论文部分内容阅读
目的采用高效阴离子交换色谱-积分脉冲安培法(high performance anion exchange chromatography with pulsed amperometric detector,HPAEC-PAD)测定四价脑膜炎球菌多糖疫苗中A、C、Y、W135群多糖含量,并对该方法进行验证。方法用三氟乙酸对四价脑膜炎球菌结合疫苗进行水解,得到各群多糖的水解产物:唾液酸、葡萄糖、半乳糖、6-磷酸葡萄糖胺(ManN-6-P),采用HPAEC-PAD法,CarboPac PA10分离柱分别测定缓冲提取液中的总唾液酸、葡萄糖、半乳糖、ManN-6-P的含量,利用归一化的峰面积计算相应唾液酸系数,再计算C群脑膜炎球菌唾液酸含量。对HPAEC-PAD法进行验证,并与传统火箭电泳法检测结果进行比较。结果四价脑膜炎球菌结合疫苗水解后得到的4群单糖均呈良好线性,分离度和峰形均良好;HPAEC-PAD法检测4群糖的日内和中间精密性的RSD均小于5%;各群多糖峰面积的回收率均在80%~120%之间;3批四价脑膜炎球菌多糖疫苗水解后,经HPAEC-PAD法测定的各群多糖含量与传统火箭电泳法测定结果相当。结论 HPAEC-PAD法检测四价脑膜炎球菌多糖疫苗中的A、C、Y、W135群多糖含量准确、简便、快速、可靠,对控制该疫苗的质量具有重要意义。
Objective To determine the polysaccharide content of A, C, Y and W135 in quadrivalent meningococcal polysaccharide vaccine by high performance anion exchange chromatography with pulsed amperometric detector (HPAEC-PAD) authenticating. Methods Tetravalent meningococcal conjugate vaccine was hydrolyzed with trifluoroacetic acid to obtain the hydrolyzate of polysaccharides: sialic acid, glucose, galactose and glucosamine 6-phosphate (ManN-6-P). HPAEC-PAD , CarboPac PA10 column were used to determine the content of total sialic acid, glucose, galactose and ManN-6-P in the buffer extract. The corresponding sialic acid coefficient was calculated by the normalized peak area, and then the meningococcus group C saliva Acid content. The HPAEC-PAD method was validated and compared with the traditional rocket electrophoresis test results. Results The four groups of monosaccharides obtained after the tetravalent meningococcal conjugate vaccine hydrolyzed showed a good linearity with good resolution and peak shape. The intra-and inter-precision RSDs of 4 groups of carbohydrates detected by HPAEC-PAD were all less than 5% The recoveries of polysaccharide peak area of all groups were in the range of 80% ~ 120%. After three batches of tetravalent meningococcal polysaccharide vaccine were hydrolyzed, the content of polysaccharide in each group determined by HPAEC-PAD method was equivalent to that of traditional rocket electrophoresis. Conclusion The detection of polysaccharides of A, C, Y and W135 in the tetravalent meningococcal polysaccharide vaccine by HPAEC-PAD method is accurate, simple, rapid and reliable, and it is important to control the quality of the vaccine.