Isolation, Purification, And Structural Elucidatuion Of Polysaccharides From The Fruting Bodies Of Flammulina Velutipes

There are three parts in this research including the optimization of Flammulina velutipes polysaccharide water extracted method, physicochemical property research and sugar structure analysis.With the univariate and orthogonal experiments, the optimization condition was confirmed:solid-to-liquid ratio was 1:20, time was 3 h, temperature was 90℃and extract with 3 times. The impact effect of polysaccharide fractions from water extract was:solid-to-liquid ratio> times> time> temperature。The polysaccharide fraction from water extract of Flammulina velutipes after ethanol precipitation with the concentration of 60% was isolated and purified using DEAE Sapharose Fast Flow and Sephacryl S gel-filtration chromatography repeatedly. Tow water-soluble homogeneous polysaccharides FVP60-B and FVP60-C2 were obtained. Lack of absorption at 260nm and 280nm by UV scanning indicated that they contained no nucleic acid and protein. IR spectra showed that the sugar characteristic groups were existed. HPLC producing a single symmetrical peak, indicated that each of them was homogeneous and their molecular weight were 1.3×104 Da及1.4×104 Da.FVP60-B was investigated by sugar analysis, methylation analysis and NMR spectroscopic methods to confirm the sugar composition, the number of sugar residues and the form of glycosyl linkage. FVP60-B was a neutral polysaccharide and composed of fucose, mannose, glucose and galactose with the molar ratio of 0.71:0.82:1.00:4.88. And the methylation showed that it contained terminalof fucose linkage,1,6 and terminal linkage,1,6 galactose linkage and 1,2,6 mannose linkage. With the NMR analysis, it's showed that most of the sugar residue were existed as a configuration, including someβ-gylcosyl.FVP60-C2 was investigated by sugar analysis and showed that it was composed of fucose, xylose, mannose, glucose and galactose with the molar ratio of 0.74:0.21:1.36:1.00:1.31.