[A sensitive method for the analysis of the glycosaminoglycan distribution pattern, applied to 9 intervertebral discs...

A method has been established for analysing the glycosaminoglycan distribution pattern in small specimens from intervertebral discs and other connective tissue materials. The procedure is based on the consecutive digestion by the specific enzymes hyaluronate lyase, chondroitin sulfate lyase AC and ABC. The quantitative determination of the resulting disaccharides was achieved by high performance liquid chromatography with a detection limit of less than 0.1 nmmol. The undigested heparan- and keratansulfate were separated from each other by thin layer chromatography and quantified by the colorimetric determination of uronic acid and hexosamine, respectively. Correlation of distribution patterns obtained by this method with those obtained by ion exchange chromatography (Dowex 1 X 2) is not possible without the additional enzymatic characterization of the ion exchange chromatographic fractions. The method was applied to 9 discs of one human spine (anulus fibrosus and nucleus pulposus separately) with the following results: 1) there were no systematic changes to be seen in the distribution patterns with respect to the location of the discs in the spine, and 2) the galactosaminoglycan/keratansulfate ratio was higher in the anulus fibrosus than in the nucleus pulposus.