Assay of keratan sulfate as anion-exchanger bound hexose

An assay for keratan sulfate in papain digests of human intervertebral disc and other tissues has been developed. The digest is applied to the acetate form of a tertiary amine acrylic anion-exchange resin, the oligosaccharide hexose is removed by washing the resin with 0.2 M sodium acetate buffer pH 5.0, then the keratan sulfate is eluted quantitatively with 1.0 M pyridinium sulfate pH 2.5 and assayed for hexose by the anthrone reaction. The keratan sulfate content of human intervertebral disc tissues ranged from 7 to 78 mumole galactose equivalents/g fresh weight; the root mean square error was 2 mumole/g; 10-25 mg of tissue were required. The separation of oligosaccharides from keratan sulfate was confirmed by gel permeation chromatography, sugar composition, ester sulfate analysis, and nuclear magnetic resonance.