Li Y.-S.,Beijingvigoo BiologicalCo. |
Liu M.-Y.,Beijingvigoo BiologicalCo. |
Chen J.,Beijingvigoo BiologicalCo. |
Lin H.-T.,Beijingvigoo BiologicalCo. |
Wang P.,Beijingvigoo BiologicalCo.
Chinese Journal of Biologicals | Year: 2013
Objective: To determine the galactose and glucose contents in groups W135 and Y meningococcal polysaccharide by high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD), and verify and preliminarily apply the method. Methods: The samples of pooled bulks of polysaccharide-protein conjugate was hydrolyzed with hydrochloric acid and loaded onto CarboPac PA10 column, then eluted by sodium hydroxide-sodium acetate gradient elution and determined by pulse amperometric detection mold. The condition for hydrolysis was optimized, and HPAEC-PAD method was verified for specificity, precision and accuracy, and preliminarily applied. Results: The condition for hydrolysis was optimized as treatment with 2 mol/L hydrochloric acid at 85°C for 1 h. Tetanus toxoid (TT) as a carrier showed no interference to detection result. The RSDs of results of reproducibility test as well as intra- and inter-assays were less than 5%. In accuracy test, the recovery rates of glucose and galactose were 90% ∼ 110%. The polysaccharide content in bulk of conjugate determined by HPAEC-PAD was approximately equal to that by routine resorcinol assay. The contents of groups W135 and Y polysaccharides in final product met the Requirements for Groups A, C, W135 and Y Meningococcal Conjugate Vaccine (Draft) (not less than 4 μg/dose). Conclusion: HPAEC-PAD method showed high specificity, precision and accuracy, which might be used for determination of groups W135 and Y meningococcal polysaccharide contents in groups A, C, W135 and Y meningococcal conjugate vaccine.