金针菇子实体多糖分离纯化及结构和免疫活性研究

Extraction, structure characteristics and immune activities of polysaccharides from the fruiting body of Flammulina velutipes

从金针菇子实体中分离纯化多糖,并对多糖结构和体外免疫活性进行研究。采用水提醇沉法从金针菇子实体中提取粗多糖,利用DEAE‐Cellulose‐52及Sephacryl S‐300HR柱层析纯化得到FVPⅠ‐a,再利用HPLC‐ELSD技术、红外及核磁共振对FVPⅠ‐a进行结构解析。在体外以促RAW264.7巨噬细胞产NO、分泌细胞因子,促进小鼠淋巴细胞增殖实验,考察FVPⅠ‐a增强免疫的能力。从金针菇子实体中分离纯化得到FVPⅠ‐a,其为分子量81.4kDa,由葡萄糖、果糖和鼠李糖组成的β构型的吡喃型杂多糖。体外免疫实验表明,FVPⅠ‐a能够促进RAW264.7巨噬细胞产生NO及分泌细胞因子(IL‐1β,IL‐6,TNF‐α),能单独的促进小鼠淋巴细胞增殖(P<0.05),并能协同增强ConA和LPS对小鼠淋巴细胞的促增殖作用(P<0.01,P<0.05)。首次从金针菇子实体中获得FVPⅠ‐a杂多糖,其在体外具有增强非特异性免疫反应及增强特异性免疫反应的能力。

The polysaccharides from the fruiting body of Flammulina velutipes were extracted, and the immunity-enhancing activities of the polysaccharides in vitro were evaluated. By water extraction and alcohol precipitation, crude polysaccharides were obtained. Through purification by DEAE-Cellulose 52 ion exchange chromatography and SephacrylS-300 HR gel filtration chromatography, a polysaccharide named as FVP I-a was obtained. HPLC-ELSD technology, IR and NMR were used to evaluate the structure of FVP I-a. NO production of RAW264.7 macrophages, cytokines secretion and lymphocyte proliferation were used as the indicator to evaluate the immunity-enhancing activities of F. velutipes polysaccharides in vitro. FVP I-a was mainly composed of glucose, a trace amount of fructose and rhamnose, and the molecular weight was 81.4kDa. IR analysis indicated that FVP I-a was a β-pyranoside polysaccharides, and NMR analysis also hinted that FVP I-a was a β-polysaccharides. FVP I-a can largely promote the NO production and augment the IL-1β, IL-6, TNF-α secretion of RAW264.7 macrophages. FVP I-a can promote lymphocyte proliferation （P〈0.05）, and synergistically enhance the effect of ConA and LPS on the proliferation of mouse lymphocytes （P〈0.01, P 〈0.05）. The FVP Ⅰ-a heteropolysaccharide was obtained from F. velutipes for the first time, and it could enhance non-specific immune response and specific immune responses in vitro.