Instability and depolymerization of the exopolysaccharide produced by Haemophilus Influenzae type B during fermentation

Abstract

Haemophilus influenzae type b (Hib) is a pathogenic bacterium and the major cause of sequalae and deaths among infants due to meningitis. Hib vaccine is constituted of the exopolysaccharide from the cell capsule and conjugated to a carrier protein. The production of the polysaccharide is complicated due to low yields of production and product recovery in the downstream. In this work, the profiles of molecular mass throughout fermentation were investigated. The molecular mass decreased along fermentation time, despite the increase in concentration. The mechanisms of hydrolysis were investigated, with three possibilities considered: alkaline hydrolysis caused by the solution used to maintain pH of the fermentation; presence of hydrolytic activity from the fermentation metabolites and spontaneous hydrolysisat the temperature and pH was evaluated in the conditions of the fermentation and with purified polysaccharide. The results have suggested that there is not significant influence of the strength of the alkali solution used for pH control neither the presence of hydrolases in the supernatant; however spontaneous hydrolysis was verified in a temperature dependent manner and confirmed with purified polysaccharide.