14型肺炎链球菌荚膜多糖纯化工艺中两种去除蛋白方法的比较

目的苯酚抽提法和脱氧胆酸钠沉淀法去除14型肺炎链球菌荚膜多糖中蛋白质的效果比较。方法将3批次14型肺炎链球菌发酵培养液经超滤、乙醇沉淀等方法初步纯化后,平分成两份,分别采用苯酚抽提法和脱氧胆酸钠沉淀法去除蛋白,通过比较多糖收获量、多糖组分检定结果、多糖分子质量、多糖抗原活性、多糖核磁共振图谱,以此评价这两种蛋白去除方法的效果。结果与苯酚抽提法相比,脱氧胆酸钠沉淀法制备的14型肺炎链球菌纯化荚膜多糖除收获量较高,蛋白和核酸杂质含量较低外,氨基己糖含量、多糖分子质量、抗原活性和多糖核磁共振图谱的检定分析结果无显著性差异(P>0.1)。结论作为14型肺炎链球菌荚膜多糖纯化工艺中的除蛋白方法,脱氧胆酸钠沉淀法优于苯酚抽提法。

Comparison of effects on the removing proteins from capsular polysaccharide of Streptococcus pneumoniae type 14 with two methods

Objective To compare effects of the removing proteins in purification process of Streptococcus pneumoniae capsular polysaccharide type 14 with two methods(phenol extraction and sodium deoxycholate precipitation).Methods After the fermentation liquid of Streptococcus pneumoniae type 14 was preliminary purified by ultrafiltration and alcohol precipitation,it was divided into two parts,and removed protein with the method of phenol extraction or sodium deoxycholate precipitation,respectively.Purified polysaccharide was analyzed including yield,biochemical characteristic,molecule weight,antigenicity and 1H-NMR spectrum.Results Compared with phenol extraction,the polysaccharide purified by sodium deoxycholate precipitation had higher yield and lower contaminant content.There was no significant difference on content of hexosamine,molecule weight,antigenicity and structure.Conclusion As the deproteinization method in purification process of Streptococcus pneumoniae capsular polysaccharide type 14,sodium deoxycholate precipitation was superior to phenol extraction.