Acyl glucuronides formed from carboxylic acids can undergo hydrolysis, acyl migration, and covalent binding to proteins. In buffers at physiological pH, the degradation of acylglucuronide of a chiral NSAID, carprofen, consisted mainly of acyl migration. Acidic pH reduced hydrolysis and acyl migration, thus stabilizing the carprofen acyl glucuronides. Addition of human serum albumin (HSA) led to an increased hydrolysis of the conjugates of both enantiomers. This protein protected R-carprofen glucuronide from migration and therefore improved its overall stability. Hydrolysis was stereoselective in favor of the S conjugate. The protein domains and the amino acid residues likely to be responsible for the hydrolytic activity of HSA were deduced from the results of various investigations: competition with probes specific of binding sites, effects of pH and of chemical modifications of albumin. Dansylsarcosine (DS), a specific ligand of site II of HSA, impaired the hydrolysis, whereas dansylamide (DNSA) and digoxin, which are specific ligands of sites I and III, respectively, had no effect. The extent of hydrolysis by HSA strongly increased with pH, indicating the participation of basic amino acids in this process. The results obtained with chemically modified HSA suggest the major involvement of Tyr and Lys residues in the hydrolysis of glucuronide of S-carprofen, and of other Lys residues for that of its diastereoisomer.
Copyright 2000 Wiley-Liss, Inc.