β-GLUCURONIDASE HYDROLYSIS OF URINARY CORTICOSTEROID CONJUGATES: THE EFFECT OF SALICYLATE GLUCURONOSIDE AS A COMPETING SUBSTRATE AND THE EFFECT OF ENZYME INACTIVATION*

Abstract

One microcurie of 4-C14-cortisol was administered intravenously to 3 normal adults receiving 6-8 Gm of acetylsalicylic acid daily, and urine collections were made at various intervals following the infusion. In some of these urine samples maximal liberation of radioactive glucuronoside was obtained only when using very high concentrations of a beef liver preparation of [beta]-glucuronidase (3000 units per milliliter of urine) and with prolonged periods (five days) of incubation at 38[degree]C. With a smaller amount of enzyme (400 units per milliliter of urine) and a shorter period of incubation (forty-eight hours at 38[degree]C), the degree of hydrolysis of corticosteroid glucuronosides was inversely related to the amount of salicylate glucuronoside present in the urine samples. The data suggest that salicylate glucuronoside acted as a competing substrate for the enzyme. When [beta]-glucuronidase from beef liver was incubated at 38[degree]C with urine from normal subjects, there was an average loss of 62 percent of the enzymatic activity after five days; approximately half of this loss occurred during the first twenty-four hours. The rate of enzyme inactivation did not appear to be altered by the presence of salicylate or its metabolites.