Racemic Drug Research Articles

Chiral Drugs. An Overview

About more than half of the drugs currently in use are chiral compounds and near 90% of the last ones are marketed as racemates consisting of an equimolar mixture of two enantiomers. Although they have the same chemical structure, most isomers of chiral drugs exhibit marked differences in biological activities such as pharmacology, toxicology, pharmacokinetics, metabolism etc. Some mechanisms of these properties are also explained. Therefore, it is important to promote the chiral separation and analysis of racemic drugs in pharmaceutical industry as well as in clinic in order to eliminate the unwanted isomer from the preparation and to find an optimal treatment and a right therapeutic control for the patient. In this article, we review the nomenclature, pharmacology, toxicology, pharmacokinetics, metabolism etc of some usual chiral drugs as well as their mechanisms. Different techniques used for the chiral separation in pharmaceutical industry as well as in clinical analyses are also examined.

Optimisation of the derivatization in cellulose‐type chiral selectors for enantioseparation by centrifugal partition chromatography

Cellulose was chemically modified with hydrophobic dodecanoyl groups followed by 3,5-dimethylphenylcarbamoyl substituents forming mixed ester/carbamate derivatives in order to improve the solubility in lipophilic solvents compared to the corresponding homosubstituted cellulose tris(3,5-dimethylphenylcarbamate). Two mixed derivatives of different degree of substitution were prepared, tested as chiral selectors (CSs) in counter-current chromatography (CCC) and compared to the homo-substituted derivative. Alternatively, cellulose tris(3,5-dichlorophenylcarbamate), was synthesised and tested with the same purpose. The racemic drugs pindolol and warfarin were used as test compounds. Biphasic organic/aqueous solvent systems, composed of methyl isobutyl ketone, t-butyl methyl ether or ethyl acetate and aqueous ammonium acetate or sodium phosphate buffer, were used. Centrifugal partition chromatography, a variety of CCC, in the classical elution mode and the pH-zone-refining displacement mode was applied. The enantioseparation of pindolol and warfarin was achieved in the latter conditions. The presence of dodecanoyl chains on the CS increased solubility in the organic solvents used. The selectivity of the mixed dodecanoyl/3,5-dimethylphenylcarbamoyl cellulose derivative with low degree of dodecanoyl groups (degree of substitution 0.3/degree of substitution 2.6, respectively) was similar to that of the homo-substituted derivative. Furthermore, the loading capacity for pindolol was increased by a factor of three compared to cellulose tris(3,5-dimethylphenylcarbamate). Nevertheless, the increasing degree of substitution with dodecanoyl groups on the CS, although improved solubility in the stationary phase, contributed negatively to the enantioselectivity, where warfarin was more affected than pindolol.

Highlighting the role of the chlorine substituents in the glycopeptide antibiotic balhimycin for chiral recognition in capillary electrophoresis

The vancomycin-type glycopeptide antibiotic balhimycin (I) and its dehaloanalogue dechlorobalhimycin (III), which is characterized by the total substitution of the two chlorine atoms of I by hydrogen, were employed as chiral selectors for the enantioresolution of 11 racemic dansyl amino acids and six 2-arylpropionic acid nonsteroidal anti-inflammatory racemic drugs by CE. The observed enantioresolution capability of I for all test analytes is clearly higher than that observed for III. This result suggests that chlorine substituents of I played a major role in the enantioresolution of these test analytes. A dimerization-based mechanism is proposed in order to explain this phenomenon. The two chlorine substituents of each monomer, which mutually penetrate into the cavity of the adjacent molecule of the dimer, are assumed to promote dimerization and as a consequence also enantioresolution.

Vancomycin as chiral selector for enantioselective separation of selected profen nonsteroidal anti‐inflammatory drugs incapillary liquid chromatography

The chiral selector vancomycin was used either as mobile phase additive or bound as a chiral stationary phase (CSP) for the stereoselective separation of seven racemic nonsteroidal anti-inflammatory drugs (NSAIDs), fenoprofen, carprofen, flurbiprofen, indoprofen, flobufen, ketoprofen, and suprofen, by capillary liquid chromatography. The effect of the type of stationary phase, the chiral column Chirobiotic V or the achiral stationary phases Nucleosil 100 C8 HD and Nucleosil 100 C18 HD, and the concentration of vancomycin in the mobile phase on separation of the drug enantiomers were evaluated. All the drugs, except flobufen, were successfully enantioseparated on Nucleosil 100 C8 HD with 4 mM vancomycin present in the mobile phase (composed of methanol and buffer) in the reversed phase mode. On the vancomycin-bonded chiral stationary phase, it was difficult to get enantioseparations of the profen NSAIDs. However, flobufen gave better enantioseparation on the vancomycin CSP. The better enantioresolution of the majority of profen derivatives on the achiral columns with vancomycin added to the mobile phase can be attributed in particular to the higher separation efficiency of this capillary chromatographic system. In addition, vancomycin dimers, formed in the mobile phase, seem to offer a better steric arrangement for stereoselective interaction to these analytes than the vancomycin bonded on the CSP. These substantial differences in the CS structure significantly influence the chiral discrimination mechanism.

Drug disposition in three dimensions: an update on stereoselectivity in pharmacokinetics

Many marketed drugs are chiral and are administered as the racemate, a 50:50 combination of two enantiomers. Pharmacodynamic and pharmacokinetic differences between enantiomers are well documented. Because of enantioselectivity in pharmacokinetics, results of in vitro pharmacodynamic studies involving enantiomers may differ from those in vivo where pharmacokinetic processes will proceed. With respect to pharmacokinetics, disparate plasma concentration vs time curves of enantiomers may result from the pharmacokinetic processes proceeding at different rates for the two enantiomers. At their foundation, pharmacokinetic processes may be enantioselective at the levels of drug absorption, distribution, metabolism and excretion. In some circumstances, one enantiomer can be chemically or biochemically inverted to its antipode in a unidirectional or bidirectional manner. Genetic consideration such as polymorphic drug metabolism and gender, and patient factors such as age, disease state and concomitant drug intake can all play a role in determining the relative plasma concentrations of the enantiomers of a racemic drug. The use of a nonstereoselective assay method for a racemic compound can lead to difficulties in interpretation of data from, for example, bioequivalence or dose/concentration vs effect assessments. In this review data from a number of representative studies involving pharmacokinetics of chiral drugs are presented and discussed.

PET neuroimaging of [ 11C]mirtazapine enantiomers in pigs

Previously, we used positron emission tomography (PET) for studying the pharmacokinetics of rac-[ 11C]mirtazapine in living brain. Our findings showed that rac-[ 11C]mirtazapine has suitable properties for PET neuroimaging. However, separate studies of enantiomers are typically required for characterizing the pharmacokinetics of a racemic drug. Therefore, we have determined the whole-body distribution and brain pharmacokinetics of S- and R-[ 11C]mirtazapine in pigs. The enantiomers of [ 11C]mirtazapine produced similar effective doses of radioactivity in most body organs, except for the brain, in which the dose was ∼ 40% higher after injection of S-[ 11C]mirtazapine than the antipode. Kinetic analyses of dynamic brain PET recordings showed that values for regional accumulation of compound ( k 3) were significantly higher for S-[ 11C]mirtazapine than for the antipode, while the values for clearance of compounds from tissue to circulation ( k 2) were consistently lower for S-[ 11C]mirtazapine than for the R-form. No reliable difference occurred in the rate of metabolism of S- and R-[ 11C]mirtazapine in the bloodstream of the pigs. The present findings indicate that enantioselective processes affect the cerebral pharmacokinetics of rac-mirtazapine.

Synthesis and application of mono-2 A-azido-2 A-deoxyperphenylcarbamoylated β-cyclodextrin and mono-2 A-azido-2 A-deoxyperacetylated β-cyclodextrin as chiral stationary phases for high-performance liquid chromatography

Two novel chiral stationary phases (CSPs) were prepared based upon the regioselective immobilizations of β-cyclodextrin (β-CD) at its C2 position to the silica support. The mono-2 A -azido-2 A -deoxyperphenylcarbamoylated β-cyclodextrin and mono-2 A -azido-2 A -deoxyperacetylated β-cyclodextrin were synthesized by selective tosylation and azidolysis followed by perfunctionalisation. The derivatised cyclodextrins were then immobilized onto the aminised silica gel via the Staudinger reaction to provide new chiral stationary phases. Their application to high-performance liquid chromatography (HPLC) enantioseparation of racemic compounds was demonstrated using β-adrenergic blockers, flavonone compounds, benzodiazepinones, antihistamines and weakly protolytic compounds, of which good separations were achieved for some racemic compounds, for instance, bendroflumethiazide ( R s 6.26), oxazepam ( R s 5.99), temazepam ( R s 2.85) and althiazide ( R s 1.13) when compared with the corresponding CSPs where the β-CD molecule was regioselectively immobilized at the C6 position. The enantiodiscriminatory properties of these CSPs were found to be affected by the orientation of the CD cavity under reversed-phase conditions, and also by the derivitising groups of the CD. The HPLC results inferred that the mono-6 A -azido-6 A -deoxyperphenylcarbamoylated CD CSP (CD bonded at C6 position to silica) exhibited slightly better chiral recognition ability than mono-2 A -azido-2 A -deoxyperphenylcarbamoylated CD CSP under the normal-phase and reversed-phase modes on the separation of 31 different racemic compounds and drugs. On the contrary, higher chiral recognition abilities were observed on the mono-2 A -azido-2 A -deoxyperacetylated CD CSP compared to mono-6 A -azido-6 A -deoxyperacetylated CD CSP.

Comparison, Applications, Advantages, and Limitations of Immobilized and Coated Amylose Tris‐(3,5‐Dimethylphenylcarbamate) Chiral Stationary Phases in HPLC

A direct high performance liquid chromatographic enantioselective separation of a set of racemic acidic drugs on the new immobilized and conventional coated amylose tris‐(3,5‐dimethylphenylcarbamate) chiral stationary phases (Chiralpak IA and Chiralpak AD, respectively) was studied using n‐hexane and 2‐propanaol (80∶20 v/v), containing TFA (0.1%) as mobile phase. The separation and elution order of the enantiomers on both columns under the same conditions were compared. The effect of the immobilization of the amylose tris‐(3,5‐dimethylphenylcarbamate) chiral stationary phase on silica (Chiralpak IA) on the chiral recognition ability was noted, as the coated phase (Chiralpak AD) possesses a higher resolving power than the immobilized one (Chiralpak IA). A few racemates, which were not or poorly resolved on the immobilized Chiralpak IA were most efficiently resolved on the coated Chiralpak AD. However, the immobilized phase withstands the prohibited HPLC solvents such as dichloromethane, ethyl acetate, tetrahydrofuran, and others when used as eluents or as a dissolving agent for the analyte itself. The versatility of the immobilized Chiralpak IA in monitoring reactions performed in dichloromethane using direct analysis techniques without further purification, workup, or removal of dichloromethane, was studied on a representative example consisting of the lipase‐catalyzed enantioselective esterification of flurbiprofen, with n‐butanol in dichloromethane as organic solvent.

Comparison, Applications, Advantages, and Limitations of Immobilized and Coated Amylose Tris‐(3,5‐Dimethylphenylcarbamate) Chiral Stationary Phases in HPLC

A direct high performance liquid chromatographic enantioselective separation of a set of racemic acidic drugs on the new immobilized and conventionally coated amylose tris‐(3,5‐dimethylphenylcarbamate) chiral stationary phases (Chiralpak IA and Chiralpak AD, respectively) was studied using n‐hexane and 2‐propanaol (80∶20 v/v) containing TFA (0.1%) as mobile phase. The separation and elution order of the enantiomers on both columns under the same conditions were compared. The effect of the immobilization of the amylose tris‐(3,5‐dimethylphenylcarbamate) chiral stationary phase on silica (Chiralpak IA), on the chiral recognition ability was noted, as the coated phase (Chiralpak AD) possesses a higher resolving power than the immobilized one (Chiralpak IA). A few racemates, which were not resolved or were poorly resolved on the immobilized Chiralpak IA were most efficiently resolved on the coated Chiralpak AD. However, the immobilized phase withstands the prohibited HPLC solvents such as dichloromethane, ethyl acetate, tetrahydrofuran, and others, when used as eluents or as a dissolving agent for the analyte itself. The versatility of the immobilized Chiralpak IA in monitoring reactions performed in dichloromethane using direct analysis techniques without further purification, workup, or removal of dichloromethane, was studied on a representative example consisting of the lipase‐catalyzed enantioselective esterification of flurbiprofen with n‐butanol in dichloromethane as organic solvent.

Chiral supercritical fluid chromatography/tandem mass spectrometry for the simultaneous determination of pindolol and propranolol in metabolic stability samples

A chiral packed-column supercritical fluid chromatographic (pSFC) system coupled to tandem mass spectrometer (MS/MS) for the rapid measurements of (R,S)-propranolol and (+/-)-pindolol in metabolic stability samples was developed. The effects of the eluent flow rates and compositions, as well as of the nebulizer temperature, on the chromatographic performance and the ionization efficiency of the analytes in positive ion mode under pSFC conditions were studied. The ionization mechanism of the analytes in the CO2/methanol atmospheric pressure chemical ionization (APCI) environments with or without the use of an additive was studied. The chiral pSFC-APCI-MS/MS approach requiring approximately 2 min per sample was applied for the simultaneous determination of two pairs of racemic drugs in in vitro samples at low nanogram per milliliter concentrations.

Enantioseparation of Some Clinically Used Drugs by Capillary Electrophoresis Using Sulfated β-Cyclodextrin as a Chiral Selector

The enantiomeric separation of 37 clinically used racemic basic drugs among 50 drugs was achieved using sulfated β-cyclodextrin (S-β-CD) as chiral selector at pH2.5 and in the reversed polarity mode. The results obtained in this study were different from the one obtained using neutral β-CD and its derivatives as chiral selectors. Using S-β-CD as chiral selector did not require the presence of the substructure 4H to achieve chiral separation as observed with β-Cyclodextrin (β-CD) and its derivatives since among the 37 separated drugs only 7 possess the 4H substructure. The chiral discrimination depends on the appropriate interaction between the analyte and the sulfated β-cyclodextrin.

Stereoselective Interactions Between Local Anesthetics and Cardiac K+ Channels

Ionic channels are membrane proteins that can be blocked by many different types of drugs such as local anesthetics, antiarrhythmics, etc. Therefore, they are considered drug targets, whose topology, at the ion channel level, has been analyzed by studying the interactions of specific ion channel blockers and site-directed mutant ion channels. Stereoselective interactions are especially interesting because they can reveal three-dimensional relationships between drugs and channels with otherwise identical biophysical and physicochemical properties. Furthermore, stereoselectivity suggests direct and specific receptor-mediated action, and identification of such stereospecific interactions may have important clinical consequences. However, ≉25% of drugs used in clinical practice are racemic mixtures, the individual enantiomers of which frequently differ in both their pharmacodynamic and pharmacokinetic profile. Furthermore, these different effects induced by one of the enantiomers of a racemic drug may contribute to the undesired effects that can be similar or different to the pharmacological effect of the racemic drug. In other cases, the enantiomers on the molecular target are opposite. In this review, we focus on the stereoselective effects of bupivacaine on different Kv channels. Bupivacaine stereoselectively blocks Kv1.5 and Kv11.1 channels, whereas non-stereoselectively it blocks Kv2.1 and Kv4.3 channels.

Kinetic studies on the intramolecular acyl migration of β-1-O-acyl glucuronides: Application to the glucuronides of (R)- and (S)-ketoprofen, (R)- and (S)-hydroxy-ketoprofen metabolites, and tolmetin by 1H-NMR spectroscopy

Conjugation of carboxylate drugs with D-glucuronic acid is of considerable interest because of the inherent reactivity of the resulting β-1-O-acyl glucuronides. These conjugates can degrade by spontaneous hydrolysis and internal acyl migration. β-1-O-acyl glucuronides and their acyl migration products can also react covalently with macromolecules with potential toxicological consequences. The spontaneous degradation of the diastereoisomeric β-1-O-acyl glucuronide metabolites of the racemic drug ketoprofen, two of its ring-hydroxylated metabolites and of tolmetin β-1-O-acyl glucuronide was investigated by 1H-NMR spectroscopy in buffer solutions, at pH 7.4 and 37°C. A plot of the logarithm of the peak integrals against time revealed first-order kinetics. Degradation rates and half-lives were calculated for each glucuronide using first-order reaction equations. Tolmetin glucuronide had the fastest degradation rate, whilst all of the ketoprofen-related glucuronides had similar degradation rates. The degradation of the diastereoisomeric glucuronides was stereoselective, with the rate for the (S)-isomer always slower compared with the (R)-isomer by approximately a factor of 2.

Elucidation of SR-propranolol transport rate and enantioselectivity through chiral activated membranes

Chiral activated membranes were prepared and tested for the enantioseparation of the racemic drug SR-propranolol. Polysulfone (PS) based membranes were investigated for this purpose, and N-Hexadecyl- l-hydroxyproline (HHP) was tested as a chiral carrier. The prepared membranes were characterized using two different membrane modules. In both cases, the concentration profile of both enantiomers of propranolol in the aqueous solutions over time is measured using the technique of capillary electrophoresis (CE). The influence of the simultaneous variation of both the concentration of the carrier in the membrane and the pH of the stripping solution, on the propranolol transport rate and the enantioselectivity across the chiral activated membranes, was studied. The propranolol transport rate through the membranes was investigated and established by response surface modelling. Clear enantioselective transport was encountered under the working conditions corresponding to the lower propranolol transport rate. In that case an alpha value of 1.7 was attained.

Enantioselective Separation of Propranolol by Chiral Activated Membranes

In this work, chiral activated membranes have been tested for the enantioseparation of the racemic drug propranolol. Polysulfone (PS)‐based membranes have been prepared for this purpose, using N‐hexadecyl‐L‐hydroxyproline (HHP) and L‐di‐n‐dodecyltartrate (L‐DDT) as chiral carriers, respectively. Kinetic experiments have been carried out using a membrane module with two rectangular channels separated by the membrane, which allows for a well‐defined hydrodynamics of the feed and stripping phases. Propranolol transport across the membrane depends on its diffusion rate from the bulk solution to the membrane surface and also on the relative amount of carrier present in the membrane. Therefore, the influence of both the solute/carrier ratio and the flow rates of the feed and stripping phases on the rate of extraction and on the selectivity of the process was evaluated for both chemical systems. Modeling of kinetic experiments has been performed, and mass transfer coefficients obtained for both systems were compared. This work has been supported by M. C. Y. T. (Project ref: PPQ2002‐04267‐C03‐01 and PPQ2002‐04201‐C02‐01) and “Fundação para a Ciência e Tecnologia” (project POCTI/FCB/43942/2001). The support by the CRUP (Portugal) and the DGICT (Spain) through the programmes “Acções Integradas Luso‐Espanholas” and “Acciones Integradas entre Espana y Portugal,” for exchange of researchers are also acknowledged. T. Gumí acknowledges the Ministerio de Educación, Cultura y Deporte for the predoctoral fellowship. Rui M.C. Viegas acknowledges the financial suport of Fundação para a Ciência e Tecnologia through the research grant PRAXIS XXI/BD/9034/96.

Enantioselective pharmacokinetics of lercanidipine in healthy volunteers

Objective: The enantioselective kinetic disposition of lercanidipine, a dihydropyridine type of third-generation calcium antagonist, was investigated in six healthy male volunteers following a single 20 mg racemic oral dose. Methods: Serial plasma samples were obtained from 0 to 24 h after drug administration. Lercanidipine enantiomers were analysed using a chiral LC–MS–MS method. Results: The following differences ( p < 0.05, Wilcoxon test) between ( S) and ( R) enantiomers were found (median): C max 2.071 ng mL −1 versus 1.681 ng mL −1; AUC 0–2412.352 ng h mL −1 versus 10.063 ng h mL −1 and Cl/ f 732.16 L h −1 versus 1891.84 L h −1. The AUC 0–∞ values for ( S)-LER were 1.21-fold higher than those for ( R)-LER. Conclusion: The pharmacokinetics of LER was enantioselective in healthy volunteers following a single dose of 20 mg of the unlabeled racemic drug.

Séparation des énantiomères : pour quelle amélioration du rapport bénéfice/risque ?

The chirality of drugs has emerged as a major theme in drug design, discovery, development and intellectual property. Single enantiomers offer some advantages and two options are available: the switch from an existing racemic drug to the single enantiomer or the development of an enantiomerically pure chiral drug. At present, the second possibility is systematically considered. The possibility of chiral switch has led to innovative drugs. However, this opportunity may be limited by several factors: the spontaneous interconversion of the two enantiomers, the pharmacokinetic profile or the selective toxicity of the most active enantiomer, and the difficulty of demonstrating the clinical superiority of one enantiomer over the racemate. Finally, the small size of the market, the policies of the drug agencies and/or the patent problem also limit the development of enantiomers. All these factors have led the pharmaceutical industry to develop new galenic forms rather than enantiomers when the patent of the racemate is expiring.

Clinical pharmacokinetic data of racemic drugs obtained by the indirect method following precolumn diastereomer formation: is the influence of racemization during chiral derivatization significant?

This report reviews the stereoselective data of a number of racemic drugs (n = 17) obtained from indirect chiral method (via diastereomer formation) in comparison to similar data generated by the application of a direct chiral method. While it was noted that the indirect method still continued to be used to characterize the stereoselective disposition of racemic drugs, the present review critically evaluates the issue of racemization that has the potential to skew the stereoselective data obtained from the indirect method. The review describes various remedies to counter and/or minimize the impact of racemization on the final outcome of the stereoselective analysis by the indirect method. On the basis of this review it could be concluded that the indirect method is a viable and important tool for gathering stereoselective data of racemic drugs used in medical practice as well for those racemic drugs still in discovery and developmental stages.

Evaluation of experimental strategies for the development of chiral chromatographic methods based on diastereomer formation.

The past two decades has seen explosive growth in the field of chirality as illustrated by the rapid progress in the various facets of this intriguing field. Firstly, it is the basic understanding of the importance of chirality and, secondly, the awareness of the therapeutic pitfalls due to failure to recognize chirality that have paved the way for a rejuvenated interest in the field. Needless to say that the impetus for chiral separation advancement and enhancement has been found to be the highest in the past decade and still continues to be an area of high focus. In this regard, both direct and indirect separation approaches have been instrumental in placing into literature stereoselective pharmacokinetic and pharmacodynamic data of plethora of drug racemates. Also, today, the cloud of uncertainty associated with the development of a chiral molecule is a thing of the past because the field is so evolved and numerous options are available for stereoselective analysis. However, the decision to advance either a single enantiomer or a racemate for development has to be made by a rational approach with adequate justification. Although indirect method of chiral separation has been well established with numerous examples of well-documented cases of stereoselective pharmacokinetic data, there is a growing need for a review that provides a strategic overview of considerations and key issues for developing chromatographic methods based on diastereomer formation. This review provides a general framework for the exploratory planning and a definitive game plan for the establishment of chiral methods based on diastereomer formation. Also, it provides an exhaustive list of applications of numerous chiral derivatization reagents that have been used in the generation of stereoselective pharmacokinetic data.

Influence of Feeding and Analytical Method on the Bioequivalence of a Racemic Drug Undergoing Enantioselective Enterohepatic Recycling

Two crossover bioequivalence trials of an enantioselectively enterohepatic recycled drug, carprofen, were conducted in dogs with the same racemic oral formulation to determine: (i) the influence of feeding patterns, and (ii) the effect of the analytical method (enantioselective vs non-enantioselective) on the statistical power of the trials. The first trial was conducted with a standard feeding protocol and the second with a special feeding protocol selected to ensure constant biliary flow into the duodenum. Using a non-enantioselective technique, 90% confidence intervals provided conclusions of bioequivalence in 100% of the cases for both area under the plasma concentration versus time curve (AUC) and maximum plasma drug concentration (Cmax) with the special feeding protocol, but only 50% for AUC and 13% for Cmax with the standard feeding protocol, suggesting that a feeding pattern that diminishes plasma drug concentration rebound for an enterohepatically recycled drug increases the power of a bioequivalence trial. Whatever the feeding protocol, an enantioselective method decreased the power of the trials for AUC but increased the power of the trials for Cmax. For an enterohepatically recycled drug, feeding pattern can influence the power of a bioequivalence trial, and the analytical technique that provides the greatest power depends on the assessed bioequivalence parameter and the feeding pattern. © 2004 Wiley-Liss, Inc. and the American Pharmacists Association