrose agar?CPDA?.Totally purified 312 endophytic fungi isolates were got and preserved by slant cryopreservation method of potato dextrose agar?PDA?.65 different morphotypes isolates were identified by morphological and molecular biology methods,the results suggested that they could be devided into genus Fusarium,Alternaria,Mucor,Aspergillus,Penicillium,Nectria,Rhizopus and Eurotiomycetes.2.Preliminary Screening of Endophyte fungi with Functional Enzyme Activity in Corydalis yanhusuoBy applying fibrinolytic enzyme,cellulase,alkaline protease,compound enzyme,catalaseand chitosanase screening medium to screen functional enzyme producing isolates from 65different morphotypes strains.The result displayed 25 strains with fibrinolytic enzyme activity,24 strains with compound enzyme activity,23 strains with protease activity,12strains with catalase activity,1 strains with cellulase activity,but no strains with chitosanase activity.The result revealed endophyte fungi within Corydalis yanhusuo had metabolic diversity,and 38 isolates indicated promising potential for deployment in biotechnological processes involving production of fibrinolytic enzyme,cellulase,alkaline protease,compound enzyme and catalase.3.The Exploration of Endophyte Fungi with Phosphate Dissolving Ability in Corydalis yanhusuoTo preliminaryly screen phosphorus solubilizing strains in use of inorganic phosphorus solid medium and solid Mengjin Na organophosphorus medium,25 of 65 isolates had inorganic phosphorus solubilizing ability,and two isolates with organophosphorus dissolving ability,it was worthy to say No.YH-84?Penicillium asturianum?strains had dual phosphorus dissolving activity.The research showed metabolic diversity of endophyte fungi within Corydalis yanhusuo and complemented the exploration of phosphate-solubilizing fungi from special ecoenvironment.The research about antibacterial activity of 25 isolates with inorganic insoluble phosphate dissolving ability was done by modified bacterial antagonism experiment against Staphylococcus aureus,Bacillus subtilis,Escherichia coli,Salmonella typhi as indicators.YH-102 showed strong antibacterial activity against four kinds of indicator bacteria,YH-84had strong antibacterial activity against Salmonella typhi,Staphylococcus aureus and Bacillus subtilis,other strains did not show strong antibacterial activity.When the operational conditions were controled at pH 8,28?,in darkness,with the inoculum of 200?L or 1000?L,four different carbon and nitrogen sources in Pikovaskaia liquid medium,the author inquiryed changes about dissolving capacity of Ca₃?PO₄?₂ of foure endophytic fungi from Corydalis yanhusuo.The outcome revealed the highest concentration of soluble phosphorus was 5.178 mg/L when the inoculum was controled at 0.2%?V/V?,nitrogen and carbon source were yeast extract and glucose respectively.With the inoculum of1%?V/V?,the concentration of soluble phosphorus in broth about foure isolates increased obviousely and reached peak at 36th houre or 40th houre after inoculation respectively.The peak of soluble phosphorus was 5.754 mg/L.Under alkaline conditions,endophytic fungi from Corydalis yanhusuo showed certain solubilizing capability during 40 houres after inoculation.