Lepista Sordida Liquid Fermentation And Polysaccharide Extraction And Preliminary Study On Immunological Activity

Lepista sordida not only has excellent characteristics of safe non-toxic, easy separation and short production cycle, but also has stable yield and quality and higher price in actual production.At present, many research studies on the culture medium lepistasordida, component analysis of the fermentation product components and metabolites have been reported.But for less,the reports about the extraction,purification and immunity from fermentation product of Lepista sordida are less.The experimental research is focused on extraction and purification of extracellular and intracellular polysaccharide from Lepista sordida mycelium and fermentation liquid,and exploring its immunological activity, the experimental results as follows:1. The fermentation conditions were optimized in the liquid fermentation: the optimum conditions of B5 strain is the speed of 150 r/min, the fermentation time is 10 days, the temperature is 25 ?, the inoculation amount was 6%; the optimum conditions of 05 strain is the speed of 120 r/min, the fermentation time is 10 days, the temperature is 30?, the inoculation amount was 6%.2. The extraction conditions were optimized in the experiments of crude polysaccharide extraction: the of extracellular polysaccharide B5 strain is the best combination of extraction temperature is 80?, 80% ethanol concentration, extraction times was 3, pH was 7, and the best combination of 05 strains of crude extracellular polysaccharide extraction is the extraction temperature is 90? and the ethanol concentration is 80%, extraction times was 3, pH value is 6.The best combination of the extraction of intracellular polysaccharide from B5 strain is the extraction temperature is 80?, the ethanol concentration is 80%, the extraction times is 3, and the pH value is 7. The best combination of the extraction of crude polysaccharide from the 05 strains is the extraction temperature is 90?, the ethanol concentration is 80%, the extraction times is 3, and the pH value is 6.3. The crude polysaccharide was obtained by the optimum liquid fermentation and optimum extraction conditions. After the removal of protein by papaya protease and Sevage binding,hydrogen peroxide oxidation method, the removal of small molecules by dialysis method andmultiple purification of dextran gel,the single and symmetrical polysaccharide peaks were obtained. The purity of the extracellular and intracellular polysaccharides of B5 strains were87.59% and 84.62%, respectively. The purity can be used for immune test.4. Preliminary study on the immune activity of purified polysaccharide showed that different concentrations of intracellular and extracellular polysaccharides significantly promote the proliferation of mouse spleen lympHocytes in vitro, promote the secretion of cultured spleen lymphocyte TNF-? factor; has increased significantly hypoimmune mice IgA, IgG, IgM in the normal mice and the secretion of serum hemolysin, have a certain role in promoting IL-4cytokines IL-2, IFN-? and TNF-? secretion on immunosuppressed mice. In summary, Lepista sordida intracellular and extracellular polysaccharide not only have certain immune efficacy, but also the immune effect of intracellular polysaccharide was better than the immune effect of extracellular polysaccharide.