Species-dependent enantioselective glucuronidation of carprofen.

1. The stereoselective glucuronidation of carprofen, a non-steroidal anti-inflammatory drug, was investigated in vitro using microsomes prepared from liver of different species (rat, dog, horse, sheep and man) or UGT2B1 expressed in fibroblasts. 2. The Km towards the drug was very similar among these species and for the two enantiomers, whereas the Vmax varied substantially according to the animal used. The rat exhibited a high stereoselective glucuronidation whereas other species, including man, presented a low stereoselectivity. The R-enantiomer was glucuronidated at a more efficient rate than its enantiomorph, and was a better substrate (in terms of Vmax/Km). 3. To explain the enantioselective disposition of carprofen in man and in the different species, the ratio of the enzymatic efficacies (Vmax/Km) were compared with the ratio of the pharmacokinetic parameters AUCs. The basic hypothesis that the intrinsic clearance reflect the enantioselective behaviour of carprofen seemed substantiated when we focused on man and rat glucuronidation, but the in vivo-in-vitro correlation was not possible in other species. 4. In conclusion, the chiral pharmacokinetics of carprofen is less dependent on the stereoselective glucuronidation than other stereoselective processes such as protein binding of carprofen, enzymatic hydrolysis, or renal elimination of glucuronides.

Enantioselectivity of the enterohepatic recycling of carprofen in the dog.

The disposition of the two enantiomers of carprofen (CPF), the (R)-CPF and the (S)-CPF, was investigated after iv administration of the racemate (4 mg/kg) in dogs equipped with a chronic bile duct catheter. Studies in dogs with diverted bile flow showed that both enantiomers were extensively excreted in bile with 74% of the (R)-enantiomer and 92% of the (S)-enantiomer from the iv administered dose being recovered in the bile as the respective glucuronide conjugates. The direct administration of acidic bile containing acyl-glucuronides of CPF in the duodenum showed that both conjugated enantiomers led to high CPF enantiomer systemic availability. However, comparison of CPF pharmacokinetics between dogs with nondiverted bile flow and dogs with diverted bile flow suggested that CPF was subjected to enantioselective enterohepatic recycling (EHC) and that only the (S)-CPF was recycled. The absence of EHC for the (R)-CPF is hypothesized to be the result of formation of glucuronidase-resistant isoglucuronides (epimers) to a greater extent for the (R)-CPF than for the (S)-CPF.

Enantioselective behaviour of drugs used in domestic animals: a review.

The chirality of drugs, with particular reference to agents used in veterinary medicine, is reviewed. Basic concepts of chirality and aspects of the methodology for the separation of enantiomers are considered. Chiral compounds are in common use in animals and their pharmacological actions and side-effects (pharmacodynamics) and absorption into and fate within the body (pharmacokinetics) are of fundamental importance; pharmacodynamic and pharmacokinetic properties of enantiomeric pairs commonly differ and this has major implications for their effective and safe therapeutic use. As examples of the particular significance of chirality in veterinary medicine, the following drug classes are reviewed; benzimidazole anthelmintics, cloprostenol, verapamil, ketamine, halogenated hydrocarbon anaesthetics and 2-arylpropionic acid anti-inflammatory drugs. The implications of chirality for drug product development and approval by registration authorities are discussed.

[Enantioselectivity in the excretion of glucuronides of carprofen in man, dogs and horses]. / Enantiosélectivité de l'excrétion des glucuronides du carprofène chez l'homme, le chien et le cheval.

After administration of the racemic drug, the stereoselective quantification of the enantiomers of free and conjugated carprofen was performed in human plasma and in plasma, urine and bile of dogs and horses. In humans, the plasma profile of free carprofen and its glucuronides is not stereoselective and the glucuronides excreted in urine are close to a racemate. In dogs and horses on the contrary, the R(-) enantiomer of the free drug is predominant in plasma, while urine and/or bile concentrations of the glucuronides are high in comparison to plasma with a strong selectivity for the S(+) enantiomer. Because glucuronidation of carprofen, as a carboxylic compound, is known to be the major metabolic pathway in most species, interspecies discrepancies in the stereoselective disposition of carprofen seem to be mainly related to the stereoselectivity in the excretion of the glucuronides. Finally, the high plasma concentrations of carprofen glucuronides in human in comparison to other animal species suggest that the former could be specifically subjected to immunological side effects in the time course of treatments by this type of compounds.

Pharmacodynamics and enantioselective pharmacokinetics of carprofen in the cat.

The pharmacodynamics and enantioselective pharmacokinetics of the arylpropionic acid non-steroidal anti-inflammatory drug, carprofen, were investigated in cats after administration of the racemic mixture (rac-carprofen) at dose rates ranging from 0.7 to 4.0 mg kg-1 intravenously and subcutaneously. A low dose of rac-carprofen (0.7 mg kg-1) partially inhibited the rise in skin temperature at a site of acute inflammation but had no effect on the ex vivo synthesis of serum thromboxane (Tx) B2. A higher dose (4.0 mg kg-1) inhibited oedematous swelling, although the response was statistically significant at only one time, and also reduced the ex vivo synthesis of serum TxB2 for 12 hours after intravenous injection or 24 hours after subcutaneous injection. The main features of carprofen pharmacokinetics were a low distribution volume, a relatively long elimination half-life, the predominance of the R(-) enantiomer and a bioavailability (after subcutaneous dosing) of 100 per cent and 92 per cent, respectively, after doses of 0.7 and 4.0 mg kg-1. On the basis of these data, it is suggested that a dose of 4.0 mg kg-1 by both intravenous and subcutaneous routes should be evaluated in clinical subjects.

Pharmacodynamics and chiral pharmacokinetics of carprofen in calves.

The non-steroidal anti-inflammatory drug, carprofen, was administered intravenously as the racemate at a dose rate of 0.7 mg kg-1 to six Friesian bull calves aged 8-10 weeks. Anti-inflammatory properties were indicated by attenuation of temperature rise at sites of intradermal injection of the irritants, carrageenin and dextran, but responses were not statistically significant at most recording times. Carrageenin- and dextran-induced swelling were not significantly reduced by carprofen. Carprofen reduced ex vivo serum thromboxane B2 synthesis but this effect was also not significant at most sampling times. Enantioselective pharmacokinetics of carprofen was demonstrated, plasma concentrations of the R(-) enantiomer predominating at all sampling times. It is concluded that inhibition of cyclo-oxygenase is unlikely to be the sole mechanism of action of carprofen in calves.

Evaluation of carprofen in calves using a tissue cage model of inflammation.

The arylpropionate anti-inflammatory drug, carprofen, was administered intravenously as the racemate at a dose rate of 0.7 mg kg-1 body weight to six Friesian bull calves aged 16-17 weeks. Anti-inflammatory and pharmacokinetic properties were investigated using a tissue cage model of inflammation based on intracaveal injection of the mild irritant, carrageenin. Carprofen displayed enantioselective pharmacokinetics, with the R(-) enantiomer predominating in plasma at all measuring times. Elimination half-life and mean residence time were shorter and volume of distribution and clearance were greater for the S(+) than for the R(-) enantiomer. Penetration of both enantiomers into transudate (non-stimulated tissue cage) was poor but penetration into exudate (carrageenin-stimulated tissue cage) was good. Carprofen failed to reduce exudate concentration of prostaglandin E2 and the reductions in 12-hydroxyeicosatetraenoic acid were non-significant at most sampling times. The long elimination half-life of both R(-) and S(+) carprofen enantiomers and their ready penetration into and slow clearance from inflammatory exudate indicate that the drug is likely to have a long duration of action in calves. The mechanism of action is unknown but it is unlikely to involve inhibition of either cyclooxygenase or 12-lipoxygenase.

Chiral inversion of fenoprofen in horses and dogs: an in vivo-in vitro study.

Fenoprofen (FPF) is a chiral non-steroid antiinflammatory drug, marketed as a racemic mixture of its R(-) and S(+) enantiomers. Its stereoselective disposition in humans and animals is due to a chiral inversion converting R(-)FPF into S(+)FPF. The first step of this reaction, which produces an acyl-CoA thioester, is catalysed by an acyl-CoA ligase. A stereospecific high performance liquid chromatography assay was used to study the disposition of FPF enantiomers in four geldings and three male beagle dogs, following intravenous doses of racemic FPF (1 mg/kg in horses), R(-)FPF (0.5 mg/kg in horses, 1 mg/kg in dogs), and S(+)FPF (0.5 mg/kg in horses, 1 mg/kg in dogs). A unidirectional stereoinversion of the R(-) enantiomer into its optical antipode (38% in horses, 90% in dogs) was demonstrated. This explained the clear enantioselective behaviour of FPF in both species. Acyl-CoA ligase activity (Km = 473.2 +/- 92.5 microM; Vmax = 23 +/- 3.3 nmol/min/mg) has also been quantified in vitro on equine hepatic microsomes, using a high performance liquid chromatography method to measure thioester formation. The present study showed that, in horses and dogs, as previously demonstrated in rats and sheep, the R(-)FPF clearance was better correlated with ligase activity than with inversion rate. A highly significant linear relationship was demonstrated between these variables.