DETERMINATION OF N-ACETYLNEURAMINIC AND N-GLYCOLYLNEURAMINIC ACIDS IN GLYCOCONJUGATES BY REVERSED-PHASE HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY WITH ULTRAVIOLET DETECTION

A rapid and sensitive method for the determination of sialic acids is described. The measurement is based on isocratic high-performance liquid chromatography, whereby N-acetylneuraminic and N-glycolylneuraminic acids are separated. The total amounts of these acids can be determined after hydrolysis and per-O-benzoylation. The conditions for hydrolysis and derivatization were optimized for measurement of these sialic acids in glycoconjugates. The benzoyl derivatives were chromatographed on a reversed-phase column with 67% (v/v) aqueous acetonitrile and the eluted peaks were monitored by UV detection. The method allows the determination of picomole amounts. the reaction was shown to give linear calibration graphs over the entire range tested. i.e., up to 160 nmol (50 μg) of each of the sialic acids. © 1990.