Racemic Drug Research Articles

Stereo-specific LC and LC-MS bioassays of antidepressants and psychotics.

Stereochemistry in drug action is gaining importance because the enantiomers often differ in their biological activity and pharmacokinetic profiles. The use of racemic drugs may contribute to adverse effects owing to the presence of either inactive or toxic enantiomers. Most of the drugs currently used to treat psychiatric disorders, including depression, contain one or more chiral centers and are mostly sold as racemates. Single-enantiomer drugs provide greater selectivity for their biological targets, improved therapeutic indices and better pharmacokinetics compared with racemates. Therefore it is of great importance to monitor body fluid/tissue levels of drugs used to treat depression and psychiatric disorders. The present manuscript gives an overview of liquid chromatographic and mass spectrometric techniques reported during 2000-2013 for enantiomeric separation of various classes of antidepressants, viz. selective serotonin reuptake inhibitors, serotonin and norepinephrine reuptake inhibitors, noradrenergic and specific serotonergic antidepressants, norepinephrine reuptake inhibitors, norepinephrine-dopamine reuptake inhibitors, nonamphetamine central nervous system stimulants, serotonin and dopamine inhibitors and γ-aminobutyric acid receptor agonists in biological matrices. Techniques used for extraction, separation and quantification are discussed.

Cyclodextrin‐functionalized silica nanoparticles with dendrimer‐like spacers for enantioselective capillary electrochromatography

In this report, β-cyclodextrin (β-CD)-functionalized silica nanoparticles (SNPs) with dendrimer-like spacers were synthesized by first grafting the SNPs with different generations of polyamidoamines (PAMAMs), followed by modifying the grafted SNPs with mono-6-deoxy-(p-tolylsulfonyl)-β-CD. The β-cyclodextrin-modified silica nanoparticle-cored PAMAMs (SNP-PAMAM-β-CDs) were studied as chiral pseudostationary phases for separating three racemic drugs, namely, chlorpheniramine, nefopam, and verapamil. The β-CD-functionalized SNPs with dendrimer-like spacers were more enantioselective than native β-CD, and the structures of the dendritic spacers were highly influential on the separation. In 20 mM NaH2 PO4 , addition of 4.00 mg/mL SNP-G1.0-β-CD (corresponding to 1.12 mM β-CD) could baseline separate the enantiomers of chlorpheniramine and nefopam, and introduction of 7.00 mg/mL SNP-G0-β-CD (corresponding to 1.68 mM β-CD) resulted in enantioseparation of the verapamil racemates. On the contrary, the enantiomers of nefopam and verapamil could not be resolved in the presence of 15 mM native β-CD. Our results suggest that SNP-PAMAM-β-CDs hold great promise for enantioselective separations.

Enantio-separation of drugs with multiple chiral centers by chromatography and capillary electrophoresis.

Some drugs with more than one chiral center are of high medicinal values. However, the chiral resolution of such types of racemate is a challenging task for scientists. The evolution of efficient chiral selectors is a hope for the future. Some papers have described such types of chiral resolution. The different techniques used for the purpose are HPLC, micellar electrokinetic chromatography, supercritical fluid chromatography, capillary electrochromatography and capillary electrophoresis. Owing to these facts, it was necessary to review the enantiomeric resolution of racemic drugs having more than one chiral center. The present article describes the chiral resolution of labetalol, formoterol, nadolol, indenolol, nebivolol and diltiazem using above cited techniques. In addition, attempts have been made to discuss the chiral recognition mechanism and future perspectives.

Etodolac, a racemic nonsteroidal anti‐inflammatory drug, exhibits low gastrointestinal toxicity: relation to the effects of the enantiomers (840.13)

Etodolac, a racemic (LR) nonsteroidal anti‐inflammatory drug (NSAID), has higher gastric safety compared to other conventional NSAIDs. However, it remains unknown whether the LR etodolac shows less adverse action in the small intestine. In the present study, we examined the effect of etodolac, both L/R and the enantiomers (S and R) on the gastric and small intestinal mucosa. Methods: Male SD rats were used. Etodolac (5‐50 mg/kg), both LR, S and R, was given p.o. in 18 h‐fasted or non‐fasted rats, and the stomach or the small intestine was examined for lesions 4 or 24 h later, respectively. Results: LR‐etodolac and S‐etodolac dose‐dependently caused lesions in both the stomach and small intestine, yet the effect of the former was much less potent compared to the latter. R‐etodolac did not cause damage even at 50 mg/kg. On the other hand, pretreatment with R‐etodolac dose‐dependently prevented the development of gastric lesions induced by indomethacin. The mucosal PGE2 content in the stomach and small intestine was significantly decreased by both LR‐etodolac and S‐etodolac, but not R‐etodolac. Conclusion: These results suggest that LR‐etodolac has less ulcerogenic properties in the gastrointestinal mucosa and showed that this safety profile of LR‐etodolac may be explained by the mucosal protective action of R‐etodolac.

CHIRAL SWITCH- AN EMERGING STRATEGY IN THERAPEUTICS

During the last decade, drug chirality, more specifically the use of single enantiomers versus racemic mixtures has been in the forefront of discussions in scientific forums.This is because the left and right handed twins of a molecule behave quite differently from each other in a biological environment. This can frequently lead to an improvement in pharmacological and therapeutic profile of the molecule/drug. This understanding of the significance of stereo-chemistry coupled with advances in chemical technologies and further nudged by regulatory requirements has helped the increase in the development of individual isomers at the expense of racemic mixtures.Apart from the development of novel stereo-selective compounds, a number of racemates have been re-evaluated as potential single enantiomer agents with  the possibility of an improved pharmacological/therapeutic profile. These have been termed as Chiral Switches and have resulted in the re-birth of a number of agents as single enantiomers and have provided significant improvements over the racemic drug. Economic considerations are also playing a part with pharmaceutical companies increasingly using chiral switching as a marketing strategy to increase the patent longevity and profitability period of a drug. However, not all these switches have resulted in therapeutic superiority and in many instances, unpredicted adverse reactions have resulted. Before a switch to clinical use of single enantiomers is made, physicians should satisfy themselves from evidence based on well-conducted clinical trials that the chiral switch is cost-effective and improves the outcomes for patients.  KEY WORDS: Chirality, Chiral Switch, Enantiomers

Understanding the Hysteresis Loop Conundrum in Pharmacokinetic / Pharmacodynamic Relationships

Hysteresis loops are phenomena that sometimes are encountered in the analysis of pharmacokinetic and pharmacodynamic relationships spanning from pre-clinical to clinical studies. When hysteresis occurs it provides insight into the complexity of drug action and disposition that can be encountered. Hysteresis loops suggest that the relationship between drug concentration and the effect being measured is not a simple direct relationship, but may have an inherent time delay and disequilibrium, which may be the result of metabolites, the consequence of changes in pharmacodynamics or the use of a non-specific assay or may involve an indirect relationship. Counter-clockwise hysteresis has been generally defined as the process in which effect can increase with time for a given drug concentration, while in the case of clockwise hysteresis the measured effect decreases with time for a given drug concentration. Hysteresis loops can occur as a consequence of a number of different pharmacokinetic and pharmacodynamic mechanisms including tolerance, distributional delay, feedback regulation, input and output rate changes, agonistic or antagonistic active metabolites, uptake into active site, slow receptor kinetics, delayed or modified activity, time-dependent protein binding and the use of racemic drugs among other factors. In this review, each of these various causes of hysteresis loops are discussed, with incorporation of relevant examples of drugs demonstrating these relationships for illustrative purposes. Furthermore, the effect that pharmaceutical formulation has on the occurrence and potential change in direction of the hysteresis loop, and the major pharmacokinetic / pharmacodynamic modeling approaches utilized to collapse and model hysteresis are detailed.

Predicting Nonlinear Pharmacokinetics of Omeprazole Enantiomers and Racemic Drug Using Physiologically Based Pharmacokinetic Modeling and Simulation: Application to Predict Drug/Genetic Interactions

The objective of this study is to develop a physiologically-based pharmacokinetic (PBPK) model for each omeprazole enantiomer that accounts for nonlinear PK of the two enantiomers as well as omeprazole racemic drug. By integrating in vitro, in silico and human PK data, we first developed PBPK models for each enantiomer. Simulation of racemic omeprazole PK was accomplished by combining enantiomer models that allow mutual drug interactions to occur. The established PBPK models for the first time satisfactorily predicted the nonlinear PK of esomeprazole, R-omeprazole and the racemic drug. The modeling exercises revealed that the strong time-dependent inhibition of CYP2C19 by esomeprazole greatly altered the R-omeprazole PK following administration of racemic omeprazole as in contrast to R-omeprazole given alone. When PBPK models incorporated both autoinhibition of each enantiomer and mutual interactions, the ratios between predicted and observed AUC following single and multiple dosing of omeprazole were 0.97 and 0.94, respectively. PBPK models of omeprazole enantiomers and racemic drug were developed. These models can be utilized to assess CYP2C19-mediated drug and genetic interaction potential for omeprazole and esomeprazole.

High-Performance Liquid Chromatographic Enantioseparation of Racemic Drugs Based on Homochiral Metal–Organic Framework

Homochiral metal-organic frameworks with fine-tuned pore sizes/walls and large surface areas are promising porous materials for enantioseparation considering the traditional zeolite molecular sieves have no chirality. Using enantiopure pyridyl-functionalized salen [(N-(4-Pyridylmethyl)-L-leucine·HBr)] as a starting material, we have prepared a noninterpenetrated three-dimensional homochiral metal organic framework {[ZnLBr]·H2O}n, which was further used as a chiral stationary phase for high-performance liquid chromatography to enantioseparate racemic drugs, showing excellent performances in enantioseparation of drugs. The metal-organic framework can be regarded as a novel molecular sieve-like material with a chiral separation function based on the relative sizes of the chiral channels and the resolved molecules.

Levomilnacipran (Fetzima)

In July 2013, the US Food and Drug Administration approved levomilnacipran extended release (ER; Fetzima), a serotonin-norepinephrine reuptake inhibitor, for the treatment of adults with major depressive disorder. Levomilnacipran is an active enantiomer of the racemic drug milnacipran that is currently approved in the United States for the treatment of fibromyalgia. This article provides an overview of the mechanism of action, pharmacokinetic properties, clinical efficacy, safety, and tolerability of levomilnacipran ER. Relevant information was identified through a search of databases using the key word levomilnacipran. Additional information was obtained from fda.gov, by a review of the reference lists of identified articles, and from posters and abstracts from scientific meetings. Levomilnacipran ER, dosed once daily, is generally well tolerated and has demonstrated favorable effects compared to placebo in clinical trials of patients with major depressive disorder. The increased potency for norepinephrine reuptake inhibition is a characteristic that may represent a novel contribution for levomilnacipran. Additional studies comparing levomilnacipran ER to other commonly prescribed antidepressants are needed to further evaluate its place in therapy.

Profiling of enantiopure drugs towards aryl hydrocarbon (AhR), glucocorticoid (GR) and pregnane X (PXR) receptors in human reporter cell lines

In the past decade, a large number of enantiopure drugs were introduced to clinical practice, since improved therapeutic effects were demonstrated for one of the enantiomers from originally racemic drug. While the therapeutic effects and safety of enantiopure drugs were tested prior to their approval, various biological enantiospecific activities of these, often “old” drugs, remain to be elucidated. In the current paper, we examined enantiospecific effects of clinically used enantiopure drugs containing one chiral center in the structure (i.e. zopiclone, tamsulosin, tolterodine, modafinil, citalopram) towards aryl hydrocarbon (AhR), glucocorticoid (GR) and pregnane X (PXR) receptors in human reporter cell lines.The cytotoxicity (IC50), agonist (EC50) and antagonist effects (IC50) of R-form, S-form and racemic mixture for each tested drugs were determined and compared in AhR-, GR- and PXR-gene reporter cell lines. Since AhR, GR and PXR are key regulators of drug metabolism, energy metabolism, immunity and play many other physiological functions, the data presented here might be of toxicological significance.

ChemInform Abstract: Molecular Chirality in Chemistry and Biology: Historical Milestones

AbstractReview: <175 refs.

GRAPHENE AS TUNABLE STATIONARY PHASE ADDITIVE FOR ENANTIOSEPARATION

Graphene-based biosensor and chiral sensor have made significant progress in recent years. Because of the similarity of the optical enantiomers, enantioseparation is perhaps the most subtle to achieve. Developing more effectively chiral separation techniques based on graphene is highly desirable. Herein, pristine graphene was prepared and then applicated to assist resolution of two racemic drugs of propranolol and ofloxacin using thin-layer chromatography (TLC). In comparison with TLC chiral separation by only using pure D-(-)-tartaric acid as a selector with relatively low degree of separation, a mixture of graphene and D-(-)-tartaric acid is more attractive, efficient and ready available. The high specific resolution ability for racemic compounds aided by graphene might allow its potential application in future chiral separation technologies.

Molecular modeling and spectroscopic studies on the interaction of the chiral drug venlafaxine hydrochloride with bovine serum albumin

This study was designed to examine the interaction of racemic antidepressant drug “S,R-venlafaxine hydrochloride (VEN)” with bovine serum albumin (BSA) under physiological conditions. The mechanism of interaction was studied by spectroscopic techniques combination with molecular modeling. Stern–Volmer analysis of fluorescence quenching data shows the presence of the static quenching mechanism. The thermodynamic parameters indicated that the hydrogen bonding and weak van der Waals interactions are the predominant intermolecular forces stabilizing the complex. The number of binding sites (n) was calculated. Through the site marker competitive experiment, VEN was confirmed to be located in subdomain IIIA of BSA. The binding distance (r=4.93nm) between the donor BSA and acceptor VEN was obtained according to Förster’s non-radiative energy transfer theory. According to UV–vis spectra and CD data binding of VEN leaded to conformational changes of BSA. Molecular docking simulations of S and R-VEN revealed that both isomers have similar interaction and the same binding sites, from this point of view S and R isomers are equal.

Online sample concentration in partial-filling chiral electrokinetic chromatography – mass spectrometry.

The concentration sensitivity of a racemic drug (chlorpheniramine maleate) in chiral capillary electrophoresis with electrospray ionization – mass spectrometric detection was improved ~500-fold via stacking. Enantiomeric separation was achieved through the use of a neutral chiral pseudostationary phase (2-hydroxpropyl-β-cyclodextrin), untreated fused-silica capillaries, and the application of a partial-filling technique to prevent the pseudostationary phase from entering the detector. A concentration factor of 50 resulted from field-enhanced sample injection(FESI). However, the higher concentration factor was achieved by combining FESI with micelle-to-solvent stacking (MSS) to increase sample load and focus the analyte band. MSS was achieved by injection of an ammonium lauryl sulfate micellar plug prior to sample injection. The sample diluent was a 20-fold dilution of the background electrolyte (50 mM ammonium acetate, pH 3.5) with 60% acetonitrile. This methodology provided a limit of detection (LOD) of as low as 5 ng/ml of the racemate.

Fast evaluation of enantioselective drug metabolism by electrophoretically mediated microanalysis: Application to fluoxetine metabolism by CYP2D6

In this work, a capillary electrophoretic methodology for the enantioselective in vitro evaluation of drugs metabolism is applied to the evaluation of fluoxetine (FLX) metabolism by cytochrome 2D6 (CYP2D6). This methodology comprises the in-capillary enzymatic reaction and the chiral separation of FLX and its major metabolite, norfluoxetine enantiomers employing highly sulfated β-CD and the partial filling technique. The methodology employed in this work is a fast way to obtain a first approach of the enantioselective in vitro metabolism of racemic drugs, with the additional advantage of an extremely low consumption of enzymes, CDs and all the reagents involved in the process. Michaelis-Menten kinetic parameters (Km and Vmax ) for the metabolism of FLX enantiomers by CYP2D6 have been estimated by nonlinear fitting of experimental data to the Michaelis-Menten equation. Km values have been found to be 30 ± 3 μM for S-FLX and 39 ± 5 μM for R-FLX. Vmax estimations were 28.6 ± 1.2 and 34 ± 2 pmol·min(-1) ·(pmol CYP)(-1) for S- and R-FLX, respectively. Similar results were obtained using a single enantiomer (R-FLX), indicating that the use of the racemate is a good option for obtaining enantioselective estimations. The results obtained show a slight enantioselectivity in favor of R-FLX.

Combination therapy in a transgenic model of Alzheimer's disease

The pathological accumulation of the β-amyloid protein (Aβ) has been closely associated with synaptic loss and neurotoxicity contributing to cognitive dysfunction in Alzheimer's disease (AD). Oligomers of Aβ42 appear to be the most neurotoxic form. Two of the most promising attempts to reduce Aβ accumulation have been with scyllo-inositol, an inositol steroisomer, that stabilizes Aβ42 peptide and prevents it from progressing to oligomers and fibrils and R-flurbiprofen, a purified enantiomer of the classical racemic non-steroidal anti-inflammatory drugs (NSAID), flurbiprofen, that retains the ability to specifically lower Aβ42. In the present study we evaluated the effects of scyllo-inositol and the combination treatment of scyllo-inositol+R-flurbiprofen on amyloid pathology and hippocampal-dependent memory function in 5XFAD mice, a model of Aβ pathology characterized by an enormous production of Aβ42. Our expectations were that the combination treatment of scyllo-inositol+R-flurbiprofen would have an additive effect in preventing Aβ accumulation and that cognition would be improved. Mice treated with scyllo-inositol exhibit 41 and 35% reduction in the deposition of the amyloid plaques stained by antibody against Aβ42 and Aβ40 respectively. Scyllo-inositol was not more effective when combined with R-flurbiprofen for the measures tested. Scyllo-inositol treated mice performed significantly better at the radial arm water maze (RAWM) task than untreated and scyllo-inositol+R-flurbiprofen treated mice.

Microfluidic chip capillary electrophoresis coupled with electrochemiluminescence for enantioseparation of racemic drugs using central composite design optimization

Optimization based on central composite design (CCD) for enantioseparation of anisodamine (AN), atenolol (AT), and metoprolol (ME) in human urine was developed using a microfluidic chip-CE device. Coupling the flexible and wide working range of microfluidic chip-CE device to CCD for chiral separation of AN, AT, and ME in human urine, a total of 15 experiments is needed for the optimization procedure as compared to 75 experiments using the normal one variable at a time optimization. The optimum conditions obtained are found to be more robust as shown by the curvature effects of the interaction factors. The developed microfluidic chip-CE-ECL system with adjustable dilution ratios has been validated by satisfactory recoveries (89.5-99% for six enanotiomers) in urine sample analysis. The working range (0.3-600 μM), repeatability (3.1-4.9% RSD for peak height and 4.0-5.2% RSD for peak area), and detection limit (0.3-0.6 μM) of the method developed are found to meet the requirements for bedside monitoring of AN, AT, and ME in patients under critical conditions. In summary, the hyphenation of CCD with the microfluidic chip-CE device is shown to offer a rapid means for optimizing the working conditions on simultaneous separation of three racemic drugs using the microfluidic chip-CE device developed.

Chiral Switch Drugs for Asthma and Allergies: True Benefit or Marketing Hype

Enantiomers are one of several possible molecular configurations present in a drug that has at least 1 chiral center. A drug containing 2 or more enatiomers is called a racemic mixture. Enatiomers are being developed from racemic mixtures as drugs in their own right often to extend patent protection of highly popular drugs. However, the therapeutic advantages of single enantiomer drugs developed for respiratory use such as levalbuterol, arformoterol, and levocetirizine over their racemate has been disappointing. In addition, single enantiomer drugs may be several fold more expensive than the racemic drug. New single enantiomer drugs, which are stable (no interconversion back to the racemate) and have fewer adverse effects and a more predictable pharmacodynamic or pharmacokinetic profile would confer a therapeutic advantage and thus would be beneficial for clinical use.

Molecular Chirality in Chemistry and Biology: Historical Milestones

AbstractBeginning early in the 19th century, developments in crystallography, optics, and chemistry in France set the stage for the discovery of molecular chirality by Louis Pasteur in 1848. He found that the crystallization of the sodium ammonium salt of ‘paratartaric acid’, a mysterious ‘isomer’ of natural (+)‐tartaric acid (TA), produced two different crystal types that were non‐superimposable mirror‐image forms of each other. He separated the two types and found their optical rotations in solution opposite in direction and equal in absolute magnitude. This led him to conclude that paratartaric acid is a combination of two mirror‐image molecule types of TA that are ‘dissymmetric’, an existing term he adapted to the connotation of today's ‘chiral’. In 1857, he found that the two enantiomers of TA were metabolized by a microorganism at drastically different rates, and thereby discovered biological enantioselectivity. In 1886, Italian chemist Arnaldo Piutti discovered D‐asparagine and found that it tasted intensely sweet, in contrast to the known L‐asparagine which had no taste. This was the discovery of stereoselectivity at biological receptors. As a result of advances in stereoselective synthesis and enantioselective chromatography during the last decades of the 20th century, in the 1990s the importance of molecular chirality in drug action and disposition began to receive serious attention from drug‐regulatory authorities and the pharmaceutical industry, the overall result of which has been the near‐complete disappearance of racemic drugs as newly introduced pharmaceuticals.

L-364,373 (R-L3) enantiomers have opposite modulating effects on IKs in mammalian ventricular myocytes

Activators of the slow delayed rectifier K⁺ current (IKs) have been suggested as promising tools for suppressing ventricular arrhythmias due to prolongation of repolarization. Recently, L-364,373 (R-L3) was nominated to activate IKs in myocytes from several species; however, in some studies, it failed to activate IKs. One later study suggested opposite modulating effects from the R-L3 enantiomers as a possible explanation for this discrepancy. Therefore, we analyzed the effect of the RL-3 enantiomers on IKs in ventricular mammalian myocytes, by applying standard microelectrode and whole-cell patch-clamp techniques at 37 °C. We synthesized 2 substances, ZS_1270B (right) and ZS_1271B (left), the 2 enantiomers of R-L3. In rabbit myocytes, ZS_1270B enhanced the IKs tail current by approximately 30%, whereas ZS_1271B reduced IKs tails by 45%. In guinea pig right ventricular preparations, ZS_1270B shortened APD90 (action potential duration measured at 90% repolarization) by 12%, whereas ZS_1271B lengthened it by approximately 15%. We concluded that R-L3 enantiomers in the same concentration range indeed have opposite modulating effects on IKs, which may explain why the racemic drug R-L3 previously failed to activate IKs. ZS_1270B is a potent IKs activator, therefore, this substance is appropriate to test whether IKs activators are ideal tools to suppress ventricular arrhythmias originating from prolongation of action potentials.