A群C群脑膜炎球菌多糖疫苗中乳糖含量HPLC检测方法的验证及应用

目的验证A群C群脑膜炎球菌多糖疫苗中乳糖含量的检测方法。方法采用色谱柱XBridge Amide(4. 6 mm×150 cm,3. 5μm),流动相为乙腈、水、三乙胺(75∶25∶0. 1)的混合液,流速为1 mL/min,柱温为40℃,示差检测器温度为40℃,进样量为50μL。同时验证该方法的系统适用性、特异性、线性范围、精密度、稳定性、准确度及定量限。采用该方法对3个厂家生产的30批A群C群脑膜炎球菌多糖疫苗中乳糖含量进行检测。结果乳糖对照品色谱峰的理论塔板数为8 123,与其他色谱峰的分离度> 1. 5;乳糖对照品及供试品溶液图谱于相应保留时间处可见乳糖峰,阴性对照溶液未见该色谱峰;乳糖浓度在0. 050 03～0. 650 4 mg/mL范围内与峰面积呈良好的线性关系,回归方程为Y=3. 484 3×10⁵X-7. 378 4×10²,R²=0. 999 97。批号为Y201409089-1、Y201501002和201410050的3批供试品溶液分别测定6次结果的RSD分别为0. 71%、1. 01%和...

Validation and application of a HPLC method for determination of lactose content in groups A and C meningococcal polysaccharide vaccine

Objective To validate a method for determination of lactose content in groups A and C meningococcal polysaccharide vaccine. Methods HPLC was used for determination of lactose content in groups A and C meningococcal polysaccharide vaccine. A XBridge Amide(4. 6 mm × 150 cm,3. 5 μm)column was used as the stationary phase,while the mixture of acetonitrile,water and triethylamine(75 : 25 : 0. 1)as the mobile phase. The flow rate was set up as 1 mL/min,while the column temperature as 40 ℃,the differential detector temperature as 40 ℃,and the sample loading as 50 μL.The method was validated for system suitability,specificity,linear range,precision and accuracy. Results The theoretical plate number of chromatographic peak of lactose control was 8 123,while the separation degree with other chromatographic peaks was more than 1. 5. Lactose peaks were observed at the corresponding retention time sites on chromatographic profiles of lactose control and test samples,while no such a peak on the profile of negative control. Lactose at a concentration range of 0. 050 03 ~ 0. 650 4 mg/mL showed good linear relationship to the peak area,of which the regression equation was Y = 3. 484 3 × 10^5 X-7. 378 4 × 10^2,with a R^2 value of 0. 999 97. The RSDs of six test results of samples with with lot numbers of Y201409089-1,Y201501002 and 201410050 were 0. 71%,1. 01% and 1. 69%,while those determined at 12 time points were 0. 25%,0. 38% and 0. 58%,respectively. The recovery rates of lactose in the three batches of samples were 101. 67%,101. 93% and 99. 43%,with RSDs of 1. 37%,1. 20% and 1. 38%,respectively. The lower limit of quantitation of lactose was 0. 4 μg. The lactose contents in 30 batches of groups A and C meningococcal polysaccharide vaccine were 8. 29 ~ 11. 05 mg/mL. Conclusion The developed method was precise,stable and accurate,which might be used for the determination of lactose content in groups A and C meningococcal polysaccharide vaccine.