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  • Title: Monosaccharide analysis of glycoconjugates by anion exchange chromatography with pulsed amperometric detection.
    Author: Hardy MR, Townsend RR, Lee YC.
    Journal: Anal Biochem; 1988 Apr; 170(1):54-62. PubMed ID: 3389518.
    Abstract:
    The method of anion exchange chromatography followed by pulsed amperometric detection (AE-PAD; Johnson, D. C., and Polta, T. Z. (1986) Chromatogr. Forum 1, 37-44) has been applied to the compositional analysis of glycoconjugates. Using 22 mM NaOH as a column effluent, underivatized fucose, galactosamine, glucosamine, galactose, glucose, and mannose were readily separated in 15 min at a flow rate of 1 ml/min. The limit of quantification of the monosaccharides was better than 100 pmol (signal to noise ratio 184:1). AE-PAD was employed to quantify the monosaccharides of several glycoproteins, glycopeptides, and oligosaccharides after hydrolysis with 2 M trifluoroacetic acid. Both neutral and amino sugars could be rapidly estimated in a single chromatographic step using AE-PAD. Complete release of N-acetylglucosamine required more vigorous hydrolysis conditions (Lee, Y. C. (1972) in Methods in Enzymology (Ginsburg, V., Ed.), Vol. 28, pp. 63-73, Academic Press, New York). In both glycopeptides and oligosaccharides, approximately one less residue of Man than predicted was determined. Both AE-PAD and liquid chromatographic analysis of borate-monosaccharide complexes with fluorometric detection (Mikami, H., and Ishida, Y. (1983) Bunseki Kagaku 32, E207-E210) gave similar quantification of mannose and other sugars. The capability of rapid, sensitive quantification of underivitized monosaccharides should facilitate structural analysis of glycoconjugates.
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