Compositional Analysis of Hyaluronan, Chondroitin Sulfate and Dermatan Sulfate : HPLC of Disaccharides Produced from the Glycosaminoglycans by Solvolysis

Abstract

An ion-exchange high-performance liquid chromatography procedure was developed for analysis of mixtures of N-acetyldermosine, N-acetylchondrosine and N-acetylhyalobiuronic acid produced quantitatively by heating dermatan sulfate, chondroitin sulfate and hyaluronan in dimethyl sulfoxide (DMSO) containing 0.1% (v/v) H₂O at 80°C for 48 h. These disaccharides were eluted from a TSK gel SAX column using 0.1 M acetic acid containing KCl, and detected fluorometrically by post column derivatization. The eluate was mixed with 2-cyanoacetamide solution and alkaline solution, and heated at 110°C for 4 min. The resultant compounds were detected fluorometrically (Ex. 335 nm and Em. 390 nm). The usefulness and practicality of the present method were verified by applications to the determination of glycosaminoglycans in tissues.