Applicability of Determination Methods for Groups Generated in Cyanate Esterification of Streptococcus Pneumoniae Capsular Polysaccharides

OBJECTIVE  To explore the applicabilities of previously reported quantitative determination methods for cyanate ester, imidocarbonate, and carbamate groups in the cyanate esterification process of Streptococcus pneumoniae capsular polysaccharides(CPS).

METHODS For the content of cyanate ester in activated polysaccharide, the perchloric acid titration method and the dimethylbarbiturate pyridine method were used. For the determination of imidocarbonate content, the hydrochloric acid hydrolysis method was used to hydrolyze imine carbonate into ammonium ions, which was measured by spectrophotometry. For the content of carbamate, enzyme-linked immunosorbent assay was used.

RESULTS Due to the insolubility of the Streptococcus pneumoniae CPS in acetic acid, the determination of cyanate ester content in the activated Streptococcus pneumoniae CPS by using perchloric acid titration required optimization of the reaction solvent. On the other hand, the cyanate ester content of activated 23F Streptococcus pneumoniae CPS determined by the dimethylbarbiturate pyridine method was (0.038±0.004)%. The imidocarbonate content of 23F Streptococcus pneumoniae CPS determined by hydrochloric acid hydrolysis method was (2.20±0.03)%. The aminoformate content of activated polysaccharide determined by enzyme-linked immunosorbent assay exceeded the detection range. This method was thought to be not suitable for the activated pneumococcal polysaccharides.

CONCLUSION The dimethylbarbiturate pyridine method can be used to determine the cyanate ester content in cyanated polysaccharides with an activation degree of (0.038±0.004)%. The hydrochloric acid hydrolysis method can accurately determine the imidocarbonate content of activated polysaccharides. Enzyme-linked immunosorbent assay is found to be not suitable for the determination of carbamate content in activated polysaccharides.