冻干b型流感嗜血杆菌结合物疫苗中磷含量测定方法建立及验证  

Development and Verification of Phosphorus Assay in Haemophilus Influenzae Type B Conjugate Vaccine

作　　者：张晓芸 孙星宇 丁逸梅[1,2] ZHANG Xiaoyun;SUN Xingyu;DING Yimei(Jiangsu Provincial Institute of Material Medica,Nanjing Tech University,Nanjing 210009,China;School of Pharmaceutical Sciences,Nanjing Tech University,Nanjing 211816,China)

机构地区：[1]南京工业大学江苏省药物研究所,南京210009 [2]南京工业大学药学院,南京211816

出　　处：《药学与临床研究》2024年第2期136-139,共4页Pharmaceutical and Clinical Research

摘　　要：目的:建立冻干b型流感嗜血杆菌(Hib)结合疫苗磷含量检测的超滤法联合紫外分光光度法,并对该方法进行验证及初步应用。方法:用去离子水将样品溶解后超滤,收集超滤管中Hib多糖,采用等体积比硫酸高氯酸消解样品,使用钼酸铵在酸性条件下与磷形成磷钼酸,与维生素C生成蓝色化合物,在825 nm下有最大吸收。对该方法进行线性、精密度、稳定性、准确度验证。用建立的方法检测3批次商业化冻干Hib结合疫苗磷含量。结果:标准磷在质量浓度1~10μg·mL^(-1)内线性关系良好(r=1.000),线性方程为Y=8.878×10^(-2)X+2.402×10^(-3),方法在低、中、高浓度水平下回收率均值分别为在97.35%、96.65%、97.42%(n=3),RSD分别为2.55%、0.95%、0.95%;重现性试验的RSD为2.81%;中间精密度RSD 2.14%;磷标准品衍生物溶液室温放置1 h稳定,RSD为0.39%。结论:该方法结果准确、可靠,可用于冻干b型流感嗜血杆菌结合疫苗磷含量检测,同时也为其他多糖蛋白结合疫苗磷含量测定提供了思路。Objective:To establish and verify an ultrafiltration method combined with ultraviolet spectrophotometry for detection of phosphorus contents in Haemophilus influenzae type b(Hib)conjugate vaccines.Methods:The sample was dissolved with deionized water and then ultrafiltrated.Hib polysaccharide in the ultrafiltration tube was collected.The sample was digested with perchloric acid and sulfuric acid(1∶1).Ammonium molybdate was used to form phosphomolybdic acid with phosphorus under acidic conditions,and then blue compound was formed with vitamin C with maximum absorption at 825 nm.The linearity,precision,stability,and accuracy of this method were verified.The established method was used to detect the phosphorus contents of three commercial batches of freeze-dried Hib conjugate vaccines.Results:The phosphorus concentrations at a range of 1-10μg·mL^(-1)showed good linear relationship,the coefficient r was 1.000,the linear regression equation was Y=8.878×10^(-2)X+2.402×10^(-3).The average recovery at low,medium,and high concentration levels were 97.35%,96.65%and 97.42%(n=3),respectively,with RSD of 2.55%,0.95%and 0.95%.The RSD of the reproducibility test was 2.81%,and the intermediate precision RSD was 2.14%.The derivative solutions of phosphorus standard were stable at room temperature for 1 hour with RSD at 0.39%.Conclusion:This method has accurate and reliable results,and can be used for the detection of phosphorus contents in freeze-dried Haemophilus influenzae type b conjugate vaccines.It also provides clues for the determination of phosphorus contents in other polysaccharide protein conjugate vaccines.

关 键 词：超滤法 冻干b型流感嗜血杆菌结合疫苗 磷 抗坏血酸 

分 类 号：R927.2[医药卫生—药学]