Analysis of mucin-derived oligosaccharides by medium-pressure gel-permeation and amino-plate thin-layer chromatography conducted in conjunction

Oligosaccharides present in mucin were labeled by reduction with NaB3H4 and separated by gel-permeation chromatography with a Toyopearl HW-40S column using 0.1 M pyridine acetate, pH 5.0, as the solvent. Each fraction was further analyzed by thin-layer chromatography (TLC) on a Funagel AMP plate, a glass plate precoated with 3-aminopropyl-bonded silica. Acetonitrile/10 mM triethylamine acetate (3/2, by volume) served as the solvent. The sites of oligosaccharides on the TLC plate could be determined according to size, anionic charge, and sugar composition. They could thus be "mapped" on the plate. In this manner, the distribution of oligosaccharides on bovine submaxillary mucin and rat gastric mucin was determined. Each radiolabeled oligosaccharide in newly synthesized rat gastric mucin, metabolically labeled with 14C]glucosamine or 35S]sulfate, was also identified by this method.