Simplified and improved methylation analysis of saccharides, using a modified procedure and thin-layer chromatography

Abstract Methylation analysis used in determining the positions of glycosidic linkages in oligo- and poly-saccharides has been greatly simplified by eliminating two of the usual reactions, reduction and acetylation of the methylated monosaccharide residues. The saccharides are methylated by the Hakomori procedure, followed by acid hydrolysis and analysis by thin-layer chromatography (TLC). The procedure is illustrated by analysis of four known disaccharides, cyclomaltohexaose, and eleven polysaccharides. 2,3,4,6- Tetra -O- methyl- d -glucose , 2,3,4,6- tetra -O- methyl- d -galactose , and 2,3,4,6- tetra -O- methyl- d -mannose , three isomeric tri -O- methyl- d -glucoses, tri -O- methyl- d -mannoses , and tri -O- methyl- d -galactoses , three isomeric di -O- methyl- d -glucoses , and mono -O- methyl- d -glucose are resolved by the TLC system. The O -methylated sugars can be quantitated directly on the TLC plate by using imaging densitometry. The new procedure permits any laboratory to carry out the analyses with a minimum of expense and without the need of dedicated, elaborate laboratory instrumentation. The procedure is sufficiently micro that it can be carried out on 1 mg of sample.