In vitro stereoselective covalent binding of carprofen glucuronides to human serum albumin: characterization of the mechanism.
The reactivity, in terms of covalent binding, of R- and S-carprofen acylglucuronides with human serum albumin (HSA) has been investigated in vitro. The irreversible binding of these metabolites to the HSA 580 mM occurred at pH 7.4 and 37 degrees C instantaneously and stereoselectively in favour of the R-enentiomer glucuronide. The amount of carprofen adducts remained stable with time up to 48 hr, and increased with the glucuronide concentration. It was not modified by addiction of imine-trapping reagents, suggesting that the reaction is not mediated by a Schiff base mechanism. Moreover the extreme rapidity of the covalent binding supports a mechanism of nucleophilic attack. Competition studies with ligands known to bind to different sites of HSA, indicated that carprofen glucuronides interacted mainly with site II. The extent of the binding of R-carprofen glucuronide increased with pH, thus suggesting the participation of an alkaline group in the process. The modification of HSA by amino-acid directed chemicals led to the conclusion that Tyr, Lys or Arg residues in site II were mainly involved.
Greige-Georges
Hélène
H
Université libanaise, Faculté de Sciences, Section II, Fanar, Lebanon.
Buronfosse
Thierry
T
Netter
Patrcik
P
Magdalou
Jacques
J
Lapicque
Françoise
F
D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000378:metabolism*; D001665:Binding Sites; D002227:Carbazoles / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D020719:Glucuronides / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007700:Kinetics; D008024:Ligands; D011485:Protein Binding; D012709:Serum Albumin / Q000737:chemistry / Q000378:metabolism*; D013237:Stereoisomerism; D013696:Temperature
D000596:Amino Acids; D000894:Anti-Inflammatory Agents, Non-Steroidal; D002227:Carbazoles; D020719:Glucuronides; D008024:Ligands; D012709:Serum Albumin; C007005:carprofen