A high-performance liquid chromatography approach for isolation and sequencing of chondroitin sulfate oligosaccharides

✍ Scribed by Stephen R. Delaney; H.Edward Conrad; Janet H. Glaser

• Publisher: Elsevier Science
• Year: 1980
• Tongue: English
• Weight: 848 KB
• Volume: 108
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(80)90689-2

✦ Synopsis

Chondroitin-6S0, from whale cartilage and chondroitin-6-SO, from shark cartilage were digested with hyaluronidase and the resulting tetra-, hexa-, octa-, and decasaccharides were purified in two stages. In Stage I the oligosaccharides from each digest were separated according to size by gel filtration on Sephadex G-50 and by subsequent preparative paper chromatography of their Na-[3H]borohydride-reduced derivatives. The Stage I oligosaccharides were mixtures of oligosaccharides having the same degrees of polymerization but differing in their proportions of 4-sulfated and 6-sulfated N-acetyl-n-galactosamine residues. The Stage I 3H-labeled oligosaccharides were further purified by high-performance liquid chromatography on a Partisil-10 SAX anion-exchange column to give relatively homogeneous Stage II products. The most prominent Stage II oligosaccharide obtained from each of the Stage I mixtures was assayed for the percentages of the 4-and 6-sulfated disaccharides at its reducing terminal and its nonreducing terminal, and in its internal positions using a newly developed high-performance liquid chromatography procedure for analysis of the saturated and unsaturated disaccharides released when the purified oligosaccharide is treated with bacterial chondroitinase. The tetra-and hexasaccharides obtained from the two starting materials were completely 4-sulfated (whale cartilage chondroitin SO*) or 6-sulfated (shark cartilage chondroitin SO,), but the octa-and decasaccharides, which appeared to be chromatographically homogeneous, contained both 4-and 6-sulfatedN-acetyl-D-galactosamine residues.