Liquid Chromatographic Enantioseparation of Newly Synthesized Fluorinated Tryptophan Analogs Applying Macrocyclic Glycopeptides-Based Chiral Stationary Phases Utilizing Core-Shell Particles.

Due to the favorable features obtained through the incorporation of fluorine atom(s), fluorinated drugs are a group with emerging pharmaceutical importance. As their commercial availability is still very limited, to expand the range of possible candidates, new fluorinated tryptophan analogs were synthesized. Control of enantiopurity during the synthesis procedure requires that highly efficient enantioseparation methods be available. In this work, the enantioseparation of seven fluorinated tryptophans and tryptophan was studied and compared systematically to (i) develop analytical methods for enantioselective separations and (ii) explore the chromatographic features of the fluorotrytophans. For enantioresolution, macrocyclic glycopeptide-based selectors linked to core-shell particles were utilized, applying liquid chromatography-based methods. Application of the polar-ionic mode resulted in asymmetric and broadened peaks, while reversed-phase conditions, together with mobile-phase additives, resulted in baseline separation for all studied fluorinated tryptophans. The marked differences observed between the methanol and acetonitrile-containing eluent systems can be explained by the different solvation abilities of the bulk solvents of the applied mobile phases. Among the studied chiral selectors, teicoplanin and teicoplanin aglycone were found to work effectively. Under optimized conditions, baseline separations were achieved within 6 min. Ionic interactions were semi-quantitatively characterized and found to not influence enantiorecognition. Interestingly, fluorination of the analytes does not lead to marked changes in the chromatographic characteristics of the methanol-containing eluents, while larger differences were noticed when the polar but aprotic acetonitrile was applied. Experiments conducted on the influence of the separation temperature indicated that the separations are enthalpically driven, with only one exception. Enantiomeric elution order was found to be constant on both teicoplanin and teicoplanin aglycone-based chiral stationary phases (L < D) under all applied chromatographic conditions.

High-performance liquid chromatographic enantioseparation of azole analogs of monoterpene lactones and amides focusing on the separation characteristics of polysaccharide-based chiral stationary phases.

High-performance liquid chromatography-based enantioseparation of newly prepared azole analogs of monoterpene lactones and amides was studied. Effects of additives and mobile phase composition were evaluated both in normal and polar organic modes. Applying amylose tris-(3,5-dimethylphenylcarbamate) selector in normal and polar organic modes acid and base additives were found to affect the peak profiles, without significantly influencing the enantiorecognition ability of the studied selector. In most cases, differences observed in retention times and enantioselectivities were lower than 10 and 20 % under normal phase and polar organic conditions, respectively. Under normal phase conditions decreased retention was observed for all the studied analytes with increased eluent polarity. Interestingly, enantioselectivity was only slightly (<10 %) influenced by the variation in the n-hexane/2-propanol ratio between 80/20 and 20/80 v/v. In polar organic mode, five different neat solvents (acetonitrile, methanol, ethanol, 1-propanol, and 2-propanol) were tested, and the best results were obtained with acetonitrile and ethanol in the case of Lux Amylose-1 column with enantioresolutions most often above 2. Based on results obtained with amylose and cellulose-based columns the amylose tris-(3,5-dimethylphenylcarbamate) selector is found to offer a superior performance both in normal and polar organic modes. When evaluating the possible effects of the selector immobilization, no striking differences were found in the normal phase. Usually, enantioselectivities and resolutions were higher (10-20 %), while retention factors of the first peaks were lower (20-30 %), on the coated-type column. In contrast, in polar organic mode, the retention characteristics and enantiorecognition ability of the coated and immobilized selectors were heavily affected by the nature of the polar solvent. Special attention has been paid to the history-dependent behavior of polysaccharide-based selectors. A confidence interval-based evaluation is suggested to help comparison of the histereticity observed in different systems. Several examples are shown to confirm that the recently discovered hysteresis is a common characteristic of polysaccharide-based selectors.

Enantioseparation of a-substituted proline analogs with macrocyclic glycopeptide-based chiral stationary phases immobilized on superficially porous particles of silica applying liquid chromatography with ultraviolet and mass spectrometric detection.

In this study, the liquid chromatography-based direct enantioseparation of the stereoisomers of α-substituted proline analogs has been investigated utilizing chiral stationary phases with UV and/or mass spectrometric (MS) detection. Macrocyclic antibiotics, such as vancomycin, teicoplanin, modified teicoplanin, and teicoplanin aglycone, all covalently immobilized to 2.7 µm superficially porous silica particles have been applied as stationary phases. Mobile phases utilizing mixtures of methanol and acetonitrile with different additives (polar-ionic mode) were optimized during method development. Best separations were achieved with mobile phases of 100% MeOH containing either 20 mM acetic acid or 20 mM triethylammonium acetate. Special attention was given to the applicability of MS-compatible mobile phases. Acetic acid was found to be advantageous as a mobile phase additive for MS detection. Enantioselective chromatographic behaviors are interpreted based on the explored correlations between the analytes' structural features and those of the applied chiral stationary phases. For the thermodynamic characterization, separations were studied in the temperature range of 5-50 °C. Generally, retention and selectivity decreased with increasing temperature, and in most cases, enthalpy-driven enantiorecognition was observed, but entropic contributions also were present. Unexpectedly, unusual shapes for the van Deemter curves were registered in the kinetic evaluations. General trends could be observed in the enantiomeric elution orders: S < R on VancoShell and NicoShell, and opposite R < S on TeicoShell and TagShell columns.

Macrocyclic glycopeptides- and derivatized cyclofructan-based chiral stationary phases for the enantioseparation of fluorinated ß-phenylalanine analogs.

The enantioseparation of five fluorinated ß-phenylalanine analogs together with the nonfluorinated α- and ß-phenylalanines has been investigated utilizing chiral stationary phases. The employed chiral selectors include macrocyclic antibiotics, such as vancomycin, teicoplanin, and teicoplanin aglycone, isopropyl carbamate functionalized cyclofructan-6, and Cinchona alkaloid-based tert.-butyl carbamate quinine, all covalently bonded to 2.7 µm superficially porous silica particles. The applied conditions included reversed-phase and polar-ionic modes where the vancomycin-, and the cyclofructan-6-based core-shell particles proved to offer suitable efficiency. Under reversed-phase conditions typical hydrophobic chromatographic behavior was observed, especially in the H2O/MeOH system. The improved selectivity with increasing MeOH content observed in polar ionic mode suggests that H-bonding may not play a major role in the chiral recognition. The stoichiometric displacement model was probed to gather information on the ionic interactions. The ion-exchange process was found to affect retention, but it has no essential contribution to chiral recognition. Without paying special attention to the optimization of the system volume of the UHPLC instrument plate heights varying in the range of 10-50 µm were obtained. In all cases, retention and selectivity decreased with increasing temperature, and enthalpy-driven enantiorecognition was observed. Elution sequences were determined in all cases.

Enantiomeric separation of newly synthesized amino, thio, and oxy derivatives of monoterpene lactones, amides, and ester applying polysaccharide-based chiral stationary phases in normal-phase mode.

New amino, thio, and oxy derivatives of monoterpene lactones, amides, and esters have been synthesized and their enantioselective separations were investigated on seven covalently immobilized polysaccharide-based chiral stationary phases. The effects of basic additives, different short-chain alcohols, and the influence of the temperature on the chromatographic behavior were studied. In addition, relationships between the structure of selector and selectand and the chromatographic parameters were explored to reveal mechanistic details of chiral recognition. Experiments were performed in the temperature range 10-50°C and thermodynamic parameters were calculated from plots of lnα versus 1/T. The separations were generally enthalpy-controlled, but entropy-driven separation was also observed. Special attention has been paid to the enantiomer elution order and several examples are shown how the structural characteristics of the selector, the nature, and the concentration of the polar modifier induce reversal of the enantiomer elution order in the case of the polysaccharide-based selectors.

Enantioselective high-performance liquid chromatographic separation of fluorinated ß- phenylalanine derivatives utilizing Cinchona alkaloid-based ion-exchanger chiral stationary phases: Enantioselective separation of fluorinated ß-phenylalanine derivatives.

The enantioselective separation of newly synthesized fluorine-substituted ß-phenylalanines has been performed utilizing Cinchona alkaloid-based ion-exchanger chiral stationary phases. Experiments were designed to study the effect of eluent composition, counterion content, and temperature on the chromatographic properties in a systematic manner. Mobile phase systems containing methanol or mixtures of methanol and acetonitrile together with acid and base additives ensured highly efficient enantioseparations. Zwitterionic phases [Chiralpak ZWIX (+) and ZWIX(-)] were found to provide superior performance compared to that by the anion-exchangers (Chiralpak QN-AX and QD-AX). A detailed thermodynamic characterization was also performed by employing van't Hoff analysis. Using typical liquid chromatographic experimental conditions, no marked effect of the flow rate could be observed on the calculated thermodynamic parameters. In contrast, a clear tendency has been revealed about the effect of the eluent composition on the thermodynamics for the zwitterionic phases.

Liquid Chromatographic Enantioseparations Utilizing Chiral Stationary Phases Based on Crown Ethers and Cyclofructans.

Natural compounds can exist in different forms, where molecules possessing chirality play an essential role in living organisms. Currently, one of the most important tasks of modern analytical chemistry is the enantioseparation of chiral compounds, in particular, the enantiomers of compounds having biological and/or pharmaceutical activity. Whether the task is to analyze environmental or food samples or to develop an assay for drug control, well-reproducible, highly sensitive, stereoselective, and robust methods are required. High-performance liquid chromatography best meets these conditions. Nevertheless, in many cases, gas chromatography, supercritical fluid chromatography, or capillary electrophoresis can also offer a suitable solution. Amino acids, proteins, cyclodextrins, derivatized polysaccharides, macrocyclic glycopeptides, and ion exchangers can serve as efficient selectors in liquid chromatography, and they are quite frequently applied and reviewed. Crown ethers and cyclofructans possessing similar structural characteristics and selectivity in the enantiodiscrimination of different amine compounds are discussed less frequently. This review collects information on enantioseparations achieved recently with the use of chiral stationary phases based on crown ethers or cyclofructans, focusing on liquid chromatographic applications.

Enantioseparation of ß2-amino acids by liquid chromatography using core-shell chiral stationary phases based on teicoplanin and teicoplanin aglycone.

Enantioseparation of nineteen ß2-amino acids has been performed by liquid chromatography on chiral stationary phases based on native teicoplanin and teicoplanin aglycone covalently bonded to 2.7 µm superficially porous silica particles. Separations were carried out in unbuffered (water/methanol), buffered [aqueous triethylammonium acetate (TEAA)/methanol] reversed-phase (RP) mode, and in polar-ionic (TEAA containing acetonitrile/methanol) mobile phases. Effects of pH in the RP mode, acid and salt additives, as well as counter-ion concentrations on chromatographic parameters have been studied. The structure of selectands (ß2-amino acids possessing aliphatic or aromatic side chains) and selectors (native teicoplanin or teicoplanin aglycone) was found to have a considerable influence on separation performance. Analysis of van Deemter plots and determination of thermodynamic parameters were performed to further explore details of the separation performance.

Enantioselective Liquid Chromatographic Separations Using Macrocyclic Glycopeptide-Based Chiral Selectors.

Numerous chemical compounds of high practical importance, such as drugs, fertilizers, and food additives are being commercialized as racemic mixtures, although in most cases only one of the isomers possesses the desirable properties. As our understanding of the biological actions of chiral compounds has improved, the investigation of the pharmacological and toxicological properties has become more and more important. Chirality has become a major issue in the pharmaceutical industry; therefore, there is a continuous demand to extend the available analytical methods for enantiomeric separations and enhance their efficiency. Direct liquid chromatography methods based on the application of chiral stationary phases have become a very sophisticated field of enantiomeric separations by now. Hundreds of chiral stationary phases have been commercialized so far. Among these, macrocyclic glycopeptide-based chiral selectors have proved to be an exceptionally useful class of chiral selectors for the separation of enantiomers of biological and pharmacological importance. This review focuses on direct liquid chromatography-based enantiomer separations, applying macrocyclic glycopeptide-based chiral selectors. Special attention is paid to the characterization of the physico-chemical properties of these macrocyclic glycopeptide antibiotics providing detailed information on their applications published recently.

Unexpected effects of mobile phase solvents and additives on retention and resolution of N-acyl-D,L-leucine applying Cinchonane-based chiral ion exchangers.

Chiral ion exchangers based on quinine (QN) and quinidine (QD), namely Chiralpak QN-AX and QD-AX as anionic and ZWIX(+) and ZWIX(-) as zwitterionic ion exchanger chiral stationary phases (CSPs) have been investigated with respect to their retention and chiral resolution characteristics. For the evaluation of the effects of the composition of the polar organic bulk solvents of the mobile phase (MP) and those of the organic acid and base additives acting as displacers necessary for a liquid chromatographic ion-exchange process, racemic N-(3,5-dinitrobenzoyl)leucine and other related analytes were applied. The main aim was to evaluate the impact of the MP variations on the observed, and thus the apparent enantioselectivity (αapp), and the retention factor. Significant differences were found using either polar protic methanol (MeOH) or polar non-protic acetonitrile (MeCN) solvents in combination with the acid and base additives as counter- and co-ions. It became clear, that the charged sites of both the chiral selectors of the CSPs and the analytes get specifically solvated, accompanied by the adsorption of all MP components on the CSP, thereby building a stagnant "stationary phase layer" with a composition different from the bulk MP. Via a systematic change of the MP composition, trends of resulting αapp and retention factors have been identified and discussed. In a detailed set of experiments, the effect of the concentration of the acid component in the MP containing MeOH or MeCN was specifically investigated, with the acid considered to be a displacer in anion-exchange type chromatographic systems. Surprisingly, all four chiral columns retained and resolved the tested N-acyl-Leu analytes with αapp values up to 21 within a retention factor window of 0.03 and 10 with pure MeOH as eluent. However, using pure MeCN as eluent, an almost infinite-long retention of the acidic analyte was noticed in all cases. We suggest that the rather different thickness of the solvation shells generated by MeOH or MeCN around the charged/chargeable sites of the chiral selector determines eventually the strength of the electrostatic selector-selectand interactions. As a control experiment we included the non-chiral N-acylglycine derivatives as analyte in all cases to support the interpretations with respect to the contribution of the enantioselective and non-enantioselective retention factor increments as a part of the observed αapp.

Cinchona-alkaloid-based zwitterionic chiral stationary phases as potential tools for high-performance liquid chromatographic enantioseparation of cationic compounds of pharmaceutical relevance.

Enantiomers of cationic compounds of pharmaceutical relevance, namely tetrahydro-ß-carboline and 1,2,3,4-tetrahydroisoquinoline analogs, were separated by high-performance liquid chromatography. Separations were performed on Cinchona-alkaloid-based zwitterionic ion exchanger type chiral stationary phases applied as cation exchangers using mixtures of methanol and acetonitrile or tetrahydrofuran as bulk solvent components containing triethylammonium acetate or ammonium acetate as organic salt additives. On the zwitterionic ZWIX(+) and ZWIX(-) columns investigated, retention and enantioseparation of the studied basic analytes were influenced by the nature and concentration of the organic components of the mobile phase. The effect of organic salt additives on the retention behavior of the studied analytes can be described by the stoichiometric displacement model related to the counterion concentration. Investigations on the structure-retention relationships were performed applying different mobile phase systems for the two types of cationic analytes. For the thermodynamic characterization, parameters such as changes in standard enthalpy (Δ(ΔH°)), entropy (Δ(ΔS°)), and free energy (Δ(ΔG°)) were calculated on the basis of van't Hoff plots derived from the ln α versus 1/T curves. In most cases, enthalpy-driven enantioseparations were observed, with a consistent dependence of the calculated thermodynamic parameters on the mobile phase composition. Elution sequences of the studied compounds were determined in all cases.

High-performance liquid chromatographic evaluation of strong cation exchanger-based chiral stationary phases focusing on stationary phase characteristics and mobile phase effects employing enantiomers of tetrahydro-ß-carboline and 1,2,3,4-tetrahydroisoquinoline analogs.

In this study, we present results obtained on the enantioseparation of some cationic compounds of pharmaceutical relevance, namely tetrahydro-ß-carboline and 1,2,3,4-tetrahydroisoquinoline analogs. In high-performance liquid chromatography, chiral stationary phases (CSPs) based on strong cation exchanger were employed using mixtures of methanol and acetonitrile or tetrahydrofuran as mobile phase systems with organic salt additives. Through the variation of the applied chromatographic conditions, the focus has been placed on the study of retention and enantioselectivity characteristics as well as elution order. Retention behavior of the studied analytes could be described by the stoichiometric displacement model related to the counter-ion effect of ammonium salts as mobile phase additives. For the thermodynamic characterization parameters, such as changes in standard enthalpy Δ(ΔH°), entropy Δ(ΔS°), and free energy Δ(ΔG°), were calculated on the basis of van't Hoff plots derived from the ln α vs. 1/T curves. In all cases, enthalpy-driven enantioseparations were observed with a slight, but consistent dependence of the calculated thermodynamic parameters on the eluent composition. Elution sequences of the studied compounds were determined in all cases. They were found to be opposite on the enantiomeric stationary phases and they were not affected by either the temperature or the eluent composition.

Polysaccharide-based chiral stationary phases as efficient tools for diastereo- and enantioseparation of natural and synthetic Cinchona alkaloid analogs.

In this study, we present results obtained on the diastereo- and enantioseparation of some basic natural and synthetic Cinchona alkaloid analogs by applying liquid chromatographic (LC) and subcritical fluid chromatographic (SFC) modalities on amylose and cellulose tris-(phenylcarbamate)-based stationary phases using n-hexane/alcohol/DEA or CO2/alcohol/DEA mobile phase systems. Seven chiral stationary phases in their immobilized form were employed to explore their stereoselectivity for a series of closely related group of analytes. The most important characteristics of LC and SFC systems were evaluated through the variation of the applied chromatographic conditions (e.g., the nature and content of the alcohol modifier, the concentration of additives, temperature). The columns Chiralpak IC and IG turned out to be the best in both LC and SFC modalities. Temperature-dependence study indicated enthalpy-controlled separation in most cases; however, separation controlled by entropy was also registered.

High-performance liquid chromatographic enantioseparation of isopulegol-based ß-amino lactone and ß-amino amide analogs on polysaccharide-based chiral stationary phases focusing on the change of the enantiomer elution order.

The enantioselective separation of newly prepared, pharmacologically significant isopulegol-based ß-amino lactones and ß-amino amides has been studied by carrying out high-performance liquid chromatography on diverse amylose and cellulose tris-(phenylcarbamate)-based chiral stationary phases (CSPs) in n-hexane/alcohol/diethylamine or n-heptane/alcohol/ diethylamine mobile phase systems. For the elucidation of mechanistic details of the chiral recognition, seven polysaccharide-based CSPs were employed under normal-phase conditions. The effect of the nature of selector backbone (amylose or cellulose) and the position of substituents of the tris-(phenylcarbamate) moiety was evaluated. Due to the complex structure and solvation state of polysaccharide-based selectors and the resulting enantioselective interaction sites, the chromatographic conditions (e.g., the nature and content of alcohol modifier) were found to exert a strong influence on the chiral recognition process, resulting in a particular elution order of the resolved enantiomers. Since no prediction can be made for the observed enantiomeric resolution, special attention has been paid to the identification of the elution sequences. The comparison between the effectiveness of covalently immobilized and coated polysaccharide phases allows the conclusion that, in several cases, the application of coated phases can be more advantageous. However, in general, the immobilized phases may be preferred due to their increased robustness. Thermodynamic parameters derived from the temperature-dependence of the selectivity revealed enthalpically-driven separations in most cases, but unusual temperature behavior was also observed.

Liquid chromatographic resolution of natural and racemic Cinchona alkaloid analogues using strong cation- and zwitterion ion-exchange type stationary phases. Qualitative evaluation of stationary phase characteristics and mobile phase effects on stereoselectivity and retention.

Liquid chromatographic (LC) and subcritical fluid chromatographic (SFC) resolution of the basic natural and synthetic Cinchona alkaloid analogues has been studied. Focus has been placed on the employment of four enantiomerically structured chiral strong cation-exchangers and four chiral diastereoisomeric Cinchona alkaloid and cyclohexyl aminosulfonic acid-based zwitterionic ion-exchangers. Except for the novel, recently synthesized racemic quinine the other investigated pairs of basic analytes are diastereomeric, but often called "pseudoenantiomeric" compounds of quinine and quinidine, cinchonidine and cinchonine, 9­epi­quinine and 9­epi­quinidine. As expected, the elution order of the resolved racemic quinine was reversed for all the eight investigated enantiomeric and (pseudo)enantiomeric pairs of chiral stationary phases, whereas this was not necessarily the case for the diastereomeric pairs of the Cinchona alkaloid related analytes. Varying the type and composition of the protic (methanol) and non-protic (acetonitrile) but polar bulk solvents in combination with organic salt additives in the mobile phase the overall retention and stereoselectivity characteristics could be triggered, leading to well performing LC and SFC systems. Thus the retention behavior of the basic analytes on both the chiral cation-exchangers and the diastereomeric zwitterionic ion-exchangers, used as cation-exchangers, could be described by the stoichiometric displacement model related to the counter-ion effect of the mobile phase additives. In addition, it became obvious that the non-protic acetonitrile compared to methanol as bulk solvent lead to a significant increase in retention, which can be associated with an increased electrostatic interaction of the charged sites due to a smaller solvation shell of the solvated cationic and anionic species. Based on the chromatographic results of the systematically selected chiral analytes and stationary phases attempts were undertaken to interpret qualitatively the observed stereoselectivity phenomena.

Enantioselective resolution of biologically active dipeptide analogs by high-performance liquid chromatography applying Cinchona alkaloid-based ion-exchanger chiral stationary phases.

Sixteen pairs of enantiomeric dipeptides were separated on four chiral ion-exchanger-type stationary phases based on Cinchona alkaloids. Anion-exchangers (QN-AX, QD-AX) and zwitterionic phases [ZWIX(+)™ and ZWIX(-)™] were studied in a comparative manner. The effects of the nature and concentrations of the mobile phase solvent components and organic salt additives on analyte retention and enantioseparation were systematically studied in order to get a deeper insight into the enantiorecognition mechanism. Moreover, experiments were performed in the temperature range 10-50 °C to calculate thermodynamic parameters like changes in standard enthalpy, Δ(ΔH°), entropy, Δ(ΔS°), and free energy, Δ(ΔG°) on the basis of van't Hoff plots derived from the ln α vs. 1/T curves. Elution sequences of the dipeptides were determined in all cases and, with a few exceptions, they were found to be opposite on the pseudoenantiomeric stationary phases as of QN-AX/QD-AX and of ZWIX(+) and ZWIX(-). The stereoselective retention mechanism is based on electrostatically driven intermolecular interactions supported by additional interaction increments mainly determined by the absolute configuration of the chiral C8 and C9 atoms of the quinine and quinidine moieties.

Enantioseparation of ß-carboline, tetrahydroisoquinoline and benzazepine analogues of pharmaceutical importance: Utilization of chiral stationary phases based on polysaccharides and sulfonic acid modified Cinchonaalkaloids in high-performance liquid and subcritical fluid chromatography.

High-performance liquid chromatographic (HPLC) and subcritical fluid chromatographic (SFC) separations of the enantiomers of structurally diverse, basic ß-carboline, tetrahydroisoquinoline and benzazepine analogues of pharmacological interest were performed applying chiral stationary phases (CSPs) based on (i) neutral polysaccharides- and (ii) zwitterionic sulfonic acid derivatives of Cinchona alkaloids. The aim of this work was to reveal the influence of structural peculiarities on the enantiorecognition on both types of CSP through the investigation of the effects of the composition of the bulk solvent, the structures of the chiral analytes (SAs) and chiral selectors (SOs) on retention and stereoselectivity. As a general tendency, valid for all polysaccharide SOs studied, the increase of the concentration of the apolar component in the mobile phase (n-hexane for LC or liquid CO2 for SFC) was found to significantly increase retention, which in most cases, was accompanied with increased selectivity and resolution. In a way, similar behaviour was registered for the zwitterionic SOs. In polar ionic mode employing eluent systems composed of methanol and acetonitrile with organic acid and base additives, moderate increases in retention factor, selectivity and resolution were observed with increasing acetonitrile content. However, under SFC conditions, an extremely high increase in retention was observed with increased CO2 content, while selectivity and resolution changed only slightly. Thermodynamic parameters derived from temperature dependence studies revealed that separations are controlled by enthalpy.

High-performance liquid chromatographic and subcritical fluid chromatographic separation of α-arylated ß-carboline, N-alkylated tetrahydroisoquinolines and their bioisosteres on polysaccharide-based chiral stationary phases.

New, pharmacologically interesting chiral amino compounds, namely, stereoisomers of α-hydroxynaphthyl-ß-carboline, benz[d]azepine and benz[c]azepine analogs as well as N-α-hydroxynaphthylbenzyl-substituted isoquinolines were enantioseparated by high-performance liquid chromatographic and subcritical fluid chromatographic methods on polysaccharide-based chiral stationary phases. Separation of the stereoisomers was optimized in both subcritical fluid chromatography and normal phase liquid chromatographic modes by investigating the effects of the composition of the bulk solvent, temperature, and the structures of the analytes and selectors. Both normal phase liquid chromatography and subcritical fluid chromatography exhibited satisfactory performance, albeit with somewhat different effectiveness in the separation of the stereoisomers studied. The optimized methods offer the possibility to apply preparative-scale separations thereby enabling further pharmacological investigations of the enantiomers.

High-Performance Liquid Chromatography Enantioseparations Using Macrocyclic Glycopeptide-Based Chiral Stationary Phases: An Overview.

Since their introduction by Daniel W. Armstrong in 1994, antibiotic-based chiral stationary phases have proven their applicability for the chiral resolution of various types of racemates. The unique structure of macrocyclic glycopeptides and their large variety of interactive sites (e.g., hydrophobic pockets, hydroxy, amino and carboxyl groups, halogen atoms, aromatic moieties) are the reasons for their wide-ranging selectivity. The commercially available Chirobiotic™ phases, which display complementary characteristics, are capable of separating a broad variety of enantiomeric compounds with good efficiency, good column loadability, high reproducibility, and long-term stability. These are the major reasons for the frequent use of macrocyclic antibiotic-based stationary phases in HPLC enantioseparations.This overview chapter provides a brief summary of general aspects of antibiotic-based chiral stationary phases including their preparation and their application to direct enantioseparations of various racemates focusing on the literature published since 2004.

Cinchona Alkaloid-Based Zwitterionic Chiral Stationary Phases Applied for Liquid Chromatographic Enantiomer Separations: An Overview.

For the early 2000s, chromatographic methods applying chiral stationary phases (CSPs) became the most effective techniques for the resolution of chiral compounds on both analytical and preparative scales. High-performance liquid chromatography (HPLC) employing various types of chiral selectors covalently bonded to silica-based supports offers a state-of-the-art methodology for "chiral analysis." Although a large number of CSPs are available nowadays, the design and development of new "chiral columns" are still needed since it is obvious that in practice one needs a good portfolio of different columns to face the challenging task of enantiomeric resolutions. The development of the unique chiral anion, cation, and zwitterion exchangers achieved by Lindner and his partners serves as an expansion of the range of the efficiently applicable CSPs.In this context this overview chapter discusses and summarizes direct enantiomer separations of chiral acids and ampholytes applying zwitterionic ion exchangers derived from Cinchona alkaloids. Our aim is to provide comprehensive information on practical solutions with focus on the molecular recognition and methodological variables.