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Vaccimonitor
On-line version ISSN 1025-0298
Abstract
VALDES-CANTERO, Ailen et al. Characterization of polyribosylribitol phosphate by high efficiency size exclusion chromatography with ultraviolet detection. Vaccimonitor [online]. 2022, vol.31, n.1, pp. 25-33. Epub Feb 01, 2022. ISSN 1025-0298.
Haemophilus influenzae type b is an important human pathogen causing some invasive diseases in children less than five years of age. Glycoconjugate vaccines based on polyribosylribitol phosphate have been licensed against this bacterium. Quimi-Hib® is the first and only vaccine against this pathogen using the chemically synthesized polysaccharide. The Active Pharmaceutical Ingredient is produced by the Center for Genetic Engineering and Biotechnology and is obtained from its conjugation to tetanus toxoid. In the present report a characterization of polyribosylribitol phosphate was performed by high performance molecular exclusion chromatography with ultraviolet detection at 215 nm. Three batches were evaluated in the study and the elution profile was determined on a SuperdexTM 75 10/300 GL Increase column with a purity percentage of 77.42 ± 8.97 and an average molecular weight of 7,381 Da ± 210.93. The main impurity present in polyribosylribitol phosphate was dimethylsulfoxide, the solvent used in the activation reaction with N-hydroxysuccinimidyl ester of β-maleimidopropionic acid. Polyribosylribitol phosphate was purified by filtration using a 2,000 Da cut-off Amicon Ultra-15 to a purity of 99.1 % and conjugated to tetanus toxoid. The yield of the conjugation reaction with the purified polysaccharide was 30.0 % ± 1.77 which shows no significant difference with the control which was 33.7%± 3.57 demonstrating that dimethylsulfoxide does not affect the performance of the conjugation reaction.
Keywords : gel chromatography; polysaccharide; tetanus toxoid; dimethylsulfoxide; conjugation; Haemophilus influenzae type b.