Development and verification of a high-performance liquid chromatography method for determination of sucrose content in oral hexavalent reassortant live attenuated rotavirus vaccine(Vero cells)
10.13200/j.cnki.cjb.004341
- VernacularTitle:口服六价重配轮状病毒减毒活疫苗（Vero细胞）中蔗糖含量高效液相色谱检测方法的建立及验证
- Author:
HE Liang
- Publication Type:Journal Article
- Keywords:
Oral hexavalent reassortant live attenuated rotavirus vaccine(Vero cells);
Sucrose content;
High-performance liquid chromatography(HPLC)
- From:
Chinese Journal of Biologicals
2024;37(11):1367-1372
- CountryChina
- Language:Chinese
-
Abstract:
Objective To develop and verify a high-performance liquid chromatography(HPLC) method for the determination of sucrose content in oral hexavalent reassortant live attenuated rotavirus vaccines(Vero cells), in order to provide a reliable method for the determination of sucrose content in vaccines with sucrose as an excipient. Methods A HPLC method for the determination of sucrose content in oral hexavalent reassortant live attenuated rotavirus vaccines(Vero cells) was developed. The chromatographic column was Luna?5 μm NH2100?(LC Colum 250 mm × 4. 6 mm), the differential detector temperature was 30 ℃, the mobile phase was acetonitrile, the flow rate was 0. 5 mL/min, the injection volume was 10 μL, and the column temperature was 30 ℃. The system suitability, specificity, linear range, precision, accuracy and robustness of the method were verified. At the same time, the consistency of the results of HPLC and polarimetry method for determining the sucrose content in the test sample was analyzed. Results The separation of lactose and sucrose in system suitability test solution was good, and the lactose did not interfere with the determination of sucrose content. The citric acid, sodium citrate, zinc chloride and calcium chloride showed no interference to the determination of sucrose content. The sucrose concentration at the range of 6-14 mg/mL showed a good linear relationship with the peak area, and the linear equation was: y = 142 189 x +2 182. 4, with the correlation coefficient(r) of 0. 999 9. The RSDs of reproducibility and intermediate precision verification were 0. 55% and 0. 49% respectively. The average recovery rates of samples with the concentration of 80%, 100%, and 120% were all between 99% and 101%. Minor changes in flow rate, column temperature, mobile phase ratio and detector temperature, or replacement of chromatographic columns of different batches had no effect on the test results, and the relative deviations(RDs) with the sucrose content measured under the original conditions were all less than 1%. The RDs of sucrose content in multiple batches of test samples determined by HPLC and polarimetry were all less than 1%. Conclusion The developed HPLC method has good specificity, precision, accuracy and robustness, and can be used for the determination of sucrose content in oral hexavalent reassortant live attenuated rotavirus vaccine(Vero cells).
