Isolation, Purification And Immunomodifying Activity Of Polysaccharides From The Fruit Bodies Of Grifola Frondosa

Maitake (Grifola frondosa) is a polypore mushroom that grows in clusters at the base of trees in temperate forests. The fungus is native to many provinces such as Hebei, Zhejiang, Sichuan, Jilin and Fujian in China. Great attentions have been paid to artificial cultivation of this rare edibe fungi for over 30 years in Zhejiang and Hebei province due to its delicious tastes and unique aroma. It is also prized as a medicinal mushroom rich in a variety of active substances. G. frondosa polysaccharide, main active ingredient, has been identified to have significant effects in immunity-improving, anti-tumor, anti-hypertensive, anti-diabetic, anti-hyperliopisis, as well as many other pharmacological activities.In this study, crude polysaccharides were purified, and then their activities were tested. In addition, structures of purified polysaccharides were analyzed using chromatographic and spectroscopic methods. Further tests were also implemented to evaluate the immune activity of polysaccharides.A water-soluble, crude polysaccharide preparation was obtained from Grifola frondosa fruit bodies by hot water (100℃) extraction followed by ethanol (75%) precipitation. Four purified polysaccharides, GFP75-2C, GFP75-2D, GFP75-3C1 and GFP75-4B, were obtained from the crude material by successive fractionation using DEAE-Sepharose Fast Flow ion exchange chromatography and Sephacryl S gel filtration chromatography. UV spectroscopy detected no protein or nucleic acid in them, which mainly consisted of fucose, galactose, and mannose.The four polysaccharides, except GFP75-2C, showed the activities to stimulate the proliferation of lymphocytes and enhance the release of NO from RAW264.7 macrophages. The immune activity of GFP75-2D which distinguished from others was carried out in-depth study. After exposure to GFP75-2D, RAW264.7 cells increased in size and formed numerous pseudopodia. Furthermore, GFP75-2D stimulated the macrophages to produce NO. Interleukin-1β(IL-1β) and TNF-a gene expression was significantly increased after exposure whereas slight changes were recorded in IL-6 gene expression. Large increases in IL-6, IL-1β and TNF-αprotein production were also recorded.