Isolation And Optimization Of Flavone-producing Endophytes From Ginkgo Biloba.L

Ginkgo bilobaLinn,belongs to the gingko family ginkgo deblade high megaphanerophyte, is a typical primitive relic plant in China,for the national secondary protection plants,is the "living fossil" in plant.Its extract contains many active ingredients for human health.The flavone(gingko bilobaL flavonoid's principal constituent are quercetin,kaempferol and isorhamnetin) is mostly researched and used in gingko's function ingredient.At present,the flavone is withdrawed from the Ginkgo bilobaL's leaf,which can cause drying up of the resources, moreover the process of produce also is restrained.Research results in recent years show that endophyte can produce some metabolites similar to those from the host plant.This feature could make endophyt became a new resource which can produce some compounds with novel structure and specific bioactivity.It has greatly potential for development and applicaton in the agriculture and the medicine industry.For above reasons,this experiment selected Ginkgo bilobaL as original materials,to isolate and identify their endophytes,and screen the bioactive strains from them.Meanwhile, we studied the bioactivities of one bioactive strain in initial stage.Result as follows:1. 126 strains endophytes were isolated from the skin,root,leaf and fruit of ginkgo bilobaL,including 71 strains bacteria,49 strains fungus and 6 strains actinomyces;2. 8 strains flavone-producing endophytes were screened by HPLC.Among the flavone- producing strains,7 strains are fungi,1 strains is actinomycetes; The results of morphologically identified are Stemphylium sp,Fusella Sacc,Alternaria sp,Fusarium sp,Gibberella sp,Sphacelia sp,Actinomyces and Dematium Pers;The output of isorhamnetin is lower,the output of quercetin and kaempferol is higher along these flavonoids.The maximum production may reach respectively 0.851mg/L and 0.817mg/L;3. These 8 strains endophytes were continuous cultived,the spores all produce the flavanone in the inside and outside.The total flavone output increased with the growth rate; 4. The strain A2 among the 8 strains was excellent with high production and growth rate. The cultural conditions of A2 strain was optimized by the orthogonal design experiment.The optimum condition for fermentation is 28℃,80mL media volume in 250mL flasks,180 rpm, inoculation 1 link,the initial pH 7.0.The best carbon and nitrogen source are glucose and ammoniumnitrate respectively,the growth factor is VC.