Achiral Environment Research Articles

Two‐dimensional liquid chromatography for direct chiral separations: a review

Separation of enantiomers remains a challenge owing to their identical physical and chemical properties in an achiral environment, and research on specialized separation techniques such as multidimensional achiral-chiral liquid chromatography continues to resolve individual enantiomers in complex samples. Recent advances in application of multidimensional liquid chromatography applied to chiral analysis are reviewed. For this reason, benefits of achiral-chiral coupling are shown, with emphasis in applications on biological and pharmaceutical fields as well as pesticide analysis. A description of standard instrumental setup in both heart-cut and comprehensive multidimensional liquid chromatography is shown. The most broadly used chiral stationary phases for multidimensional liquid chromatography are summarized. An extensive overview of different interface designs applied to complex samples is presented.

Supercritical fluid chromatography in research laboratories: Design, development and implementation of an efficient generic screening for exploiting this technique in the achiral environment

A new strategy for exploiting SFC in the achiral arena has been designed, developed and successfully implemented in our Analytical Technologies laboratories, in support of Drug Discovery. The development of a generic analytical SFC/MS screening protocol, following extensive evaluation of different stationary phase, modifier and gradient elution combinations, has enabled fast analytical method development for structurally diverse compounds within both specialised and high-throughput purification environments. Application of this approach into the daily routine has provided data on a large number and a wide variety of reaction mixtures. The results confirm the scope and generality of the strategy with more than 85% of targets being successfully resolved from their impurities.

Accuracy and sensitivity of the determination of rate constants of interconversion in achiral and chiral environments by dynamic enantioselective electrophoresis

The method for determination of rate constants of interconversion of enantiomers in chiral and achiral environments of a dynamic enantioseparation system was investigated in order to reveal its accuracy, sensitivity and robustness. Two significantly different enantioseparation systems were selected, one with a single (well-defined) chiral selector (CS) and the second with a mixture of CSs, and the rate constants of interconversion for these two systems were compared statistically. While the rate constants of interconversion in the chiral environment were found to be significantly different, the rate constants in achiral environment were confirmed to be statistically the same. The accuracy of the method was independent of experimental conditions. Influence of a CS and temperature on the rate of interconversion were discussed within the scope of determined thermodynamic parameters and statistical evaluation. A certain temperature may exist at which two different types of CSs influence the rate of interconversion equally while the extent of their influence may largely differ at other temperatures.

The Absolute Configuration of Palladium(II) and Ruthenium(II) Pseudochiral Centers in either Chiral or Achiral Environments

The meso-dithioxamide H-((R)-1-(1-phenyl)ethyl)-NSC-CSN-((S)-1-(1-phenyl)ethyl)-H (H(2)-mesoDTO) bonds [(η(6)-p-cymene)chlorido-ruthenium(II)](+) or [(η(3)-allyl)-palladium(II)](+) fragment and provides the C(s) symmetrical complexes [(η(6)-p-cymene)ClRu(H-mesoDTO κ-S,S Ru)] (1) and [(η(3)-allyl)palladium(H-mesoDTO κ-S,S Pd)] (2). These complexes are pseudochiral, and each of them exists as a mixture of two symmetrical meso forms. The improper symmetry of [(η(3)-allyl)palladium(H-mesoDTO κ-S,S Pd)] has been broken in two different ways: (i) by changing the symmetrical allyl moiety with a η(3)-crotyl frame or (ii) by substituting the residual amidic hydrogen in the dithiooxamidate ligand with a M(PR(3))Cl(+) fragment (M = Pd or Pt and PR(3) = triorganophosphine). As a consequence, a chiral plane is added to the pseudochiral palladium center, and two pairs of enantiomers are formed in each case. Furthermore, [(η(6)-p-cymene)chlorido-ruthenium(II)](+) and [(η(3)-allyl)-palladium(II)](+) fragments have been joined by means of the binucleating meso-dithiooxamidate ligand in a κ-S,S Ru κ-N,N Pd coordination mode. The resulting C(s)-symmetrical complex [(η(6)-p-cymene)ClRu(μ-mesoDTO κ-S,S Ru κ-N,N Pd)Pd(η(3)-allyl)] (8) possesses two pseudochiral metal centers, and it is therefore a mixture of four isomeric meso forms. All of these isomers in a chloroform solution interconvert in that both palladium and ruthenium invert their configurations. A mechanism of epimerization for both palladium and ruthenium is proposed. The absolute configurations of pseudochiral palladium in [(η(3)-allyl)(c)-Pd(μ-((R)-1-(1-phenyl)ethyl)-NSC-CSN-((S)-1-(1-phenyl)ethyl) κ-N,N (c)-Pd κ-S,S (A,C)-Pd)(A,C)-Pd(tri(n)propyl-phosphine)Cl] (6) and of pseudochiral palladium and ruthenium in [(η(3)-allyl)(c)-Pd(μ-((R)-1-(1-phenyl)ethyl)-NSC-CSN-((S)-1-(1-phenyl)ethyl) κ-N,N (c)-Pd κ-S,S(s)-Ru)(s)-Ru(η(6)-isopropyltoluene)Cl] (8) are provided. A suitable stereochemical notation is proposed for bimetallic complexes containing pseudochiral centers in either a chiral or an achiral environment.

Chiral Polychlorinated Biphenyl Transport, Metabolism, and Distribution: A Review

Chirality can be exploited to gain insight into enantioselective fate processes that may otherwise remain undetected because only biological, but not physical and chemical transport and transformation processes in an achiral environment will change enantiomer compositions. This review provides an in-depth overview of the application of chirality to the study of chiral polychlorinated biphenyls (PCBs), an important group of legacy pollutants. Like other chiral compounds, individual PCB enantiomers may interact enantioselectively (or enantiospecifically) with chiral macromolecules, such as cytochrome P-450 enzymes or ryanodine receptors, leading to differences in their toxicological effects and the enantioselective formation of chiral biotransformation products. Species and congener-specific enantiomer enrichment has been demonstrated in environmental compartments, wildlife, and mammals, including humans, typically due to a complex combination of biotransformation processes and uptake via the diet by passive diffusion. Changes in the enantiomer composition of chiral PCBs in the environment have been used to understand complex aerobic and anaerobic microbial transformation pathways, to delineate and quantify PCB sources and transport in the environment, to gain insight into the biotransformation of PCBs in aquatic food webs, and to investigate the enantioselective disposition of PCBs and their methylsulfonyl PCBs metabolites in rodents. Overall, changes in chiral signatures are powerful, but currently underutilized tools for studies of environmental and biological processes of PCBs.

Stereoselectivity in Drug Metabolism: Molecular Mechanisms and Analytical Methods

About 50% of therapeutic drugs are currently administered as a racemate, a mixture of equal proportions of two enantiomers. In an achiral environment, the enantiomers of a chiral drug show identical chemical and physical properties. However, they can present different chemical and pharmacological behavior in a chiral environment such as in the body. The interaction of two enantiomers with a chiral macromolecule, such as an enzyme or receptor, is three dimensional in nature, forming diastereomeric complexes resulting in a chiral recognition process. Moreover, when administered as a racemate, two enantiomers can display the pharmacokinetic processes (absorption, distribution, metabolism and excretion) in a stereoselective manner. Among these processes, stereoselectivity plays a central role in the metabolism due to the involvement of the enzymatic system. Thus, the purpose of the current review is to present important aspects related to stereochemistry of drug metabolism, with emphasis on molecular mechanisms involved in enzyme mediated reactions, such as those catalyzed by cytochrome P450, uridine 5'-diphospho (UDP)-glucuronosyltransferases and sulfotransferases. Additionally, recent advances regarding the analysis of chiral drugs and their metabolites employing different analytical techniques (high-performance liquid chromatography-HPLC and capillary electrophoresis-CE) will also be outlined.

Model of CE enantioseparation systems with a mixture of chiral selectors: Part II. Determination of thermodynamic parameters of the interconversion in chiral and achiral environments separately

The theoretical assumption that a multi-CS enantioseparation system, the model of which was described in Part I of this work, can be treated as a separation system with only one CS is confirmed by a set of experiments. The model assumes that each individual analyte–CS interaction is fast, fully independent on other interactions and the analyte: CS ratio is 1:1 and that the analyte is present in its concentration small enough not to considerably change the concentration of free CSs. An enantioselective environment in affinity capillary electrophoresis is created using a commercially available mixture of highly sulfated β-cyclodextrines as chiral selectors (CSs) and lorazepam as an analyte that undergoes interconversion during the separation process. Dependencies of the electrophoretic mobilities of the two enantiomers on concentration of the CSs mixture are proved to follow the proposed multi-CS model. Global rate constants of interconversion are determined at various temperatures and concentrations of the CSs mixture. In accord with the proposed theory, linear dependencies of the global rate constants on CSs concentration are achieved. Intercepts and slopes of these plots correspond to local rate constants of interconversion in achiral (without the mixture of CSs in background electrolyte (BGE)) and chiral (with the CSs mixture in BGE) environments, respectively. The experimentally obtained electropherograms show an excellent fit with those resulting from the computer simulation based on our model.

Molecular Basis for Water-Promoted Supramolecular Chirality Inversion in Helical Rosette Nanotubes

Helical rosette nanotubes (RNTs) are obtained through the self-assembly of the GwedgeC motif, a self-complementary DNA base analogue featuring the complementary hydrogen bonding arrays of both guanine and cytosine. The first step of this process is the formation of a 6-membered supermacrocycle (rosette) maintained by 18 hydrogen bonds, which then self-organizes into a helical stack defining a supramolecular sextuple helix whose chirality and three-dimensional organization arise from the chirality, chemical structure, and conformational organization of the GwedgeC motif. Because a chiral GwedgeC motif is predisposed to express itself asymmetrically upon self-assembly, there is a natural tendency for it to form one chiral RNT over its mirror image. Here we describe the synthesis and characterization of a chiral GwedgeC motif that self-assembles into helical RNTs in methanol, but undergoes mirror image supramolecular chirality inversion upon the addition of very small amounts of water (<1% v/v). Extensive physical and computational studies established that the mirror-image RNTs obtained, referred to as chiromers, result from thermodynamic (in water) and kinetic (in methanol) self-assembly processes involving two conformational isomers of the parent GwedgeC motif. Although derived from conformational states, the chiromers are thermodynamically stable supramolecular species, they display dominant/recessive behavior, they memorize and amplify their chirality in an achiral environment, they change their chirality in response to solvent and temperature, and they catalytically transfer their chirality. On the basis of these studies, a detailed mechanism for supramolecular chirality inversion triggered by specific molecular interactions between water molecules and the GwedgeC motif is proposed.

Precolumn diastereomerization and micellar electrokinetic chromatography on a plastic microchip: Rapid chiral analysis of amino acids

Precolumn derivatization and chiral separation of DL-amino acids based on diastereomerization have been performed on an integrated poly(dimethylsiloxane) microchip. Diastereomeric derivatives were formed in a microfabricated precolumn reactor by the reaction of amino acid enantiomers with o-phthaldialdehyde/2,3,4,6-tetra-O-acetyl-1-thio-beta-D-glucopyranose (OPA/TATG), and separated by MEKC in an achiral environment without chiral selectors in the running buffer. Optimized precolumn reactions and chiral separations of amino acids were achieved within 2.5 min. Resolutions of diastereomers of OPA/TATG-amino acids were in the range of 2.5-6.1 at optimized separation conditions. Simultaneous separation of a mixture of five chiral amino acids was successfully performed in a single run in less than 100 s.

An Engaging Illustration of the Physical Differences among Menthol Stereoisomers

The differences and similarities in the physical behavior of enantiomers and diastereomers can easily be demonstrated using the commercial stereoisomers (?)-menthol, (+)-menthol, (+)-isomenthol, and (+)-neomenthol. Thin-layer chromatography and melting point determinations clearly show that diastereomers have different physical properties from enantiomers and each other, but that enantiomers have identical physical properties in achiral environments. By obtaining a mixed melting point and optical rotations the difference in enantiomers can be observed. The exercise serves to reinforce the consequences of stereochemistry in terms of physical properties, as well as relating the differences in structure that lead to different types of stereoisomers. An attention-getting demonstration, suitable for the classroom or laboratory, of the melting point depression of a mixture of two enantiomers is also featured.

Amphiphilic Homochiral Oligopeptides Generated via Phase Separation of Nonracemic α‐Amino Acid Derivatives and Lattice‐Controlled Polycondensation in a Phospholipid Environment

AbstractRacemic S‐ethyl thioesters of Nε‐stearoyllysine (= S‐ethyl (R,S)‐2‐amino‐6‐(stearoylamino)hexanethioate) and S‐ethyl thioesters of γ‐stearyl glutamic acid (=stearyl (R,S)‐4‐amino‐5‐(ethylsulfanyl)‐5‐oxopentanoate) self‐assemble as separated two‐dimensional crystalline monolayers within an achiral phospholipid environment of racemic 1,2‐dipalmitoylglycerol (DPG) and 1,2‐dipalmitoylglycero‐3‐phosphoethanolamine (DPPE), as demonstrated by grazing‐incidence X‐ray‐diffraction (GIXD) measurements performed on the surface of H2O. Lattice‐controlled polycondensation within these crystallites with deuterium‐enantiolabeled monomers was initiated by injecting aqueous solutions of Ag+ or I2/KI beneath the monolayers, which yielded mixtures of diastereoisomeric oligopeptides containing up to six to eight repeating units, as analyzed by MALDI‐TOF mass spectrometry. Analysis of the diastereoisomeric distribution showed an enhanced relative abundance of the oligopeptides with homochiral sequences containing three or more repeating units. Within the DPPE monolayers, the nucleophilic amino group of the phospholipid operates as an initiator of polymerization at the periphery of the monomer two‐dimensional crystallites. Enhanced relative abundance of enantiomerically enriched homochiral oligopeptides was obtained by the polycondensation of nonracemic monomers. This enhancement indicated a phase separation into racemic and enantiomorphous monomer crystallites within the phospholipid environment, although this separation could not be observed directly by GIXD. A possible role that might have been played by crystalline assemblies for the abiotic generation and amplification of oligopeptides with homochiral sequences is discussed.

The significance of induced circular dichroism.

[reaction: see text] Achiral molecules or ions can display induced circular dichroism (ICD) within their absorption bands on association with chiral inducing molecules. The reverse process is equally important, although less appreciated: chiral inducing molecules, such as tartrates, can show significantly differing CD bands in different achiral environments. Therefore true ICD due to achiral molecules can only be demonstrated in the spectral region outside the absorption bands of the chiral inducing molecules. The case of tartrate-trisimidazoline helical assembly in aqueous solvent is discussed.

Pharmacology of chiral compounds: 2-arylpropionic acid derivatives.

Molecules exist as three dimensional structures. Therefore they can exist in symmetrical and asymmetrical forms. Molecules with an asymmetric centre are chiral. If the molecule and its mirror image are non-superimposable, the relationship between the two molecules is enantiomeric and the two stereoisomers are enantiomers. Since enantiomers have very similar or identical physicochemical properties, it is very difficult to distinguish between them in an achiral environment. However, once in a chiral environment, as in the body, they exhibit clear differences. In fact, most of the physiological processes in nature are stereospecific. Stereospecificity can occur in pharmacokinetic processes, in particular that utilise a carrier protein, receptor or enzyme. In addition, stereoselectivity occurs in pharmacodynamic processes and the differences between enantiomers can be either qualitative and quantitative. 2-arylpropionic acid derivatives (2APAs - profens) are an important subgroup within the class of NSAIDs. These are chiral compounds marketed as racemic mixtures. Some members of the group in an species-dependent manner undergo a special type of metabolic transformation leading to partial inversion to the optical antipode through a specific conjugation with CoA (coenzyme A) and subsequent epimerization. This metabolic inversion has not only pharmacological consequences (related to clinical effect) but also toxicological consequences such as, formation of hybrid triglycerides and even inhibition of fatty acid beta-oxidation. Differences on inversion rate between compounds and species will be discussed as well as its modification by different patho-physiologic processes such as, inflammation.

Sulfonic and Oxanilic Acid Metabolites of Acetanilide Herbicides: Separation of Diastereomers and Enantiomers by Capillary Zone Electrophoresis and Identification by 1H NMR Spectroscopy

Biological degradation of acetanilide herbicides in soil results in the formation of the ethanesulfonic acid (ESA) and oxanilic acid (OXA) derivatives. These molecules exist in two (alachlor), four (acetochlor), and eight (metolachlor) stereoisomeric forms. Due to the hindered s-cis/s-trans isomerization of the amide bond, most of the diastereomers of the ESA and OXA compounds were separable by capillary zone electrophoresis (CZE). Using γ-cyclodextrin as chiral selector in CZE, complete separation of all four isomers of enantiomerically enriched (5S)-metolachlor OXA was achieved. The enantiomers of acetochlor ESA, acetochlor OXA, and racemic metolachlor OXA were partially separated. The proton nuclear magnetic resonance (1H NMR) spectra showed separated signals for all diastereomers in achiral environment and enantiomers with γ-cyclodextrin as chiral selector. The assignment of these signals to the corresponding absolute configurations was used for the identification of observed CZE peaks. Rate constants for the amide s-cis/s-trans isomerization were evaluated. The time scales for this isomerization at room temperature are seconds for the parent herbicides and several hours for the metabolites. Rotation of the phenyl ring with respect to the amide moiety was not observed. CZE allows the determination and quantification of the different isomers of ESA and OXA metabolites and will facilitate the investigation of the probable stereoselective degradation of acetanilide herbicides in the environment.

Stopped‐flow multidimensional gas chromatography: A new method for the determination of enantiomerization barriers

With the stopped-flow multidimensional gas chromatographic technique enantiomerization barriers of ΔG#(T)=70–200 kJ/mol can easily be determined. First, the racemic mixture is separated by gas chromatography on a chiral stationary phase, then one enantiomer is transferred into a heated empty reactor column where enantiomerization is performed on-line in an achiral environment in the gas phase, and finally the enantiomerized fraction is separated in a second chiral column. The enantiomerization barrier ΔG (T) can be calculated from the enantiomerization time, the observed enantiomeric ratio, and the enantiomerization temperature. Moreover, by calculating ΔG#(T) at different temperatures, ΔH# and ΔS# are accessible. With this method atropisomeric polychlorinated biphenyls, a chiral allene, and a chiral aziridine were investigated. ©1999 John Wiley & Sons, Inc. J Micro Sep 11: 475–479, 1999

Stopped-flow multidimensional gas chromatography: A new method for the determination of enantiomerization barriers

With the stopped-flow multidimensional gas chromatographic technique enantiomerization barriers of ΔG#(T)=70–200 kJ/mol can easily be determined. First, the racemic mixture is separated by gas chromatography on a chiral stationary phase, then one enantiomer is transferred into a heated empty reactor column where enantiomerization is performed on-line in an achiral environment in the gas phase, and finally the enantiomerized fraction is separated in a second chiral column. The enantiomerization barrier ΔG (T) can be calculated from the enantiomerization time, the observed enantiomeric ratio, and the enantiomerization temperature. Moreover, by calculating ΔG#(T) at different temperatures, ΔH# and ΔS# are accessible. With this method atropisomeric polychlorinated biphenyls, a chiral allene, and a chiral aziridine were investigated. ©1999 John Wiley & Sons, Inc. J Micro Sep 11: 475–479, 1999

Pharmacological importance of stereochemical resolution of enantiomeric drugs.

Drug enantiomers have identical properties in an achiral environment, but should be considered as different chemical compounds. This is because they often differ considerably in potency, pharmacological activity and pharmacokinetic profile, since the modules with which they interact in biological systems are also optically active. Within biological systems, the metabolism of one isomer may be via a different pathway or occur at a different rate from that of the other isomer. Preferential binding of one isomer to plasma proteins may cause differences in circulating free drug and hence alter concentrations at active sites. Interactions of both isomers may differ at the active sites through which pharmacological action is mediated. Actions and levels of activity of the stereoisomers in vivo may also differ. All the pharmacological activity may reside in a single enantiomer, whereas several possibilities exist for the other enantiomer-- it may be inactive, have a qualitatively different effect, an antagonistic effect or produce greater toxicity. Two isomers may have nearly identical qualitative pharmacological activity, qualitatively similar pharmacological activity but quantitatively different potency, or qualitatively different pharmacological activity. To avoid adverse effects and optimise the therapeutic value of enantiomeric drugs, it is necessary that methods for the resolution of racemates be evolved and devolved to determine isomeric purity, establish the effectiveness of isomers of the drug, and detect the presence of an enantiomer with lower therapeutic activity and undesirable adverse effects. Even if a drug is given as a pure enantiomer, methods to discriminate between enantiomers are required because racemisation can occur both in vitro and in vivo. Methods developed for resolution of drug enantiomers should facilitate routine testing of single isomers and their metabolites, studies of pharmacological, toxicological and clinical effectiveness, routine analysis of racemates, pure enantiomers or intermediates in manufacturing processes, and investigation of the potential for inversion of an enantiopure drug substance during the early stages of drug development and therapeutic drug monitoring.

Vibrational spectroscopy of ldll-mixtures of amino acids

In equimolar mixtures of l- and dl-proline, the vibrational spectra of the ldll-mixture is not an average between that of the l- and the dl-isomers. Instead the dl-mixture is clearly intermediate between the l- and the ldll-spectra. Adding l-isomer to the dl-isomer, the solid state spectra becomes less similar to that of the l-isomer. Replacing respectively the ll-half and the dl-half with a second amino acid, each set can significantly alter the conformation of proline and/or the second amino acid, even though both sets ( ldll- and lldl-mixtures) have identical structural composition. Large changes in spectral properties between ldll- and lldl-amino acids are consistent with diastereoismeric interactions between isomers. Infrared frequencies for chiral structures in an achiral localized environment are not predictive of the frequencies for the same chemical structures in a chiral environment.

Two-group Raman optical activity revisited

In the circular differential Raman scattering observed in biological and other large polyfunctional molecules, many spectral features may be attributed to conferred chirality, in the following sense. Although a given vibrational transition may occur within a group in an intrinsically achiral local environment, it is coupling with another achiral but dissymmetrically placed group that generates the chiral response. Where two groups so coupled are chemically dissimilar, the effect originates in interference between one- and two-centre scattering mechanisms. The one-centre mechanism entails vibrational transition by a group as it undergoes conventional Raman scattering. The two-centre mechanism involves mediation of the chiral influence of a second group on this transition by Förster-type radiationless energy transfer. Where quantum-mechanical interference generates a differential Raman signal, the circular intensity differential depends on the inverse square of the distance between the groups so coupled. This distance dependence may be understood as originating from a combination of two factors. One is the linear distance dependence characterising Raman optical activity due to direct interference between transitions at distinct sites, which arises in the case of chemically identical groups. The other is the inverse cubic distance dependence associated with the probability amplitude for Förster energy migration. The Raman optical activity of any group with no chemical equivalents in its vicinity should thus be interpreted as resulting from a sum of inverse-square couplings with other chromophores. The two-group model for Raman optical activity is critically assessed, possible ways to improve upon the model are considered, and the result for the differential scattering intensity is recast in a new form that is more general and also more consise than has hitherto been presented.

Chiral selection in poly(C)-directed synthesis of oligo(G)

Theories of the origin of optical asymmetry in living systems place fundamental importance on the amplification of optical asymmetry by an autocatalytic process. The replication of a polynucleotide is one obvious choice for such an autocatalytic growth mechanism. If an optically homogeneous polynucleotide could replicate by directing the polymerization of monomers of the same handedness, while excluding monomers of the opposite handedness, its chiral descendants would come to dominate what was once an achiral environment. Recently, two highly efficient template-directed reaction systems have been developed for the oligomerization of activated guanosine mononucleotides (Fig. 1) on a poly(C) template. The synthesis of L-guanosine 5'-mononucleotide makes it possible to study chiral selection in these systems. We report here that poly(C)-directed oligomerization of activated guanosine mononucleotides proceeds readily if the monomers are of the same optical handedness as the template, and is indeed far less efficient if the monomers are of the opposite handedness. However, in template-directed reactions with a racemic mixture, monomers of the opposite handedness to the template are incorporated as chain terminators at the 2'(3') end of the products. This inhibition raises an important problem for many theories of the origin of life.