Extraction, Modification, Structural Characterization And Immunological Activity Of Polysaccharide From P

Rhizopogon rubescens(Tul.)Tul is a rare edible fungus with the certain anti-cancer activity,and polysaccharide is one of its effective ingredients,and there is no research report on the polysaccharide of Rhizopogon rubescens(Tul.)Tul.In this study,the fruit body of Rhizopogon rubescens(Tul.)Tul was used as the raw material to obtain the pure polysaccharide of Rhizopogon rubescens(Tul.)Tul after extraction,separation and purification,and its physical and chemical properties,primary structure and high-level structure are analyzed.To study the immunomodulatory activity of Rhizopogon rubescens(Tul.)Tul acid polysaccharide and carboxymethyl Rhizopogon rubescens(Tul.)Tul acid polysaccharide in mice,aiming to provide a certain experimental basis in the development and application of regulators.The main research results are as follows:1.Separation,purification and carboxymethyl modification of polysaccharides:Ultrasonic crushing assisted by hot water extraction is used to extract crude polysaccharide from Rhizopogon rubescens(Tul.)Tul(CPRR),Sevage removes protein to obtain refined polysaccharide from Rhizopogon rubescens(Tul.)Tul(RPRR),and small molecules are removed by dialysis.And DEAE-Cellulose 52 and Sephadex G-75 were used to separate and purify the polysaccharide from Rhizopogon rubescens(Tul.)Tul,and finally obtain the purified polysaccharide from Rhizopogon rubescens(Tul.)Tul(PPRR)with a purity of 92.06% and the yield was 0.68%.The acid refined polysaccharide from Rhizopogon rubescens(Tul.)Tul(RPRR-S)was purified by DEAE-Cellulose52 column to obtain carboxymethyl acid refined polysaccharide from Rhizopogon rubescens(Tul.)Tul(CM-RPRR-S),the degree of substitution is determined to be 0.352..2.Analysis of the physicochemical properties of polysaccharides: The total sugar,protein,uronic acid and sulfuric acid content of PPRR were determined by chemical methods,they were 92.06%,0.59%,2.09% and 1.98%,respectively,without flavonoids and total phenols.UV spectrum scanning PPRR found no nucleic acid,which contains a small amount of protein.3.Structural characterization of PPRR: Infrared spectroscopy shows that PPRR is mainly composed of pyranosides;High performance liquid chromatography shows that PPRR is composed of glucose,xylose and fructose,with the contents of 85.068%,5.872%and 9.060%,respectively;Nuclear magnetic resonance spectroscopy shows PPRR is α-configuration and β-configuration,both of which are pyran type,containing →3)-α-DXylp→6-β-D-Glcp polysaccharide fragment;Congo red experiment shows that there is a triple helix structure in PPRR;X-ray diffraction shows that the PPRR spectrum is irregularly dispersed peaks,indicating that PPRR exists in an amorphous structure;Scanning electron microscopy shows that the PPRR structure is relatively loose,mostly in sheet-like structures,mixed with a small amount of strand-like structures;The thermogravimetric-differential heat analysis shows that PPRR has a good thermal stability.4.Study on the immunomodulatory activity of RPRR-S and CM-RPRR-S: The experimental results show that both RPRR-S and CM-RPRR-S can improve the coat color,activity and physical condition of immunosuppressive mice;they can improve the damage of the spleen and thymus and protect the cell structure;they both can significantly increase the secretion of IL-2,IFN-γ and TNF-α(P<0.05),significantly reduce the secretion of IL-6 and IL-12(P<0.05);they both can significantly increase the number of red blood cells and platelets in the blood of suppressed mice(P<0.05),which has a certain protective effect on the bone marrow;they both can significantly activate macrophages and enhance the phagocytic ability(P<0.05).Among the above indicators,CM-RPRR-S shows stronger immunomodulatory activity than RPRR-S.In summary,RPRR-S and CM-RPRR-S have specific and non-specific immune functions in immunosuppressive mice,and the modified polysaccharide has the stronger immunomodulatory activity.