ASCORBIC ACID INTERFERENCE WITH ESTIMATION OF BETA-GLUCOSAMINIDASE ACTIVITY

Study of the interaction of ascorbic acid with β-N-acetylglucosaminidase obtained from cultured human dermal fibroblasts demonstrated that ascorbate is a noncompetitive inhibitor of the enzyme. When the substrate, p-nitrophenyl N-acetyl-β-glucosaminide, was incubated in the presence of ascorbate in concentrations above 10 μg/ml with no enzyme present, a chromogen with maximal absorbance at 376 mμ was released. The p-nitrophenol released by the enzyme as well as pure p-nitrophenol absorbs light maximally at 400 mμ. Nonenzymic cleavage of the glycosidic linkage of the substrate by ascorbic acid is postulated. Preincubation with active or heat-denatured enzyme reduced the ability of ascorbate to hydrolyze the substrate but did not influence its ability to inhibit the enzyme at the concentrations studied. © 1968.