Study On The Liquid Fermentation Of Acid-Tolerant Mycelia And Biological Activities Of Exopolysaccharide From Lentinus Edodes

The xylotrophic basidiomycete Lentinus edodes (L. edodes), commonly known as one of the most important edible and medicinal mushrooms, contains a highly functional biomacromolecule-lentinan which improves the human immune system and shows anti-cancer, antioxidant, and anti-viral activities. Up to now, because of the limitation in polysaccharides content of fruit body, more and more interest has been focused on research of liquid fermentation and the isolation and purification of exopolysaccharide (EPS) from L. edodes. Aimed at developing the open culture technology in acid condition of L. edodes mycelia and the future industrial production, the study was mainly focused on screening acid-tolerant L. edodes mutants and optimizing further the fermentation conditions.Atm-02, an optimal mutant that can meet the acid condition, was selected for open culture by ultraviolet irradiation and selective medium (pH=2.0) in turn. The optimal fermentation media by single factor tests was:sucrose 30～40 g/L, beef extract 7.0 g/L, KH2PO4 4.0 g/L, MgSO4 2.0 g/L, vitamin B1 0.15 g/L, and the optimal fermentation conditions were:culture temperature 26℃, loading volume 100 mL of 250 mL flask, rotation speed 180 rpm and inoculation volume 10%(v/v).The results showed that the maximum EPS production was up to 0.172±0.023 g/100 mL in open culture.EPS from Atm-02 was first purified by DEAE-52 ion exchange column, and then the neutral polysaccharide fraction and acidic polysaccharide fraction were gained. The yield ratios were 12.9% and 40.9%, respectively. Subsequently, the results of paper chromatography and Sephadex G-100 gel filtration column chromatogram indicated that these polysaccharide fractions were both single component.The structure of EPS was preliminarily studied. The neutral polysaccharide was a kind of polysaccharide with the structure of β-D-pyran glucan while the acidic one contained a-D-furan glucan, and they didn’t contain protein and nucleic acid by UV spectrum and IR spectrum. After pre-column derivatization, the high performance liquid chromatographic analysis showed that the neutral polysaccharide fraction was composed of glucose, mannose, rhamnose, galactose, xylose and arabinose; whereas the acidic one was mainly composed of glucose and negligible mannose.Different purity of EPS showed that they all had certain antioxidant activities. The crude polysaccharide and deproteinization polysaccharide showed the greatest reducing powers, with 43.60±0.31 mg Vc/g and 36.78±0.26 mg Vc/g, respectively; they both had a stronger DPPH free radical and ·OH free radical scavenging effect than neutral polysaccharide and acidic polysaccharide.Furthermore, the fermentation filtrate,0.5% crude protein polysaccharide,0.5% deproteinization polysaccharide and 0.5% neutral polysaccharide from Atm-02 all played an important role on the growth promotion of Glycyrrhiza uralensis Fisch. The results indicated that the effect of fermentation filtrate on the shoot growth was more efficient than that of any other treatments, whereas 0.5% deproteinization polysaccharide solution effectively enhanced the root growth.