Chiral Liquid Chromatography Research Articles

Exploring the Effects of Optically Active Solvents in Chiral Chromatography on Polysaccharide-Based Columns.

Exploring the effectiveness of optically active solvents as mobile-phase modifiers in chiral liquid chromatography (LC) can offer an additional new tool to tune the chiral selectivity. Hence, the potential of l-ethyl lactate (LEL), a biobased solvent of this nature, was explored for its distinctive interactions with both the mobile phase and analytes, as anticipated from its chiral nature. The findings reveal that LEL provides distinct selectivity compared to commonly used modifiers in chiral LC. Reversed-phase LC (RPLC)-type chiral separations were therefore compared under various conditions whereby LEL was partially or completely replacing common achiral solvents such as acetonitrile (ACN) and methanol (MeOH). An increase in chiral resolution was obtained in 8 of 16 test compounds. For 5 of them a decrease was obtained, and 3 test solutes did not offer satisfactory results under any of the tested conditions on the polysaccharide columns. When LEL was combined with methanol instead of ACN, worse results were obtained, presumably due to its protic nature. Moreover, LEL demonstrates excellent compatibility with salt additives and is fully miscible with aqueous phases. Interestingly, a steeper increase in chiral resolution is observed for LEL, as compared to ACN at lower temperatures. While LEL is somewhat hindered by its higher UV absorbance, it paves the way toward more simplified chiral screening platforms, whereby chiral solutions can be found for fewer columns and greener solvents such as LEL are incorporated. Finally, to elucidate the impact of chiral interactions between the solvent and analytes, the influence of d-ethyl lactate (DEL) was compared with that of LEL. The results revealed different interactions between the stereoisomers of ethyl lactate (EL) and chiral analytes, demonstrating an influence of optically active solvents on enantioseparations.

(S)-ML-SA1 Activates Autophagy via TRPML1-TFEB Pathway.

Autophagic flux plays a crucial role in various diseases. Recently, the lysosomal ion channel TRPML1 has emerged as a promising target in lysosomal storage diseases, such as mucolipidosis. The discovery of mucolipin synthetic agonist-1 (ML-SA1) has expanded our understanding of TRPML1's function and its potential therapeutic uses. However, ML-SA1 is a racemate with limited cellular potency and poor water solubility. In this study, we synthetized rac-ML-SA1, separated the enantiomers by chiral liquid chromatography and determined their absolute configuration by vibrational circular dichroism (VCD). In addition, we focused on investigating the impact of each enantiomer of ML-SA1 on the TRPML1-TFEB axis. Our findings revealed that (S)-ML-SA1 acts as an agonist for TRPML1 at the lysosomal membrane. This activation prompts transcription factor EB (TFEB) to translocate from the cytosol to the nucleus in a dose-dependent manner within live cells. Consequently, this signaling pathway enhances the expression of coordinated lysosomal expression and regulation (CLEAR) genes and activates autophagic flux. Our study presents evidence for the potential use of (S)-ML-SA1 in the development of new therapies for lysosomal storage diseases that target TRPML1.

LC-MS analysis of chiral amino acids in human urine reveals D-amino acids as potential biomarkers for colorectal cancer

Colorectal cancer (CRC) is a common malignant tumor in the gastrointestinal tract. Changes in amino acid metabolites have been implicated in tumorigenesis and disease progression. Biomarkers on the basis of chiral amino acids, especially D-amino acids, have not been established for early diagnosis of CRC. Quantification of chiral amino acids, especially very low concentrations of endogenous D-amino acids, is technically challenging. We report here the quantification of L- and D-amino acids in urine samples collected from 115 CRC patients and 155 healthy volunteers, using an improved method. The method of chiral labeling, liquid chromatography, and tandem mass spectrometry enabled separation and detection of 28 amino acids (14 L-amino acids, 13 D-amino acids and Gly). Orthogonal partial least squares discriminant analysis identified 14 targeted variables among these chiral amino acids that distinguished the CRC from the healthy controls. Binary logistic regression analysis revealed that D-α-aminobutyric acid (D-AABA), L-alanine (L-Ala), D-alanine (D-Ala), D-glutamine (D-Gln) and D-serine (D-Ser) could be potential biomarkers for CRC. A receiver operating characteristic curve analysis of combined multi-variables contributed to an area under the curve (AUC) of 0.995 with 98.3 % sensitivity and 96.8 % specificity. A model constructed with D-AABA, D-Ala, D-Gln, and D-Ser achieved an AUC of 0.988, indicating important contributions of D-amino acids to the association with CRC. Further analysis also demonstrated that the metabolic aberration was associated with age and the development of CRC, D-methionine (D-Met) was decreased in CRC patients with age over 50, and D/L-Gln in patients at stage IV was higher than patients at stage I. This study provides the signature of D-amino acids in urine samples and offers a promising strategy for developing non-invasive diagnosis of CRC.

Optimization of Liquid Crystalline Mixtures Enantioseparation on Polysaccharide-Based Chiral Stationary Phases by Reversed-Phase Chiral Liquid Chromatography.

Enantioseparation of nineteen liquid crystalline racemic mixtures obtained based on (R,S)-2-octanol was studied in reversed-phase mode on an amylose tris(3-chloro-5-methylphenylcarbamate) (ReproSil Chiral-MIG) and a cellulose tris(3,5-dichlorophenylcarbamate) (ReproSil Chiral-MIC). These polysaccharide-based chiral stationary phase (CSP) columns for High-Performance Liquid Chromatography (HPLC) were highly effective in recognizing isomers of minor structural differences. The mobile phase (MP), which consists of acetonitrile (ACN)/water (H2O) at different volume ratios, was used. The mobile phases were pumped at a flow rate of 0.3, 0.5, or 1 mL·min-1 with a column temperature of 25 °C, using a UV detector at 254 nm. The order of the elution was also determined. The chromatographic parameters, such as resolution (Rs), selectivity (α), and the number of theoretical plates, i.e., column efficiency (N), were determined. The polysaccharide-based CSP columns have unique advantages in separation technology, and this study has shown the potential usefulness of the CSP columns in separating liquid crystalline racemic mixtures belonging to the same homologous series.

The End of the Beginning of Mechanical Stereochemistry.

ConspectusStereochemistry has played a key role in the development of synthetic chemistry for the simple reason that the function and properties of most molecules, from medicine to materials science, depend on their shape and thus the stereoisomer used. However, despite the potential for rotaxanes and catenanes to display unusual forms of stereochemistry being identified as early as 1961, this aspect of the mechanical bond remained underexplored and underexploited; until 2014 it was only possible to access chiral rotaxanes and catenanes whose stereoisomerism is solely attributable to the mechanical bond using chiral stationary phase high performance liquid chromatography, which limited their production on scale and thus inhibited the investigation of their properties and applications. Furthermore, the stereogenic units of such molecules and analogues were often poorly described, which made it hard to fully articulate both what had been achieved in the field and what problems were left to solve. Relatively recently, methods to access rotaxanes and catenanes that display mechanical stereochemistry selectively have been developed, making these intriguing structures available for study in a range of prototypical applications including catalysis, sensing, and as chiral luminophores.In this Account, we briefly discuss the history of mechanical stereochemistry, beginning in 1961 when the potential for mechanical stereoisomerism was first identified, before defining how mechanical stereochemistry arises from a structural point of view. Building on this, using simple stereochemical arguments, we confirm that the complete set of unique stereogenic units of two-component rotaxanes and catenanes have finally been identified and categorized unambiguously, with the last being identified only in 2024. After pausing to discuss some of the stereochemical curiosities that arise when molecules contain both covalent and mechanical stereogenic units, and the potential for stereoisomerism to arise due to co-conformational movement, we use our stereochemical framework to summarize our efforts to develop conceptually general approaches to [2]catenanes and [2]rotaxanes containing all of the possible mechanical stereogenic units. In particular, we highlight how the nature of a mechanical stereogenic unit affects the available strategies for their stereoselective synthesis. We finish by highlighting recent prototypical chemical applications of interlocked molecules that rely on their mechanical stereochemistry, before discussing future directions and challenges.Taken together, we propose that the transition of such molecules from being hard to make and poorly described, to being available in high stereopurity using clearly articulated methodological and stereochemical concepts suggests that the field is finally maturing. Thus, we are now coming to the end of the beginning of mechanical stereochemistry. The stage is now set for such molecules to play a functional role in a range of areas, indeed in any chemical or physical application where control over molecular shape is required.

Development and validation of a sensitive and robust chiral method for (R)-propylene oxide: From chiral gas chromatography to chiral liquid chromatography with pre-column derivatization

Propylene oxide (PO) is an important chemical commodity and a chiral building block to prepare pharmaceutical compounds. It is crucial to develop a sensitive and robust chiral method that can be used to assess the enantiomeric purity of PO and safeguard the quality and safety of pharmaceutical products in commercial quality control (QC) laboratories. A chiral gas chromatography (GC) method based on direct enantioseparation of PO enantiomers on a 50-meter Chiraldex™ A-TA open capillary column was successfully developed for determination of enantiomeric impurity (S)-PO in (R)-PO. However, it was revealed that the chiral GC method lacked adequate robustness for routine use. Consequently, a chiral high-performance liquid chromatography (HPLC) method was developed through pre-column derivatization of PO with 2-naphthalenethiol followed by enantioseparation on a Lux Cellulose-1 (250 mm × 4.6 mm, 5 µm) column. The chiral HPLC method has been demonstrated excellent specificity, sensitivity, accuracy and robustness through method validation and robustness evaluation, thus establishing its suitability for routine use in QC laboratories.

Separation of 4-substituted 5-methylpiracetam stereoisomers on polysaccharide-based chiral stationary phases

Monitoring of stereoisomer content in samples containing chiral compounds with multiple chiral centers is an important and challenging task in chiral liquid chromatography. 4-Phenyl-5-methylpiracetam, containing two chiral centers, was used as model compound to explore the stereorecognition ability of polysaccharide-based chiral selectors. Initially, the chromatographic behavior of racemic 4-phenyl-5-methylpiracetam on nineteen commercially available polysaccharide-based chiral columns was tested with a mobile phase, consisting of a mixture of ethanol and n-hexane. The obtained results were analysed, and nine chiral phases were selected for further investigation of seven newly synthesized 4-substituted 5-methylpiracetam chiral separations. Our subsequent studies showed that chiral stationary phases based on amylose tris(5-chloro-2-methylphenylcarbamate) and amylose tris[(S)-α-methylbenzylcarbamate] posess higher ability for chiral recognition of stereoisomers of the studied analytes.

Novel insights into the dependence of adsorption-desorption kinetics on particle geometry in chiral chromatography

The existence of slow adsorption-desorption kinetics in chiral liquid chromatography is common knowledge. This may significantly contribute to worsening the efficiency and kinetic performance of a chromatographic run, especially when high flow rates are employed. Many attempts and protocols have been proposed to access this term, the so-called cads\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$c_{ads}$$\\end{document}, but they are based on different (theoretical) assumptions. As a consequence, no official method is available for the estimation of the adsorption-desorption kinetics term. In this work, a novel approach to access cads\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$c_{ads}$$\\end{document} is presented. This procedure combines experimental results obtained with kinetic and thermodynamic measurements. The investigations have been performed on two zwitterionic teicoplanin chiral stationary phases (CSPs) based on 1.9 μ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\mu $$\\end{document}m fully porous and 2.0 μ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\\mu $$\\end{document}m superficially porous particles (FPPs and SPPs), using Z-D,L-Methionine as probe molecule. Kinetic studies have been performed through the combination of both stop-flow and dynamic measurements, while adsorption isotherms have been calculated through Inverse Method. This study has confirmed that, on both particle formats, analyte diffusion on the surface of the particle is negligible, meaning that adsorption is localized, and it has been demonstrated that adsorption-desorption kinetics is strongly dependent on particle geometry and, in particular, on the loading of chiral selector. These findings are fundamental not only to unravel novel aspects of the complex enantiorecognition mechanism but also to optimize the employment of CSPs for ultra-fast and preparative applications.Graphical abstract

Comprehensive Stereoselectivity Assessment of Toxicokinetics, Tissue Distribution, Cytotoxicity, and Environmental Fate of Chiral Pesticide Propiconazole.

Propiconazole (PRO) has been widely used in the treatment of fungal infection in fruits, vegetables, cereals, and seeds. In this study, a newly established chiral liquid chromatography tandem mass spectrometry method was applied to the systemic stereoselectivity evaluation of PRO enantiomers, including toxicokinetics, tissue distributions, cytotoxicity, accumulation, and degradation. Our results showed that both trans (+)-2S,4S-PRO and cis (-)-2S,4R-PRO had lower Cmax and AUC0-∞ and higher CLz/F values in plasma and lower accumulation concentrations in the liver, heart, and brain. In cytotoxic assays, cis (-)-2S,4R-PRO exhibited the lowest cytotoxicity in PC12 neuronal, N9 microglia, SH-SY5Y neuroblastoma, and MRC5 lung fibroblast cell lines. Moreover, the Eisenia fetida incubation experiment revealed that the accumulations of both trans (+)-2S,4S-PRO and cis (-)-2S,4R-PRO were higher than those of their antipodes in E. fetida. In summary, our findings first suggested that the application of cis (-)-2S,4R-PRO for agriculture would hugely reduce the environmental risk.

Chiral LC-MS/MS method development for the enhanced separation and determination of pranoprofen enantiomers and its application to a stereoselective pharmacokinetic study

The objective of this study is to develop a quick and precise chiral liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) technology that operates in reversed phase mode. Following oral administration of 5.0 mg/kg rac-pranoprofen, the suggested approach was completely validated and successfully used to the stereoselective detection of pranoprofen enantiomers in rat plasma for the first time. Both the polysaccharide derivatized chiral stationary phase and a homemade β-cyclodextrin (β-CD) derivatized based chiral column were assessed. The latter one, a per-4-chlorophenylcarbamate-β-cyclodextrin bonded chiral stationary phase (CPCDP) made in our laboratory, allowed for the highly sensitive detection, relatively short retention time and complete separation (resolution 2.0) of the pranoprofen enantiomers. A solid phase extraction (SPE) procedure on C18 cartridge was optimized for the extraction of S-(+)-, R-(−)-pranoprofen and internal standard (S-(+)-ketoprofen, IS) from rat plasma in detail. Acetonitrile with 0.1 % formic acid in aqueous phase (60:40, v:v) at a flow rate of 0.6 mL/min was the ideal mobile phase condition. The outcome of pharmacokinetic study showed that the values of maximum plasma concentration (Cmax) and the area under plasma drug concentration–time curve to infinity (AUC0-∞) for S-(+)-pranoprofen were 1.79 and 1.90 times greater than R-(−)-isomer, respectively. Altogether, this work is the first to examine the stereospecific pharmacokinetics of pranoprofen enantiomers in vivo, whose data provided potential possibility for the development of pure isomer with superior pharmacological activity and safety to replace racemic application in the clinic.

Separation and quantitation of valacyclovir enantiomers using stability‐indicating chiral liquid chromatography method with a chiral stationary phase of amylose tris‐(3,5‐dimethylphenylcarbamate)

AbstractThe present research developed and validated a new stability‐indicating technique for the stereo‐selectively enantiomers of the antiviral nucleoside analog Valacyclovir hydrochloride (VAL). The chiral separation was performed using normal‐phase high‐performance liquid chromatography (HPLC) with a chiral stationary phase consisting of amylose tris(3‐chloro‐5‐methylphenylcarbamate) and a mobile phase of “n‐hexane, methanol, ethanol, and diethylamine”, flow rate of 0.60 mL/min, column temperature of 30°C, injection volume of 10‐μL, detection wavelength of 254‐nm, and run time of 25‐min. The enantiomers (S‐enantiomer, L‐isomer, R‐enantiomer, and D‐isomer) of Valacyclovir were separated with a resolution of 4.8 and no interference. The validation parameters verified for the proposed method, linearity in a range of 0.1002–24.3486 μg/mL (0.02–4.86%) with a regression coefficient of 0.999, and the accuracy was determined with excellent recoveries ranging from 94.38%–109.97%. The concentrations established for the detection limit and quantitation limit were 0.01% and 0.02%, respectively. The forced degradation experiments were used to assess the stability‐indicating qualities. D‐Valacyclovir impurity was successfully evaluated in release and stability samples of VAL in drug substance and tablet dosage forms using the proposed normal phase chiral HPLC approach.

Comprehensive reversed-phase×chiral two-dimensional liquid chromatography coupled to quadrupole-time-of-flight tandem mass spectrometry with post-first dimension flow splitting for untargeted enantioselective amino acid analysis.

This work describes a comprehensive achiral × chiral two-dimensional liquid chromatography separation for enantioselective amino acid analysis coupled to electrospray ionization-tandem mass spectrometry detection using data-independent acquisition. Flow splitting after the first and second dimension separation was utilized for volumetric flow reduction and for enabling a multi-detector approach (with ultraviolet, fluorescence, charged aerosol, and MS detection), respectively. Derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate provided a chromophore, a fluorophore, and an efficient mass tag for efficient ionization in positive electrospray ionization-mass spectrometry. Chiral columns often have limitations in terms of their chemoselectivity, which may be a problem when complex sample mixtures with structurally related compounds need to be separated. It can be alleviated by a reversed-phase×chiral two-dimensional-liquid chromatography setup, in which the first dimension provides the chemoselectivity and a chiral tandem column constituted of quinine-carbamate derived weak anion-exchanger and zwitterionic ion-exchanger in the second dimension separation of D- and L-amino acid enantiomers. The method was used to control the stereointegrity of the therapeutic peptide octreotide. After hydrolysis, all amino acid constituents were detected with the correct configuration and composition. Some options for flow splitting and integration of destructive detectors in the first dimension separation are outlined.

Valbenazine isomers and enantiomer determination by chiral normal phase liquid chromatography.

A novel, simple, specific, rapid, enantioselective normal phase chiral high-performance liquid chromatographic method with amylose-based Chiral Pak IG-3(250 × 4.6 mM) 3.0 μM column was developed and validated for separation and quantification of isomers and enantiomer of Valbenazine. The mobile phase composed of n-Heptane, isopropyl alcohol, dichloromethane, ethanol, and diethylamine in the ratio of 70:10:15:5:0.1 (V/V/V/VV) with a gradient flow rate was applied. The injection volume was 10 μl, and detection was carried out using a photodiode array detector at 282 nM. The column compartment was set at 35°C. The resolution between the enantiomer and isomers was found to be more than 2.0. The method was linear over the concentration range of limit of quantitation to 250% for isomers and enantiomers. The method was found to be robust with column temperature. The proposed chiral method is applicable for the determination of isomers and enantiomer of Valibenazine and was successfully used in the quality control of bulk drug manufacturing and pharmaceuticals.

Temperature Responsive × Fast Chiral Comprehensive Liquid Chromatography: a New 2D-LC Platform for Resolving Mixtures of Chiral Isomers.

Chiral resolution of solutes occurring in mixtures of unrelated species is of relevance in life sciences and in pharmaceutical analysis. While this is conceptually achievable by comprehensive two-dimensional liquid chromatography (LC × LC), few approaches exist whereby the second dimension comprises the chiral separation. The latter is preferable in combination with a conventional reversed phase type of separation in the first dimension as it offers an extension of a conventional achiral analysis. The implementation of such rapid chiral analyses in the second dimension was, thus far, limited by the challenging transfer of the first dimension mobile phase to the second dimension while still achieving chiral separation. In this study, the combination of temperature-responsive and reversed-phase chiral liquid chromatography is assessed in terms of enantioselective separation of a broad range of pharmaceutical compounds. Applying temperature-responsive liquid chromatography (TRLC) in the first dimension allows for analyses to be performed under purely aqueous conditions, which then allows for complete and more generic refocusing of (organic) solutes prior to the second dimension. This offers an enhanced ability to employ fast and broad compositional gradients over the chiral dimension, which broadens the applicability of the technique. In the proposed platform, seven chiral columns (superficially porous and fully porous columns (comprising both polysaccharide and macrocyclic antibiotic phases)) and four mobile phase gradients were screened on a pharmaceutical test mixture. The platform was shown to be able to offer the necessary resolving power for the molecules at hand and offers a new approach for chiral screening of mixtures of unrelated compounds.

When van Deemter Goes Upside Down in Chiral Chromatography

The occurrence of convex-upward van Deemter curves is rare but not unusual in chiral liquid chromatography (LC). In this article, this behaviour, experimentally observed for the more retained enantiomer of a chiral sulfoxide on a polysaccharide-based chiral stationary phase (CSP), has been investigated from an experimental viewpoint, finding that this species is characterized by a very strong, localized adsorption on the CSP. The observation of the behaviour of other chiral sulfoxides with different chemical structures has suggested correlations between molecular properties and specific interactions on the CSP.

Comparison of core-shell versus fully porous particle packings for chiral liquid chromatography productivity.

A loading and productivity study was done using three racemates on vancomycin and teicoplanin-bonded chiral stationary phases of different particle formats. Two columns were packed with 2.7 μm superficially porous particles and two columns were packed with identically bonded 5 μm fully porous particles. The last two columns were packed with specially synthesized 4.5 μm vancomycin and teicoplanin superficially porous particles. The loading of different chiral compounds showed that the columns filled with 2.7-μm chiral stationary phases were inappropriate for preparative separations due to their very low permeability which precluded high flow rates. However, columns containing 4.5 μm superficially porous (core-shell) particles were as effective for small-scale preparative chiral separations as columns filled with classical 5 μm fully porous particles. Comparing the 4.5 μm superficially porous particles and 5 μm fully porous particles teicoplanin columns, the observed respective productivities of 270 and 265 mg/g chiral phase/h for 5-methyl-5-phenyl hydantoin enantiomers were obtained. Particular attention was given to the peculiar case of the mianserin enantiomeric separation on vancomycin columns that gave observed productivities of 200 and 205 mg/g chiral phase/h on the 4.5 μm superficially porous particles and 5 μm fully porous particles, respectively.

Enantioseparation and a comprehensive spectroscopic analysis of novel synthetic cathinones laterally substituted with a trifluoromethyl group

Recently, the number of structural modifications of synthetic cathinones has been growing making them the second largest group of new psychoactive substances in Europe. Although they are abused because of their various psychoactive effects, some compounds from this group also serve as pharmaceuticals. Since synthetic cathinones are chiral molecules with one chiral center, their biological, toxicological, and pharmacological properties may significantly differ according to their absolute configuration and enantiomeric excess. In this study, we have synthesized two substances bearing a pharmacologically interesting trifluoromethyl group and developed a chiral liquid chromatography method using a polysaccharide chiral stationary phase to separate the corresponding enantiomers of both these drugs. Subsequently, we utilized molecular spectroscopic methods including chiroptical (electronic circular dichroism and vibrational circular dichroism) and non-polarizable (infrared and ultraviolet absorption) spectroscopies. In combination with density functional theory calculations, we have obtained stable conformers of selected enantiomers in solution and their relative abundances, which we used to simulate their spectra. The experimental and calculated data have been used to elucidate the 3D structure of the enantiomerically pure compounds and assign the absolute configuration of all prepared compounds.

A sensitive direct chiral liquid chromatography tandem mass spectrometry method for the enantio—Selective analysis of imeglimin in formulation

A sensitive direct chiral liquid chromatography tandem mass spectrometry method for the enantio—Selective analysis of imeglimin in formulation

Microspherical polystyrene-divinylbenzene particles hybridized with eremomycin stabilized gold nanoparticles as a stationary phase for chiral liquid chromatography

A novel enantioselective adsorbent was obtained by hybridization of microspherical polystyrene-divinylbenzene (PS-DVB) macroporous particles with eremomycin-stabilized gold nanoparticles (GNPs). Macrocyclic antibiotic eremomycin was used as a stabilization agent to obtain GNPs which were then characterized by transmission electron microscope. The average diameter of obtained nanoparticles is about 16.6 nm. Eremomycin-stabilized nanoparticles were successfully embedded into the porous polymer structure with a resulting chiral selector content of 37.5 pmol/g. The obtained PS-DVB composite containing GNPs with immobilized eremomycin was studied by scanning electron microscopy and diffuse reflectance spectroscopy. The values of the specific surface area (500 m2/g) and porosity of the adsorbent (0.39 cm3/g) are measured using nitrogen adsorption at low temperatures. The obtained composite material was used as a chiral stationary phase of liquid chromatography. A good separation enantio-selectivity to amino acids, their derivatives and beta-blockers under RPC (reversed-phase) and HILIC (Hidrophilic Interaction Liquid Chromatography) mode is demonstrated. The results obtained revealed that the prepared Eremo@GNP@PS-DVB composite is promising for use as a stationary phase in HPLC.

Effect of mobile-phase modifiers on the enantioselective retention behavior of methyl mandelate with an amylose 3,5-dimethylphenylcarbamate chiral stationary phase under reversed-phase conditions.

In this study, methanol, ethanol, n-propyl alcohol, isopropyl alcohol, acetone, and tert-butanol were used as organic modifiers in reversed-phase mode chiral liquid-chromatography to systematically investigate the effects of mobile phase components on the enantioselective retention behavior of methyl mandelate with immobilized amylose 3,5-dimethylphenylcarbamate-based sorbent called Chiralpak IA. A two-site enantioselective model was used to obtain information on the recognition mechanisms by observing the dependence of the enantioselectivity and retention factor difference on the modifier content. Similar enantioselective retention behaviors were observed for all modifiers, and characteristic modifier concentration points (PL , PM , and PH ) were identified. At modifier concentrations up to PM , the weakened hydrophobic environment resulted in polymer structural relaxation, which changed the recognition mechanisms. By contrast, at concentrations beyond PH , considerably different enantioselectivity behaviors were observed, indicating that the existence of dipole-dipole interaction, which was stronger at higher modifier concentrations, contributed to the retention mechanisms. The concentrations at which these characteristic points occurred were dependent on the carbon number of the modifier molecule. Modifiers with more carbon numbers facilitated the transition in the enantioselective behaviors. These results demonstrated that the proposed method can provide a physically consistent quantitative description of enantioselective retention behavior in reversed-phase mode.