疫苗中铝含量紫外分光光度测定法的建立及验证  

Development and verification of a novel method for determination of aluminium content in vaccine by ultraviolet spectrophotometry

作　　者：王平 张学峰 陈美丽 冯帆 邵帅 刘梅影 WANG Ping;ZHANG Xue-feng;CHEN Mei-li;FENG Fan;SHAO Shuai;LIU Mei-ying(National Vaccine and Serum Institute,Beijing 101111,China)

机构地区：[1]国药中生生物技术研究院有限公司,北京101111 [2]北京生物制品研究所有限责任公司,北京100176

出　　处：《中国生物制品学杂志》2021年第9期1094-1099,共6页Chinese Journal of Biologicals

摘　　要：目的建立疫苗中铝含量的紫外分光光度测定法,并进行验证。方法采用Tiron试剂衍生氯化铝,紫外分光光度法进行检测,筛选方法的最佳检测波长,并绘制标准曲线。同时采用氢氧化铝标准溶液优化样品处理方法中的干烤温度(120、140、160、180、200、220℃)及1 mol/L盐酸加入量(10、20、50、100、200、300、400和500μL)。对建立的方法进行准确性、精密性及专属性验证。应用建立方法检测含铝佐剂疫苗中铝含量,并与传统滴定法进行比较。结果确定衍生物的检测波长为310 nm。氯化铝标准品在1~10μg/mL范围内,与A310呈良好的线性关系,回归方程为Y=0.009 41 X+0.009 97,R=0.998 29。样品处理最适干烤温度为200℃,1 mol/L盐酸最适加入量为50μL。氯化铝的加样回收率在100.36%~108.40%之间;重复性及中间精密性RSD均<6%;非佐剂疫苗成分、硫柳汞溶液和磷酸缓冲盐溶液对测定无干扰。氢氧化铝佐剂、百白破疫苗与实验用乙型肝炎疫苗铝含量紫外分光光度法与传统滴定法测定结果差异较小;实验用流感病毒裂解疫苗(佐剂)单支检测9支均值与滴定法结果一致。结论紫外分光光度法具有良好的准确性和精密性,且操作简便、安全,避免了人为因素干扰,实现了单支疫苗的铝含量检测,为研发过程中铝佐剂制备工艺、抗原吸附工艺和分装工艺优化提供了有效的监测手段。Objective To develop and verify a method for determination of aluminium content in vaccine containing aluminium adjuvant by ultraviolet spectrophotometry.Methods Aluminium chloride was derived with Tiron and determined by ultraviolet spectrophotometry.The detection wavelength was optimized,and a standard curve was plotted.Meanwhile,the temperature for drying(120,140,160,180,200 and 220 ℃) and the added volume of 1 mol/L hydrochloric acid(10,20,50,100,200,300,400 and 500 μL) in pretreatment of samples were optimized with standard aluminum hydroxide solution.The developed method was verified for accuracy,precision and specificity.The aluminium contents in vaccines containing aluminium adjuvant were determined by the developed method,and the results were compared with those by traditional titration method.Results The detection wavelength of the derivative was determined as 310 nm.The standard aluminium chloride at concentrations of 1~10 μg/mL showed good linear relationship to the A310 value,with a regression equation of Y = 0.009 41 X + 0.009 97(r = 0.998 29).The optimal temperature for drying in pretreatment of samples was 200 ℃,while the optimal added volume of 1 mol/L hydrochloric acid was 50 μL.The recovery rate of sample in aluminum chloride was 100.36%~108.40%.Both the RSDs of test results for reproducibility and intermediate precision were less than 6%.Non-adjuvant vaccine components,thiomersal solution and phosphate buffer solution showed no interference to the determination result.There was little difference between the aluminum contents in aluminum hydroxide adjuvant,DPT and experimental hepatitis B vaccine determined by ultraviolet spectrophotometry and by traditional titration method.The mean of determination results of nine single dose influenza vaccine was consistent with that by traditional titration method.Conclusion Ultraviolet spectrophotometry showed high accuracy and precision,which was easy to handle and safe,overcame the interference of human factors and realized the detection of alumin

关 键 词：紫外分光光度法 铝佐剂 

分 类 号：Q599[生物学—生物化学]