An extracellular microbial polysaccharide composed of 2-acetamido-2-deoxy- D-glucose and 2-acetamido-2-deoxy- D-glucuronic acid: Radiochemical and gas-chromatographic analysis of the products of methanolysis

Abstract

When the polysaccharide from the black yeast NRRL Y-6272, composed of 2-acetamido-2-deoxy- D-glucose ( 1) and 2-acetamido-2-deoxy- D-glucuronic acid ( 2), is hydrolyzed, extensive humin formation occurs by decomposition of component residues, especially the hexosaminuronic acid. Methanolysis avoids this decomposition by forming stable methyl glycosides amenable to quantitation by both radiochromatographic techniques and gas chromatography. Unlike hydrolysis, which results in incomplete depolymerization, refluxing methanol-HCl ( M, 16–24 h) completely depolymerizes polysaccharide Y-6272 to the methyl glycosides of its component sugars. Use of 14C-methanol-HCl allows quantitation of 1 and 2 by counting the individual 14C-methyl glycosides after separation by paper chromatography. As the methyl glycosides derived from the hexosaminuronic acid in polysaccharide Y-6272 consist of both a methyl ester and a lactone, for quantitation it was necessary to convert these two glycoside forms into a common derivative of known 14C-methyl content by treatment with mild alkali. Methanolysis by using radioisotopes affords an extremely valuable method for detecting and quantitating amino sugars in polysaccharides; it is rapid and sensitive and it should be especially applicable for analyzing other polysaccharides and proteins that contain constituents labile to normal hydrolytic conditions.