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目的:研究人工培养蛹虫草子实体多糖的分离纯化方法。方法:多糖经脱色、醇析、除蛋白及乙醇分级沉淀,采用Seph-adex G100柱层析纯化。结果:60℃脱色9h脱色率达70.72%。醇析最优条件为:无水乙醇,24h,4倍乙醇。Sevag法抽提30min重复6次除蛋白率达84.09%;抽提1h重复3次、6次除蛋白率分别为81.14%、84.47%,多糖损失率分别为27.03%、43.16%。木瓜蛋白酶除蛋白最优条件为70℃,1:10,2 h,除蛋白率为25.67%。粗多糖经乙醇分级沉淀得2种粗多糖成分,分别经Sepha-dex G100柱层析可进一步纯化为3种较纯多糖成分。结论:采用乙醇分级沉淀及Sephadex G100柱层析纯化粗多糖可得到3种较纯多糖成分,纯化效果较好。
Objective: To study the isolation and purification of polysaccharides from Cordyceps militaris cultured by artificial cultivation. Methods: The polysaccharides were purified by Seph-adex G100 column chromatography by decolorization, alcoholysis, protein removal and ethanol fractionation. Results: The decolorization rate reached 70.72% at 60 ℃ for 9h. The optimum conditions of alcoholysis: ethanol, 24h, 4 times ethanol. Sevag method extracted six times in 30min, the protein removal rate was 84.09%; extraction 1h repeated three times, 6 times the protein removal rates were 81.14%, 84.47%, polysaccharide loss rates were 27.03%, 43.16%. The optimal conditions of papain protein removal were 70 ℃, 1: 10,2 h, and the removal rate of protein was 25.67%. Crude polysaccharides precipitated by ethanol to give two kinds of crude polysaccharides, respectively, by Sephadex G100 column chromatography can be further purified to three pure polysaccharides. Conclusion: Purification of crude polysaccharides by ethanol fractional precipitation and Sephadex G100 column chromatography can provide three kinds of pure polysaccharides with better purification effect.