重组乙肝疫苗纯度高效液相色谱法的研究与应用  

Research and application of HPLC for the purity of recombinant hepatitis B vaccine

作　　者：周志平 韩斌 董洁 ZHOU Zhi-ping;HAN Bin;DONG Jie(Hualan Biological Bacterin Co.Ltd,Xinxiang 453003,China)

机构地区：[1]华兰生物疫苗有限公司,河南新乡453003

出　　处：《生物技术》2020年第5期453-456,518,共5页Biotechnology

摘　　要：[目的]建立检测重组乙肝疫苗(汉逊酵母)纯度的高效液相色谱法(HPLC)。[方法]采用TSK gel G5000PW色谱柱,优化流动相、流速、柱温及p H等参数,确定最佳试验条件;并与TSK gel G3000SW色谱柱检测乙肝纯度的方法进行比较。[结果]确定了最佳试验条件:流动相为0.01 mmol/L PB(磷酸缓冲液),流速为0.6 m L/min,柱温为室温(20℃),p H为7.0,检测波长280 nm;乙肝表面抗原(HBs Ag)在色谱柱TSK gel G5000PW和TSK gel G3000SW的保留时间分别为8.5 min、17.5 min;检测时间分别为20 min、40 min。[结论]建立的HPLC方法专属性、重复性良好,更省时、高效,可用于HBs Ag的纯度检测。[Objective]To develop a HPLC for determination of purity of surface antigen of recombinant hepatitis B vaccine(Hansenula polymorpha).[Method]The TSK gel G5000 PW column was used and a series of experiment parameters such as mobile phase,flow rate,column temperature and p H for determination the purity of recombinant hepatitis B vaccine were optimized and the optimal conditions were chosen,the established method was compared with the method by TSK gel G3000 SW chromatographic column.[Result]The established optimal conditions were 0.01 mmol/L PB(Phosphate Buffered)as mobile phase,0.6 mL/min as flow rate,room temperature(20℃)as column temperature,7.0 as pH,and 280 nm as detection wavelength.The retention time of surface antigen(HBsAg)in TSK gel G5000 PW and TSK gel G3000 SW were 8.5 min,17.5 min and the testing time were 20 minutes,40 minutes.[Conclusion]The developed method showed high specificity,repeatability,efficiency,timesaving and it can be used for determination the purity of HBsAg.

关 键 词：高效液相色谱法 乙型肝炎病毒 重组乙型肝炎疫苗 纯度 乙肝表面抗原 

分 类 号：Q657.7[生物学—生物物理学] R392.3[医药卫生—免疫学]