【摘要】 目的 建立尺寸排阻色谱-多角度激光光散射（size exclusion chromatography-multiangle laser-light scattering,SEC-MALLS）联用测定脑膜炎球菌多糖及多糖蛋白结合物分子量的方法，并进行验证及初步应用，以期进一步提高该品种质量控制水平。方法 建立SEC-MALLS联用方法检测脑膜炎球菌多糖及多糖蛋白结合物分子量，色谱条件：流动相为0.9%氯化钠溶液，色谱分离柱为Shodex OHpak LB-806 HQ(300 mm×8.0 mm,13μm)，流速为0.5 mL/min，柱温为35℃。对方法的系统适用性、准确性、精密性、耐用性进行验证，并用该方法检测不同厂家来源的样品，同时与现行药典质量标准方法进行相关性分析。结果 普鲁兰糖标准品除STD P-100以外，STD P-50 STD P-800重均分子量（Mw）实测值与标示值相对误差和精密度小于5.0%，可作为系统适应性样品；重复性、中间精密度RSD均不大于5.0%。多糖及多糖蛋白结合物在配制完成48 h内稳定性良好。A、C、Y、W135群多糖平均Mw分别为257.5、377.3、399.7、305.5 kDa;A、C群多糖蛋白结合物分别为6 438、23 360 kDa。71批多糖样品中有5批次A、C群结果超趋势，其余批次批内和批间差异均在均值±3 SD的范围以内。SEC-MALLS法与现行药典质量标准方法测定结果除Y群多糖分配系数（K_D）与Mw呈一定的负相关性外，两种方法在其余多糖及多糖蛋白结合物中相关性并不显著。结论 SEC-MALLS方法能直接测定脑膜炎球菌疫苗多糖及多糖蛋白结合物分子量，具有操作简单、快速、准确、能一定程度反映分子结构信息等优点，有利于提升该关键质量参数的质量控制水平。更多还原

【Abstract】 Objective To establish a method for the determination of molecular weight of meningococcal polysaccharide and polysaccharide-protein conjugates by size exclusion chromatography-multiangle laser-light scattering(SEC-MALLS), and to verify and apply the method, so as to improve the quality control level of meningococcal vaccines.Methods A SEC-MALLS method was established to detect the molecular weight of polysaccharide and polysaccharide-protein conjugates of meningococcus. The chromatographic conditions were as follows: 0. 9% sodium chloride solution as the mobile phase; Shodex OHpak LB-806 HQ(300 mm × 8. 0 mm, 13 μm) as the column; the flow rate at 0. 5 mL/min; the column temperature at 35 ℃. The systematic applicability, accuracy, precision and durability of the method were verified, and the samples from different manufacturers were detected by this method. In addition, the correlation between SEC-MALLS and the existing pharmacopoeia quality standard methods was analyzed.Results Except for STD P-100, the relative error and precision between the measured and marked values of weight-average molecular weight(Mw) of STD P-50 to STD P-800 were less than 5. 0%, indicating that the pullulan standard could be used as the systematic adaptive sample. The RSDs of reproducibility and intermediate precision were not more than 5. 0%. The polysaccharide and polysaccharide-protein conjugates had good stability within 48 h after preparation. The average Mw of polysaccharides from group A, C, Y and W135 were 257. 5, 377. 3, 399. 7and 305. 5 kDa, while the polysaccharide-protein conjugates of group A and group C were 6 438 and 23 360 kDa, respectively. Among the 71 batches of polysaccharide samples, the results of group A and group C in five batches exceeded the trend, and the inner-and inter-batch differences of other batches were within the range of mean ± 3 SD. Except the KDvalue of polysaccharide in group Y had a certain negative correlation with Mw, there was no significant correlation between the two methods in the remaining polysaccharide and polysaccharide-protein conjugates.Conclusion SEC-MALLS can directly determine the molecular weight of polysaccharide and polysaccharide-protein conjugates of meningococcal vaccine with the advantages of fast, accurate and simple operation, and can reflect the molecular structure information to a certain extent,which is conducive to improve the quality control of this key quality parameter.更多还原