PREPARATION OF OLIGOSACCHARIDES FROM THE ENZYMATIC HYDROLYSATES OF KONJAC GLUCOMANNAN BY LOW-PRESSURE LIQUID CHROMATOGRAPHY

摘要

The present report describes a method for separation of oligosaccharides including their isomers from the hydrolysates of Konjac glucomannan by the low pressure liquid chromatography with the activated charcoal column.rnThe procedures of experiment were briefly described as folio whig. The activated charcoal was boiled in the 20% acetic acid and was then washed until neutrality with stilled water. The activated charcoal powder was packed into a column (10 × 200 mm) which was connected to a low-pressure pump (Kontron LC 410). After the column was washed with 200 ml of stilled water, the solution (1 ml) of the enzymatic hydrolysate, containing 0.2 g as total sugar content was introduced in the column head which was washed with 500 ml of water to remove salts and mono-saccharide. The oligosaccharides were eluted successively with 800 ml each of 5%, 7.5%, 10% and 15% ethanol in the water with flow-rate of 1.5 ml/min. The eluent was fractionated to 10 ml in each tube using a fraction collector. The sugar composition in the tubes was examined by modified thin-layer chromatography-orchiol-sulfuric acid. The developed solvent system of acetonitrile-water (7:2) was employed with multi-step developments to allow the separation of di-, tri-saccharides and isomers. The each fraction of oligosaccharide was identified by methylation analysis, fast atom bombardment mass spectrometry.