The influence of aging on the stereoselective pharmacokinetics of propranolol in the rat.

The influence of aging on the pharmacokinetics and the tissue distribution of (R)- and of (S)-propranolol was studied in 3-, 12-, and 24-month-old rats. After both iv and oral administration of rac-propranolol, the plasma concentrations were higher for the (R)- than for the (S)-enantiomer. For the tissue concentrations, the reverse was true. The free fraction of (S)-propranolol in plasma was about 4 times larger than that of (R)-propranolol, and this is the main factor responsible for the differences in kinetics between the two enantiomers. There was a suggestion for a difference in tissue binding between the two enantiomers. With aging, the plasma and tissue concentrations of both enantiomers increase, probably due to a decrease in blood clearance. Tissue binding did not change much with aging. Notwithstanding the marked differences between the kinetics of the propranolol enantiomers, the changes which occur with aging affect both enantiomers to the same degree.     

Stereoselective Accumulation of Propranolol Enantiomers in K562 and K562/ADR Cells

The stereoselective uptake of propranolol enantiomers was investigated by using the K562 and K562 adriamycin‐resistant cell line (K562/ADR) as a model. An enantioselective RP‐HPLC method was applied to determine the accumulation of propranolol (PPL) stereoisomers in K562 and K562/ADR cells. The concentration, time and temperature dependent studies showed that the accumulation of S‐(?)‐PPL was higher than R‐(+)‐PPL in K562 cells and uptake of R‐(+)‐PPL was significantly higher than that of S‐(?)‐PPL in K562/ADR cells. The results indicate the enantioselective accumulation of propranolol enantiomers in K562 and K562 / ADR cells. Chirality 25:361–364, 2013. © 2013 Wiley Periodicals, Inc.     

Enantioselective retardation of rac-propranolol from matrices containing cellulose derivatives

The dissolution characteristics of propranolol enantiomers from tablet formulations containing cellulose, or one of eight cellulose derivatives, were determined under a range of conditions. The derivatives examined were: cellulose tris(phenylcarbamate) (1), cellulose tris(2,3-dichlorophenylcarbamate) (2), cellulose tris 2,4-dichlorophenylcarbamate (3), cellulose tris(2,6-dichlorophenylcarbamate) (4), cellulose tris(2,3-dimethylphenylcarbamate) (5), cellulose tris(3,4-dichlorophenylcarbamate) (6), cellulose tris (3,5-dichlorophenylcarbamate) (7), cellulose tris(3,5-dimethylphenylcarbamate) (8). In water at 25°C, the release rates of (-)R-propranolol were generally greater than those of (-)-S-propranolol, although these differences were not always statistically significant; only compounds 5 and 8 demonstrated significant enantioselectivity. Using compound 8 in further experiments, statistically significant stereoselective dissolution of propranolol HCl was observed in buffer pH 7.4 at 25°C (intrinsic dissolution rates: 0.41 ± 0.01 mgcm²min⁻¹ for R-propranolol and 0.30 ± 0.02 mgcm²min⁻¹ for S-propranolol; P = 0.003). The cumulative amounts of enantiomers released at every time point were also found to be statistically significant (mean ratio R:S 1.25 ± 0.05). The observed low stereoselectivity of 8 with propranolol base was probably attributable to low solubility in pH 7.4 buffer, although stereoselective release did increase with time. This suggested that there is a relationship between stereoselectivity and contact time in an aqueous environment. Results also suggested that increased temperature may affect the release process as well as stereoselective interactions of 8 with individual enantiomers. To conclude, differential release of rac-propranolol from cellulose derivative matrices has been demonstrated, which supports the principle of stereoselective retardation as a potential means of stereoselective drug delivery for solid dosage forms. Chirality 9:307–312, 1997. © 1997 Wiley-Liss, Inc.     

Stereoselective pharmacokinetics of oxprenolol,propranolol, and verapamil: Species differences and influence of endotoxin

The influence of endotoxin-induced inflammation was studied on the pharmacokinetics of the enantiomers of the racemic drugs oxprenolol, propranolol, and verapamil in rabbits and dogs. Enantioselective pharmacokinetics were seen for oxprenolol and propranolol in the rabbit and for propranolol and verapamil in the dog. In the dog, the enantioselective differences in plasma concentrations are due to differences in both protein binding and metabolism, whereas in the rabbit the differences are due solely to differences in metabolism. In both species endotoxin treatment increases the plasma concentrations of the enantiomers of the three drugs; both protein binding and metabolism are influenced. In rabbits and in dogs, the influence of endotoxin on the disposition of the three drugs is less enantioselective than was previously observed in the rat. © 1995 Wiley-Liss, Inc.     

Toxicokinetics of a single intravenous dose of rac-propranolol versus optically pure propranolol in the rat

Conscious male Wistar SPF Riv:TOX rats were dosed intravenously with 2.5, 5, or 10 mg/kg rac-propranolol·HCl, or with 5 mg/kg of either (-)-(S)- or (+)-(R)-propranolol·HCl. Disposition of (-)-(S)- and (+)-(R)-propranolol after dosing of rac-propranolol was linear in the dose range examined. Total plasma clearance was not changed in animals dosed with the individual enantiomers compared to the animals that were dosed with rac-propranolol. However, for (-)-(S)-propranolol both volume of distribution and elimination half-life decreased, whereas for (+)-(R)-propranolol increases were observed for these characteristics, in animals dosed with the individual enantiomers. Our observations suggest that the (+)-(R)-enantiomer competes with (-)-(S)-propranolol for plasma protein binding sites, resulting in lower plasma protein binding of the (-)-(S)-enantiomer when the racemate is administered. From recent toxicological experiments, it was concluded that rac-propranolol is more toxic than the individual enantiomers in the rat, when dosed iv at the same total mass. It is concluded that the observed potentiation of toxic effects of propranolol enantiomers when administered as a racemate can at least partly be explained by a pharmacokinetic interaction. © 1995 Wiley-Liss, Inc.     

Toxic doses of rac-, (−)-(S)- and (+)-(R)-propranolol in rats and rabbits

The contribution of the individual enantiomers ([+]-[R]- and [−]-[S]-propranolol) to rac-propranolol intoxication was studied in anaesthetized, spontaneously breathing (SB) rats and artificially ventilated (AV) rats and rabbits. In the SB rat, propranolol (30 mg.kg⁻¹.h⁻¹ i.v.) decreased heart rate and mean arterial blood pressure and caused hypoventilation, serious hypoxaemia, respiratory acidosis, and death by respiratory arrest. Survival time (ST) in the (+)-(R)-propranolol group (ST 91 ± 5 min) was significantly longer than in the rac-propranolol group (ST 68 ± 6 min). In AV rats and rabbits toxic doses of rac-, (−)-(S)- and (+)-(R)-propranolol, 30 mg.kg⁻¹.h⁻¹ and 15 mg.kg⁻¹.h⁻¹ i.v., respectively, induced comparable effects on haemodynamic variables as in the SB rat. Artificial ventilation lengthened ST by a factor of three to four in rats. In the AV rat, ST's were not significantly different between the rac-, (−)-(S)- and (+)-(R)-propranolol groups. In the rabbit, as in the SB rat, ST in the (+)-(R)-propranolol group was significantly longer than ST's in the rac- and (−)-(S)-propranolol groups. The acute respiratory acidosis in SB rats and the prolonged ST in AV rats suggest that respiratory failure is the primary and cardiovascular failure the secondary cause of death in propranolol intoxication. The potentiation of the toxic effect of the enantiomers observed after dosing the racemate instead of the pure enantiomers could not be explained by a stereoselective difference in plasma propanolol concentration. © 1996 Wiley-Liss, Inc.     

Stereoselective disposition of S- and R-licarbazepine in mice

The stereoselective disposition of S-licarbazepine (S-Lic) and R-licarbazepine (R-Lic) was investigated in plasma, brain, liver, and kidney tissues after their individual administration (350 mg/kg) to mice by oral gavage. Plasma, brain, liver, and kidney concentrations of licarbazepine enantiomers and their metabolites were determined over the time by a validated chiral HPLC-UV method. The mean concentration data, attained at each time point, were analyzed using a non-compartmental model. S-Lic and R-Lic were rapidly absorbed from gastrointestinal tract of mouse and immediately distributed to tissues supplied with high blood flow rates. Both licarbazepine enantiomers were metabolized to a small extent, each parent compound being mainly responsible for the systemic and tissue drug exposure. The stereoselectivity in the metabolism and distribution of S- and R-Lic was easily identified. An additional metabolite was detected following R-Lic administration and S-Lic showed a particular predisposition for hepatic and renal accumulation. Stereoselective processes were also identified at the blood-brain barrier, with the brain exposure to S-Lic almost twice that of R-Lic. Another finding, reported here for the first time, was the ability of the mouse to perform the chiral inversion of S- and R-Lic, albeit to a small extent.     

Enantioselective inhibitory effect of nicardipine on the hepatic clearance of propranolol in man

The influence of a single oral dose of 30 mg nicardipine on the pharmacokinetics of (R)- and (S)-propranolol, given orally as rac-propranolol 80 mg, was studied in 12 healthy volunteers. The plasma concentrations were higher for the (S)-enantiomer than for the (R)-enantiomer. The Cl_o and the Cl′_intr of (S)-propranolol were significantly lower than the Cl_o and Cl′_intr of (R)-propranolol. The unbound fraction of (R)-propranolol was significantly higher than that of (S)-propranolol. Coadministration of nicardipine significantly increased the AUC and C_max and significantly decreased the Cl_o and Cl′_intr for unbound drug of (R)- and (S)-propranolol. These changes were more important for (R)- than for (S)-propranolol. The protein binding was not altered by nicardipine. The enantioselective effect of nicardipine on the metabolic clearance of propranolol appears to be due to an interaction at the level of the metabolizing enzymes. The effect on blood pressure of rac-propranolol was little affected when nicardipine was coadministered with rac-propranolol, and its bradycardic effect was reduced. © 1994 Wiley-Liss, Inc.     

Stereoselective disposition and tissue distribution of carvedilol enantiomers in rats.

After intravenous bolus injection of rac-carvedilol at 2 mg/kg to the rat, the (+)-(R)- and (-)-(S)-enantiomer levels in the blood and tissues (liver, kidney, heart, muscle, spleen, and aorta) were measured by stereospecific HPLC assay. As compared with the (+)-(R), the (-)-(S) had a larger Vdss (3.32 vs. 2.21 liter/kg), MRT (33.4 vs. 25.6 min), and CLtot (96.1 vs. 83.8 ml/min/kg). AUC comparison after iv and po administration showed systemic bioavailability of the (-)-(S) to be about half that of its antipode, explained by the fact that the free fraction of the (-)-(S) in blood was 1.65-fold greater than that of the (+)-(R). Tissue-to-blood partition coefficient values for the (-)-(S) were 1.6- to 2.1-fold greater than those for the (+)-(R) in all tissues, showing that the (-)-(S) accumulates more extensively in the tissues. These results were consistent with the greater Vdss for the (-)-(S) estimated from systemic blood data. The stereoselective tissue distribution of carvedilol enantiomers results from an enantiomeric difference in plasma protein binding rather than in tissue binding.     

Myocardial uptake of thiopental enantiomers by the isolated perfused rat heart

Myocardial uptake of thiopental enantiomers by an isolated perfused rat heart preparation was examined after perfusion with protein-free perfusate. Outflow perfusate samples were collected at frequent intervals for 20 min during single-pass perfusion with 10 μg/ml racemic thiopental (washin phase) and for another 45 min during perfusion with drug-free perfusate (washout phase). (+)- and (−)-thiopental concentrations were assayed by chiral high-performance liquid chromatography. Heart rate, perfusion pressure, and electrocardiogram were also monitored. During the washin phase, there was no significant difference between the mean values of the equilibration rate constants of (+)- and (−)-thiopental enantiomers (0.44 ± 0.07 min⁻¹ and 0.43 ± 0.09 min⁻¹, respectively, P > 0.05). Mean volumes of distribution of (+)- and (−)-thiopental enantiomers were similar (6.34 ± 1.20 and 6.45 ± 1.29 ml/g for the washin phase and 7.22 ± 0.71 and 7.47 ± 0.81 ml/g for the washout phase, respectively, P > 0.05). This indicates that tissue accumulation of thiopental enantiomers in the isolated perfused rat heart was not stereoselective. Uptake of thiopental by the heart was perfusion flow rate-limited and independent of capillary permeability. These findings suggest that myocardial tissue concentration of racemic thiopental should be an accurate predictor of myocardial drug effect. © 1996 Wiley-Liss, Inc.     

Chiral Assay of Atenolol Present in Microdialysis and Plasma Samples of Rats Using Chiral CBH as Stationary Phase

Two different enantioselective chiral chromatographic methods were developed and validated to investigate the disposition of the β₁-receptor antagonist atenolol in blood and in brain extracellular fluid of rats (tissue dialysates). System A for the plasma samples was a one-column chromatographic system with a Chiral CBH column with an aqueous buffer as mobile phase into which cellobiose was added for selective regulation of the retention of the internal standard, (S)-metoprolol. The plasma samples were analysed after a simple extraction procedure. The limit of quantitation was 0.2 μg/ml for the atenolol enantiomers. The repeatability of the medium concentration quality control plasma sample (6.0 μg rac-atenolol/ml) was 11–18% for the enantiomers. The dynamic linear range of the plasma samples was 0.5–20 μg/ml. For system B, since atenolol is an extremely hydrophilic drug, the tissue dialysate sample required a much more sensitive system as compared to the plasma samples. A coupled column system was used for peak compression of the enantiomers in the eluate after the separation on the Chiral CBH column, hence increasing the detection sensitivity. The limit of quantification was 0.045 μg/ml for the atenolol enantiomers in artificial CSF. The repeatability of the medium concentration quality control samples (0.1 and 4.0 μg rac-atenolol/ml in artificial CSF and Hepes Ringer, respectively) was 2.8–9.3% for the two enantiomers. The dynamic linear range of the brain samples was 0.05–1.0 and 0.5–20 μg/ml in artificial CSF and Hepes Ringer, respectively. Chirality 9:329–334, 1997. © 1997 Wiley-Liss, Inc.     

High-performance liquid chromatographic determination of (S)- and (R)-propanolol in human plasma and urine with a chiral β-cyclodextrin bonded phase

The determination of propanolol enantiomers in microsamples of human plasma and urine by HPLC using a chiral stationary phase is described. After extraction from 200 μl of plasma or urine with racemic alprenolol as internal standard (I.S.), the enantiomers are separated on a β-cyclodextrin column with a polar organic mobile phase and determined by fluorescence detection. The retention times of I.S. and propranolol enantiomers are about 12–13 min and 16–18 min, respectively. Peak resolutions are 1.4 for I.S. and 2.2 for propranol. The use of alprenolol as I.S. improves significantly the coefficients of variation (C.V.: 0.6–4.2%). Sensitivity is approximately 1.5 ng/ml per propranolol enantiomer. The assay is applied to pharmacokinetic studies of racemic propranolol in human biological fluids. The (S)-propranolol levels are always higher than the (R)-antipode concentrations in plasma and urine.     

Stereoselective plasma protein binding and target tissue distribution of clausenamide enantiomers in rats

Stereoselective differences in pharmacokinetics between clausenamide (CLA) enantiomers have been found after intravenous and oral administration of each enantiomer to rats. The differences could be associated with protein binding of CLA enantiomers. By equilibrium dialysis methods, the binding of CLA enantiomers to rat plasma protein was investigated. The results showed that mean percentages of (-) and (+)CLA in the bound form were 28.5% and 38.0%, respectively, indicating that the unbound fraction of (-)CLA was higher than that of (+)CLA, which provided an explanation for stereoselective pharmacokinetics of CLA enantiomers in rats. The results also showed that there were species differences in plasma protein binding of (-)-isomer between rats (28.5%) and rabbits (47.2%). Furthermore, effects of plasma protein binding on the distribution of CLA enantiomers to their possible target tissues were observed. The amount of (-)CLA in brain was greater than that of (+)CLA 15 min after administration of each enantiomer to rats. But the results were reverse at 4 h postdose. Further studies in distributional kinetics showed that (-)CLA had a more rapid absorption and distribution to hippocampus, cortex, and cerebellum than (+) CLA. (+)CLA had greater values for T(max), t(1/2) (beta), and AUC(0) (-->infinity), and smaller ones for CL/F and V(d)/F than its antipode. The data indicated that the distribution of (-) and (+)CLA in their target tissues was stereoselective. The stereoselective distribution might be involved in the metabolism and transport of two enantiomers in the central nerve system.     

Influence of endotoxin on the stereoselective pharmacokinetics of oxprenolol,propranolol, and verapamil in the rat

The influence of endotoxin-induced inflammation on the enantioselective pharmacokinetics of propranolol, oxprenolol, and verapamil, which bind to α₁-acid glycoprotein, was studied in the rat. The racemic mixtures were given orally. In the control animals, for propranolol and oxprenolol, the plasma concentrations of the (R)-enantiomer were higher than those of the (S)-enantiomer, while for verapamil the reverse was true. Protein binding and intrinsic clearance are the main factors responsible for this enantioselectivity. After endotoxin treatment, for the three drugs tested the plasma concentrations and the plasma binding of both enantiomers were significantly increased. This effect was more pronounced for (R)-propranolol, (R)-oxprenolol, and (S)-verapamil than for their respective antipodes. The enantioselective effect of endotoxin on the plasma concentrations of the drugs studied seems mainly due to the enantioselective increase in binding to α₁-acid glycoprotein. © 1994 Wiley-Liss, Inc.     

Solubility, metastable zone width, and racemic characterization of propranolol hydrochloride

Characterization of the racemic species, which can be a racemic compound, a racemic conglomerate, or a pseudoracemate (solid solution), is a prerequisite for the design of crystallization resolution processes. It is useful to determine the solid/liquid equilibrium solubility of the enantiomer mixtures for crystallization operation. For the beta-blocker drug propranolol hydrochloride, Gibbs free energy of formation of racemic compound and entropy of mixing of the (R)- and (S)- enantiomers in the liquid state for racemic conglomerate were calculated. The structural differences between (R, S)-propranolol hydrochloride and its (S)-enantiomer were further investigated by powder X-ray diffraction patterns, infrared spectra, and solid-state NMR spectra. The solubility and metastable zone width of (R, S)- propranolol hydrochloride in a mixed solvent of methanol and acetone were determined by cooling crystallization over the temperature range 3.5-42.5 degrees C. The ternary solubility diagram of (R)-, (S)-propranolol hydrochloride was constructed using the same mixed solvent. The diagram will be useful as a guide for choosing crystallization operation conditions to produce pure enantiomers.     

THE FLUOROMETRIC DETERMINATION OF 5-METHOXYTRYPTAMINE IN MAMMALIAN TISSUES AND FLUIDS

Abstract— Using a sensitive and specific fluorometric procedure involving selective extraction, reaction of the extracts with o -phthalaldehyde (OP), separation of the OP derivatives by TLC, and determination of fluorescent characteristics and intensities, we have detected and measured 5-methoxytryptamine, (5-MT) in various central and peripheral tissues and fluids of the rat, dog, baboon, and man.
Distribution of 5-MT in peripheral tissues of the rat seemed to parallel that of 5-HT, with highest levels being found in the gastrointestinal (GI) tract and Harderian gland, regions that are rich in 5-HT and have been reported to contain systems capable of methylating 5-HT. 5-MT was detected in the lung, plasma, kidney, spleen, and heart of the rat. 5-MT was present in the CNS of all species examined. No marked interspecies differences were observed. In the rat CNS, the regional distribution of 5-MT did not parallel that of 5-HT indicating that the systems for the synthesis, uptake, or transport of 5-MT might be different than for 5-HT. Pretreatment of rats with iproniazid resulted in a 50% increase in whole brain 5-MT. Reserpine pretreatment had no effect, indicating that the storage or release mechanisms for 5-MT are different than for the conventional amine transmitters. 5-MT was detected in human CSF and urine but not in plasma. These data indicate that 5-MT, a compound with potent pharmacological properties, is more widely distributed in the mammalian body than had previously been supposed.     

Steady‐State Plasma Concentration of Donepezil Enantiomers and Its Stereoselective Metabolism and Transport In Vitro

The aim of the present study was to elucidate the differences in the plasma concentration of two enantiomers of donepezil in Chinese patients with Alzheimer's disease (AD) and investigate in vitro stereoselective metabolism and transport. Donepezil enantiomers were separated and determined by LC‐MS/MS using D5‐donepezil as an internal standard on a Sepax Chiralomix SB‐5 column. In vitro stereoselective metabolism and transport of donepezil were investigated in human liver microsomes and MDCKII‐MDR1 cell monolayer. Pre‐dose (Css‐min) plasma concentrations were determined in 52 patients. The mean plasma level of (R)‐donepezil was 14.94 ng/ml and that of (S)‐donepezil was 23.37 ng/ml. One patient's plasma concentration of (R)‐donepezil was higher than (S)‐donepezil and the ratio is 1.51. The mean plasma levels of (S)‐donepezil were found to be higher than those of (R)‐donepezil in 51 patients and the ratio of plasma (R)‐ to (S)‐donepezil varies from 0.34 to 0.85. In the in vitro microsomal system, (R)‐donepezil degraded faster than (S)‐donepezil. V_max of (R)‐donepezil was significantly higher than (S)‐donepezil. The P‐gp inhibition experiment shown that the P_app of the two enantiomers was higher than 200 and the efflux ratios were 1.11 and 0.99. The results of the P‐gp inhibition identification experiment showed IC50 values of 35.5 and 20.4 μM, respectively, for the two enantiomers. The results indicate that donepezil exhibits stereoselective hepatic metabolism that may explain the differences in the steady‐state plasma concentrations observed. Neither (R)‐ nor (S)‐donepezil was a P‐gp substance and the two enantiomers are highly permeable through the blood–brain barrier. Chirality 25:498–505, 2013. © 2013 Wiley Periodicals, Inc.     

Fiber-based liquid-phase micro-extraction of mebeverine enantiomers followed by chiral high-performance liquid chromatography analysis and its application to pharmacokinetics study in rat plasma

The applicability of two-phase liquid-phase micro-extraction (LPME) in porous hollow polypropylene fiber for the sample preparation and the stereoselective pharmacokinetics of mebeverine (MEB) enantiomers (an antispasmodic drug) in rat after intramuscular administration were studied. Plasma was assayed for MEB enantiomer concentrations using stereospecific high-performance liquid chromatography with ultraviolet detection after a simple, inexpensive, and efficient preconcentration and clean-up hollow fiber-based LPME. Under optimized micro-extraction conditions, MEB enantiomers were extracted with 25 μl of 1-octanol within a lumen of a hollow fiber from 0.5 ml of plasma previously diluted with 4.5 ml alkalized water (pH 10). The chromatographic analysis was carried out through chiral liquid chromatography using a DELTA S column and hexane-isopropyl alcohol (85:15 v/v) containing 0.2% triethylamine as mobile phase. The mean recoveries of (+)-MEB and (-)-MEB were 75.5% and 71.0%, respectively. The limit of detection (LOD) was 3.0 ng/ml with linear response over the concentration range of 10-2500 ng/ml with correlation coefficient higher than 0.993 for both enantiomers. The pharmacokinetic studies showed that the mean plasma levels of (+)-MEB were higher than those of (-)-MEB at almost all time points. Also, (+)-MEB exhibited greater t(max) (peak time in concentration-time profile), C(max) (peak concentration in concentration-time profile), t(1/2) (elimination half-life), and AUC(0-240 min) (area under the curve for concentration versus time) and smaller CL (clearance) and V(d) (apparent distribution volume) than its antipode. The obtained results implied that the absorption, distribution, and elimination of (-)-MEB were more rapid than those of (+)-MEB and there were stereoselective differences in pharmacokinetics.     

Stereoselective distribution of hydroxychloroquine in the rabbit following single and multiple oral doses of the racemate and the separate enantiomers

Hydroxychloroquine (HCQ) stereoselective distribution was investigated in rabbits after 20 mg/kg po of racemic-HCQ (rac-HCQ) and 20 mg/kg po of each enantiomer, 97% pure (?)-(R)-HCQ and 99% pure (+)-(S)-HCQ. Concentrations were 4 to 6 times higher in whole blood than in plasma. Melanin did not affect plasma and whole blood levels since concentrations did not differ between pigmented and nonpigmented animals. After single and multiple doses of the separate enantiomers, only 5–10% of the antipode could be measured, in blood or plasma. Therefore, there was no significant interconversion from one enantiomer into the other. Following rac-HCQ, plasma (+)-(S)-levels always surpassed (?)-(R)-ones while in whole blood, (?)-(R)-HCQ concentrations were always the highest. When the enantiomers were administered separately, blood concentrations achieved after (?)-(R)-HCQ were higher, especially after multiple doses. These observations suggest that (?)-(R)-HCQ is preferentially concentrated by cellular components of blood. This enantioselective distribution of HCQ could be secondary to a stereoselective protein binding to plasma proteins, although a more specific binding of (?)-(R)-HCQ to blood cells cannot be ruled out. Since in whole blood (?)-(R)-HCQ is retained in cellular components, metabolism would favour the more available (+)-(S)-enantiomer. © 1994 Wiley-Liss, Inc.     

Determination of protein binding for novel 2-(2-hydroxypropanamido)-5-trifluoromethyl benzoic acid enantiomers to rats,dogs, and humans plasma by UPLC-MS/MS

R-/S-2-(2-hydroxypropanamido)-5-trifluoromethyl benzoic acid (R-/S-HFBA) is a novel COX inhibitor with remarkable anti-inflammatory and antiplatelet aggregation activities, but no gastrointestinal toxicity. In our previous study, the different pharmacokinetic profiles of the two enantiomers in rats were observed after administration of R-HFBA and S-HFBA. Stereoselective protein binding of the two enantiomers may be a reason for the different pharmacokinetic behaviors. In this study, we developed and validated an UPLC-MS/MS method for determining stereoselective binding of HFBA enantiomers to rat, dog, and human plasma in vitro. Chromatographic separation was achieved by gradient elution with a flow rate of 0.4 mL/min. MS/MS detection was operated in positive electrospray using multiple reaction monitoring (MRM) mode. The method was proved to be linear over the concentration range of 0.005 to 10 μg/mL with a lower limit of quantification of 0.005 μg/mL. The developed method was successfully employed to the plasma protein binding study of HFBA enantiomers. Equilibrium dialysis method was applied to assess drug-plasma protein interactions. The results showed that the enantiomers were both extensively bound to three species plasma and protein binding of R-/S-HFBA was concentration dependent. R-HFBA and S-HFBA showed significant species difference among rat, dog, and human plasma and stereoselective plasma protein binding.