SEPARATION OF PYRIDYLAMINO OLIGOSACCHARIDES BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY ON AN AMINE-BEARING SILICA COLUMN

Several neutral and sialylated pyridylamino (PA) oligosaccharides were separated on an amine-bearing silica column, PALPAK Type N. Neutral PA-oligosaccharides were fractionated according to the number of sugar residues by amine adsorption. Sialylated PA-oligosaccharides were separated by ion-exchange chromatography. An amine-bearing column was eluted with a mobile phase consisting of acetonitrile and water containing acetic acid titrated to pH 7.3 with triethylamine (TEAA buffer). A mixture of neutral and sialylated PA-oligosaccharides was separated by double-mode HPLC with a solvent program of decreasing acetonitrile concentration (70 to 51%) with a constant TEAA buffer concentration (0.03 M), and then an increasing TEAA buffer concentration (0.03 to 0.49 M) with a constant acetonitrile concentration (51%). This HPLC technique was applied to the comparative oligosaccharide pattern analysis of human Asn-linked oligosaccharides of normal and pathological IgG by hydrazinolysis. The result indicated clearly that oligosaccharides of IgG myeloma proteins have different core structures and ratios of sialic acid than those of IgG normal proteins. (C) 1994 Academic Press, Inc.