Purification,Structural Characterization And Anti-obesity Effect Of Polysaccharide From Lactarius Deliciosus Gray Mycelia

Lactarius deliciosus Gray is a kind of precious edible fungus.At present,its fruiting body has been found to have a variety of biological activities,and the main functional components are concentrated in polysaccharides.However,the fine structure of mycelia polysaccharide has not been characterized and the mechanism of anti-obesity activity has not been elucidated.In this study,the mycelia of Lactarius deliciosus Gray were obtained by liquid fermentation.A novel polysaccharide was extracted from the mycelia of Lactarius deliciosus Gray by hot water extraction and alcohol precipitation,and then it was purified.The polysaccharide structure was characterized,and its anti-obesity effect was studied.It provides a theoretical basis for the utilization and the development of Lactarius deliciosus Gray mycelia polysaccharides.The main findings are as follows:Taking the biomass of Lactarius deliciosus Gray mycelia as the index,the formula of the fermentation medium of Lactarius deliciosus Gray mycelia was optimized.The results of single factor and orthogonal experiment showed that the best fermentation conditions for Lactarius deliciosus Gray mycelia were:glucose 2.5%,yeast extract 0.5%,KH₂PO₄+Mg SO₄ 0.06%,and the biomass of mycelia was 1.36 g/100 m L.Taking the yield of the polysaccharide from Lactarius deliciosus Gray mycelia as the index,the extraction of polysaccharide from Lactarius deliciosus Gray mycelia was optimized.The results of single factor and orthogonal experiment showed that the optimal extraction conditions of polysaccharide from Lactarius deliciosus Gray mycelia were as follows:extraction temperature 80℃,extraction time 4 h,Material-liquid ratio1:40 g/m L,and the yield was 8.11%.The dried Lactarius deliciosus Gray mycelia was collected and subjected to hot water extraction,ethanol precipitation,protein removal by Sevag method,dialysis bag dialysis,and vacuum freeze-drying to obtain the crude polysaccharide LDP,and the yield of LDP was 6.84%.LDP was separated by DEAE-52 cellulose ion-exchange column chromatography to obtain the elution fraction LDP-C.The fractions were collected and further purified by Sephadex G200 to obtain the pure polysaccharide LDP-CP,and the sugar content of LDP-CP was 96.58%,detected by phenol-sulfuric acid method,and the molecular weight was 2.33×10³ k Da measured by high performance liquid chromatography.The specific rotation is+41^o.The monosaccharide composition analysis showed that LDP-CP was composed of mannose,glucose and galactose in the relative molar ratio 1.33:1.06:1.00,respectively.Through infrared analysis,periodate oxidation,Smith degradation,methylation analysis and nuclear magnetic resonance spectroscopy,the glycosidic bonds of LDP-CP were as follows:→3)-β-D-Manp-（1→,→2,4）-α-D-Manp-(1→,α-D-Manp-（1→,→4）-β-D-Glcp-（1→,→2）-β-D-Manp-（1→,→4,6）-α-D-Manp-（1→,→4）-α-D-Galp-（1→,→2,3）-α-D-Glcp-(1→andα-D-Glcf-(1→.The results of animal experiments showed that:Lactarius deliciosus Gray mycelia polysaccharide could significantly reduce the weight growth trend of obese mice,reduce serum triglyceride,total cholesterol and low-density lipoprotein cholesterol levels,and increase serum high-density lipoprotein cholesterol levels in obese mice.The results of western blotting experiments showed that LDP could enhance the expression of AMPKα1protein,reduce the level of FAS,regulate the production of PPARαand lipid metabolism,and reduce fat accumulation,thereby reducing the body weight of mice.