Quantitative Determination of Saccharides in Dietary Glyconutritional Products by Anion-Exchange Liquid Chromatography with Integrated Pulsed Amperometric Detection

Abstract

A new technique for the assay of carbohydrates is described in which separation and quantification of neutral saccharides, aminosaccharides, glycuronic acids, and disaccharides may be accomplished in less than 50 min of total run time. This method involves optimized anion-exchange liquid chromatography coupled with integrated pulse amperometric detection. Complex carbohydrates from various sources, including dietary supplements, were hydrolyzed in a dilute solution of trifluoroacetic acid, freeze-dried, and reconstituted in water containing 2-deoxygalactose as the internal standard. The solution was filtered and separated on CarboPac PA20 column. The eluted saccharides were detected by oxidation on a gold electrode with quadruple-pulsed integrated amperometry. The calibration plots for the saccharides were linear with an average correlation coefficient of 0.999. Method precision regarding peak retention time and resolution used in the peak identifications was verified. With this method, previously difficult-to-separate saccharides, such as galactosamine, glucosamine, and N-acetylglucosamine, were successfully resolved from the neutral saccharides rhamnose, arabinose, and galactose. Mannose was also resolved from xylose, and de-acetylation of aminosaccharides prior to separation was not necessary. This technique provides an accurate and efficient means to assay carbohydrates in dietary supplements, which new federal regulations will soon mandate.