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目的:建立用层析法去除脑脊髓膜炎球菌A群荚膜多糖样品中的杂蛋白的方法,以替代传统的苯酚抽提。方法:A群流脑多糖粗糖经过新型多模式离子交换填料Capto adhere,使用低盐上样高盐洗脱,洗脱的目的峰再经过Sephardex G-25凝胶过滤脱盐换成水,即为流脑多糖精糖原液。结果:采用上述方法制备的精制多糖样品中杂蛋白占固总的比例低于0.2%,符合WHO现行规程中的杂蛋白比例低于1%的要求,核酸、内毒素等生化指标均符合现行规程,整个层析过程的回收率大于65%。结论:层析法去除A群流脑多糖粗糖中的杂蛋白是有效的,比传统的苯酚抽提的方法更有利于环保,易于操作,易于放大。
OBJECTIVE: To establish a method for the removal of heterologous proteins in the sample of A group of capsular polysaccharide from M. meningitidis by chromatography, which can replace traditional phenol extraction. Methods: A group of cerebrospinal fluid crude polysaccharides through a new multi-mode ion exchange packing Capto adhere, using low salt loading high salt elution peak of the elution through Sephardex G-25 gel filtration desalination replaced by water, that is, the flow Brain polysaccharide refined sugar solution. Results: According to the current regulations of WHO, the proportion of hybrid proteins in the refined polysaccharides samples accounted for less than 0.2% of the total solids, which meets the requirement of less than 1% of the hybrid proteins in the current WHO regulations. The biochemical indexes such as nucleic acids and endotoxins all met the current regulations , The recovery rate of the whole chromatographic process is more than 65%. CONCLUSION: Chromatography is effective in removing impurity from crude cereal polysaccharides of Group A melatonin, which is more environmentally friendly, easier to operate and easier to enlarge than the traditional phenol extraction method.