The micro-assay of the disaccharide isomers of chondroitin sulphate

Abstract

Connective tissue extracts containing the 4- and 6-sulphated isomers of chondroitin sulphate can be measured at the microgram level (approximately 10 microgram) using colorimetric assays. The chondroitin sulphates are depolymerised to disaccharides using chondroitin ABC lyase (EC 4.2.2.4). The 4- and 6-sulphated disaccharides after treatment with acid periodate are determined using a thiobarbituric acid procedure. An alkaline borate p-dimethylaminobenzaldehyde reaction is used to measure 6-sulphated disaccharide, the 4-sulphated disaccharide not forming a chromogen. Unsulphated disaccharide causes interference in both assays and if present is separated prior to assay using paper electrophoresis fractionation.