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目的建立肺炎链球菌18C型多糖结合物中游离多糖含量的测定方法。方法采用不同条件预处理样品,将结合多糖与游离多糖分离,用改良蒽酮法测定结合物中总糖与游离多糖的浓度,计算出结合物中游离多糖的含量,并对检测方法进行验证。结果采用5mol/L氯化钠溶液、85%乙醇溶液及-20℃冻存72h预处理样品,结合多糖与游离多糖的分离率及实验的有效性均最高。采用本方法测定肺炎链球菌18C型多糖结合物中游离多糖含量,结合多糖与游离多糖分离率的变异系数CV值为1.7%,实验有效性变异系数CV值为5.6%;检测3个浓度供试品的平均回收率分别为98.6%、98.5%和97.0%;检测1批多糖结合疫苗中游离多糖含量的CV值为4.2%。结论已建立肺炎链球菌18C型多糖结合物中游离多糖含量的检测方法,该方法可靠性强,准确性高,精密性良好。
Objective To establish a method for the determination of free polysaccharide in Streptococcus pneumoniae 18C polysaccharide conjugate. Methods The samples were pretreated with different conditions, the polysaccharide was separated from the free polysaccharide, the total polysaccharide and free polysaccharide concentration in the conjugate was determined by modified anthrone method, and the content of free polysaccharide in the conjugate was calculated. The detection method was also validated. Results The samples were pretreated with 5mol / L sodium chloride, 85% ethanol and frozen at -20 ℃ for 72h. The separation efficiency of polysaccharide and free polysaccharide was the highest. The method was used to determine the content of free polysaccharides in Streptococcus pneumoniae 18C polysaccharide conjugate. The coefficient of variation (CV) of polysaccharide-free polysaccharide separation rate was 1.7%. The coefficient of variation (CV) was 5.6% The average recoveries of products were 98.6%, 98.5% and 97.0%, respectively. The CV value of free polysaccharide in one batch of polysaccharide conjugate vaccine was 4.2%. Conclusion The detection method of free polysaccharide in Streptococcus pneumoniae type 18C polysaccharide conjugate has been established. The method is reliable, accurate and accurate.