A Study On The Separation And Flavoniods-Producing Of Ginkgo Biloba L. Endophytic Fungi

Endophytic fungi now its definition refers to some parasitic microbial do not cause the obvious symptoms of disease in plant tissue during a stage of life history, including those temporarily harmless latent pathogenic microorganisms and mycorrhizal fungi. When research reports Taxomyces andreanae is capable of producing taxol endophytic fungi which isolated from Taxus brevifolia, aroused great interest in the study of antitumor activity of Endophytic fungi parasited in Gymnosperm. In order to solve the resources problem, in recent years the medicinal plant endophytic bacteria become a hot topic in new medicinal plants field researchIn this paper, experimental materials based on Chinese endemic plants of Ginkgo biloba L, separated the Endophytic fungi of Ginkgo biloba root, stem, leaf tissue which sampling in Anhui Shucheng and Hefei area, experiment materials use 75% ethanol doing the surface sterilization, the result of repeated experiments show that, the best disinfection time of Different Ginkgo tissueparts respectively:Ginkgo biloba root 3min, ginkgostem 2min, ginkgo leaf 1min. Under this condition, using the method of tippicking, the fungi from Ginkgo biloba root, stem, leaf tissues were isolated and purified by a total of 60 strains of endophytic fungi, according to bacterial colony morphology, color come into the preliminary morphological analysis, ruled out the same colony, finally obtained 20 strains of Endophytic Fungi in Ginkgo biloba.Identification by morphological, microscopic photography revealed hyphae of endophytic fungi in the growth process is mostly transparent, and no vibration, belong to the basic Trichothecium sheath sp., Verticillium these three genera.In this paper, through a combination of colorimetric method, Aluminium chloride colorimetry and thin layer chromatography do the preliminary qualitative analysis on isolated endophytic fungi ferment, so as to screen out the flavone producing strains. Experiments show that, if the flavonoids contained in the fermentation products, fermentation products and aluminum chloride reaction, part show the yellow green fluorescence. Fermentation product and sodium hydroxide, part of a yellow appearance. Fermentation products and boric acid reaction, part appears brown yellow. Otherwise, there is no such phenomenon.Then doing the quantitative analysis of flavonoid producing strain fermentation by ultra high pressure liquid chromatography, which screened out the high flavone producing strains, experiments show that:after fermentation and complex sieve flavone producing capacity of 5 strains more than 5 mg/mL.Experiments were conducted to activity identification of 5 strains of high flavonoids strains fermentation products, inhibiting effect intensity on Escherichia coli, Bacillus subtilis, Staphylococcus aureus were measured. Experiments show that, five kinds of fermentation liquor has antibacterial activity, and the antimicrobial effect of strain No.2013015 is the best, the following experiment choose No.2013015 as the experimental strains.This paper for conditions of liquid fermentation, explored the optimization from two aspects of the fermentation culture medium and culture conditions, the optimal conditions to produce flavonoids obtained in the liquid fermentation strains. Experiments show that, the system best flavone producing strain fermentation medium conditions are as follows:3% glucose,0.5% peptone,0.4% yeast extract,0.3%KH2PO4,0.01%MgSO4,7H2O; the optimum culture conditions for:the initial pH value 7, the optimum fermentation temperature of 28, the rotate speed is 140 r/min, the volume of liquid 125mL/250mL; this provides the basis theory for the realization of industrial production, it has very good reference value and guiding significance.The demand of Ginkgo biloba flavonoids drug is increasing, almost in a state of shortage, just rely on the extraction of Flavonoids from Ginkgo biloba leaves, will result in the lack of resources,also damage the ecological environment. However, the fungus has the characteristics of easy culture, grow fast, easy to control, so it has broad application prospects, can find a new way for the development of Ginkgo flavonoids.