Kinetic investigation of the degradation of hyaluronan by hyaluronidase using gel permeation chromatography.

A gel permeation chromatography (GPC) system for the analysis of hyaluronan was set up and calibrated by means of the universal calibration method. Using GPC, a kinetic assay of the action of hyaluronidase on hyaluronan has been developed. Applying Michaelis-Menten theory and the direct linear plot we have estimated the kinetic constants of the enzyme. By evaluating the decrease of the various molecular mass averages during the enzymatic reactions, we have given a demonstration of the at random degradation of hyaluronan by hyaluronidase in the initial stage of the reaction.