Ferrocene derivatives with planar chirality and their enantioseparation by liquid-phase techniques.

In the last decade, planar chiral ferrocenes have attracted a growing interest in several fields, particularly in asymmetric catalysis, medicinal chemistry, chiroptical spectroscopy and electrochemistry. In this frame, the access to pure or enriched enantiomers of planar chiral ferrocenes has become essential, relying on the availability of efficient asymmetric synthesis procedures and enantioseparation methods. Despite this, in enantioseparation science, these metallocenes were not comprehensively explored, and very few systematic analytical studies were reported in this field so far. On the other hand, enantioselective high-performance liquid chromatography has been frequently used by organic and organometallic chemists in order to measure the enantiomeric purity of planar chiral ferrocenes prepared by asymmetric synthesis. On these bases, this review aims to provide the reader with a comprehensive overview on the enantioseparation of planar chiral ferrocenes by discussing liquid-phase enantioseparation methods developed over time, integrating this main topic with the most relevant aspects of ferrocene chemistry. Thus, the main structural features of ferrocenes and the methods to model this class of metallocenes will be briefly summarized. In addition, planar chiral ferrocenes of applicative interest as well as the limits of asymmetric synthesis for the preparation of some classes of planar chiral ferrocenes will also be discussed with the aim to orient analytical scientists towards 'hot topics' and issues which are still open for accessing enantiomers of ferrocenes featured by planar chirality.

Comparative enantioseparation of planar chiral ferrocenes on polysaccharide-based chiral stationary phases.

Planar chiral ferrocenes are well-known compounds that have attracted interest for application in synthesis, catalysis, material science, and medicinal chemistry for several decades. In spite of the fact that asymmetric synthesis procedures for obtaining enantiomerically enriched ferrocenes are available, sometimes, the accessible enantiomeric excess of the chiral products is unsatisfactory. In such cases and for resolution of racemic planar chiral ferrocenes, enantioselective high-performance liquid chromatography (HPLC) on polysaccharide-based chiral stationary phases (CSPs) has been used in quite a few literature articles. However, although moderate/high enantioselectivities have been obtained for planar chiral ferrocenes bearing polar substituents, the enantioseparation of derivatives containing halogens, or exclusively alkyl groups, remains rather challenging. In this study, the enantioseparation of ten planar chiral 1,2- and 1,3-disubstituted ferrocenes was explored by using five polysaccharide-based CSPs under multimodal elution conditions. Baseline enantioseparations were achieved for nine analytes with separation factors (α) ranging from 1.20 to 2.92. The presence of π-extended systems in the analyte structure was shown to impact affinity of the most retained enantiomer toward amylose-based selectors, observing retention times higher than 80 min with methanol-containing mobile phases (MPs). Electrostatic potential (V) analysis and molecular dynamics (MD) simulations were used in order to study interaction modes at the molecular level.

Our research cooperation with Professor Yoshio Okamoto.

This article summarizes our cooperation with the research group of Prof. Yoshio Okamoto at Nagoya University during the period of time between 1992 and 2005. Although the text deals entirely with enantioseparations in high-performance liquid chromatography, capillary electrophoresis, and capillary electrochromatography, this is not a detailed review in any of these areas. The text highlights selected aspects of these techniques, which have been the subject of our joint research and in part their reflection in follow-up research by our and other research groups. Together with more systematically studied topics, aspects such as ultrafast separation of enantiomers, uncommonly high separation factor of enantiomers and other related issues are also addressed.

Characteristic and complementary chiral recognition ability of four recently developed immobilized chiral stationary phases based on amylose and cellulose phenyl carbamates and benzoates.

To date, various immobilized chiral stationary phases (CSPs) have been developed. The immobilized CSPs have opened up possibilities not only maintaining the high chiral recognition abilities as well as corresponding coated ones but also affording high durability to various mobile phase. This report directed to investigate enantioseparation of recently launched four immobilized CSPs with cellulose and amylose backbones under normal phase liquid chromatography conditions. Their chiral recognition abilities were compared with previously developed six immobilized CSPs. Particularly, we focused on the complementarity for chiral recognitions. Among them, amylose tris(3-chloro-5-methylphenylcarbamate) CSP, namely, CHIRALPAK IG, showed notable chiral recognition abilities to various racemates. As expected, the investigated immobilized CSPs represented remarkable durability to wide range of mobile phases, whereas the corresponding coated CSPs could not be run due to the irreversible degradation. Taking advantage of unrestricted solvent compatibility, chiral separation selectivities were improved for some racemates.

Enantioseparations of polyhalogenated 4,4'-bipyridines on polysaccharide-based chiral stationary phases and molecular dynamics simulations of selector-selectand interactions.

2'-(4-Pyridyl)- and 2'-(4-hydroxyphenyl)-TCIBPs (TCIBP = 3,3',5,5'-tetrachloro-2-iodo-4,4'-bipyridyl) are chiral compounds that showed interesting inhibition activity against transthyretin fibrillation in vitro. We became interested in their enantioseparation since we noticed that the M-stereoisomer is more effective than the P-enantiomer. Based thereon, we recently reported the enantioseparation of 2'-substituted TCIBP derivatives with amylose-based chiral columns. Following this study, herein we describe the comparative enantioseparation of both 2'-(4-pyridyl)- and 2'-(4-hydroxyphenyl)-TCIBPs on four cellulose phenylcarbamate-based chiral columns aiming to explore the effect of the polymer backbone, as well as the nature and position of substituents on the side groups on the enantioseparability of these compounds. In the frame of this project, the impact of subtle variations of analyte and polysaccharide structures, and mobile phase (MP) polarity on retention and selectivity was evaluated. The effect of temperature on retention and selectivity was also considered, and overall thermodynamic parameters associated with the analyte adsorption onto the CSP surface were derived from van 't Hoff plots. Interesting cases of enantiomer elution order (EEO) reversal were observed. In particular, the EEO was shown to be dependent on polysaccharide backbone, the elution sequence of the two analytes being P-M and M-P on cellulose and amylose tris(3,5-dimethylphenylcarbamate), respectively. In this regard, a theoretical investigation based on molecular dynamics (MD) simulations was performed by using amylose and cellulose tris(3,5-dimethylphenylcarbamate) nonamers as virtual models of the polysaccharide-based selectors. This exploration at the molecular level shed light on the origin of the enantiodiscrimination processes.

Enantioseparation of 5,5'-Dibromo-2,2'-Dichloro-3-Selanyl-4,4'-Bipyridines on Polysaccharide-Based Chiral Stationary Phases: Exploring Chalcogen Bonds in Liquid-Phase Chromatography.

The chalcogen bond (ChB) is a noncovalent interaction based on electrophilic features of regions of electron charge density depletion (σ-holes) located on bound atoms of group VI. The σ-holes of sulfur and heavy chalcogen atoms (Se, Te) (donors) can interact through their positive electrostatic potential (V) with nucleophilic partners such as lone pairs, π-clouds, and anions (acceptors). In the last few years, promising applications of ChBs in catalysis, crystal engineering, molecular biology, and supramolecular chemistry have been reported. Recently, we explored the high-performance liquid chromatography (HPLC) enantioseparation of fluorinated 3-arylthio-4,4'-bipyridines containing sulfur atoms as ChB donors. Following this study, herein we describe the comparative enantioseparation of three 5,5'-dibromo-2,2'-dichloro-3-selanyl-4,4'-bipyridines on polysaccharide-based chiral stationary phases (CSPs) aiming to understand function and potentialities of selenium σ-holes in the enantiodiscrimination process. The impact of the chalcogen substituent on enantioseparation was explored by using sulfur and non-chalcogen derivatives as reference substances for comparison. Our investigation also focused on the function of the perfluorinated aromatic ring as a π-hole donor recognition site. Thermodynamic quantities associated with the enantioseparation were derived from van't Hoff plots and local electron charge density of specific molecular regions of the interacting partners were inspected in terms of calculated V. On this basis, by correlating theoretical data and experimental results, the participation of ChBs and π-hole bonds in the enantiodiscrimination process was reasonably confirmed.

Chiral separation of novel iminonaringenin derivatives.

A series of 4-iminonaringenin derivatives 2-6 have been prepared in good overall yields from a condensation reaction between naringenin and primary amines. The structures of all products were confirmed by ultraviolet, infrared, proton nuclear magnetic resonance, and carbon-13 nuclear magnetic resonance spectroscopic techniques. These derivatives were analyzed by high-performance liquid chromatography using polysaccharide-based chiral stationary phases, namely, Chiralpak IB and Chiralcel OD, using various mobile phases. 2-Propanol showed a high enantioselectivity for naringin and its derivatives using achiral column containing immobilized polysaccharides (Chiralpak IB).

Chiral separation of lenalidomide by liquid chromatography on polysaccharide-type stationary phases and by capillary electrophoresis using cyclodextrin selectors.

Complementary techniques were applied for the investigation of the chiral recognition and enantiomeric resolution of lenalidomide using various cyclodextrins and polysaccharides as chiral selectors. The high-performance liquid chromatography enantioseparation of the anticancer drug was achieved using polysaccharide-type chiral stationary phases in polar organic mode. Elution order and absolute configuration were elucidated by combined circular dichroism spectroscopy and time-dependent density functional theory calculations after the isolation of pure enantiomers. Chiral selector dependent and mobile-phase dependent reversal of the enantiomer elution order was observed, and the nonracemic nature of the lenalidomide sample was also demonstrated. Eight anionic cyclodextrins were screened for their ability to discriminate between the uncharged enantiomers by using capillary electrophoresis. Only two derivatives presented chiral interactions, these cases being interpreted in terms of apparent stability constants and complex mobilities. The best results were delivered by sulfobutylether-ß-cyclodextrin, where quasi-equal stability constants were recorded and the enantiodiscrimination process was mainly driven by different mobilities of the transient diastereomeric complexes. The optimized high-performance liquid chromatography (Chiralcel OJ column, pure ethanol with 0.6 mL/min flow rate, 40°C) and capillary electrophoresis methods (30 mM sulfobutylether-ß-cyclodextrin, 30 mM phosphate pH 6.5, 12 kV applied voltage, 10°C) were validated for the determination of 0.1% (R)-lenalidomide as a chiral impurity, which could be important if a racemic switch is achieved.

Screening Approach for Chiral Separation of ß-Aminoketones by HPLC on Various Polysaccharide-Based Chiral Stationary Phases.

Nine ß-aminoketones were synthesized via Mannich reaction when benzaldehyde was condensed with some primary amines and acetophenone. The purified compounds were identified by using spectroscopic methods. The enantiomeric separation of these derivatives was carried out by high-performance liquid chromatography (HPLC) using several coated and immobilized polysaccharide stationary phases, namely, Chiralcel(®) OD-H, Chiralcel(®) OD, Chiralcel(®) OJ, Chiralpak(®) AD, Chiralpak(®) IA, and Chiralpak(®) IB using different mobile phases composed of n-hexane and alcohol mixed in various ratios or pure ethanol or isopropanol. The retention behavior and selectivity of these chiral stationary phases were examined in isocratic normal phase mode. The results indicate that cellulose derivatives have higher enantioselectivity than amylose derivatives for the separation of racemic ß-amino ketones.

High-performance liquid chromatographic enantioseparation of amino alcohol analogues possessing 1,2,3,4-tetrahydroisoquinoline skeleton on polysaccharide-based chiral stationary phases.

The stereoisomers of 1,2,3,4-tetrahydroisoquinoline amino alcohol analogues and derivatives thereof were separated in normal-phase mode on chiral stationary phases based on preprepared silica coated with cellulose tris-(3,5-dimethylphenyl carbamate), cellulose tris-(3-chloro-4-methylphenyl carbamate), cellulose tris-(4-methylbenzoate) or cellulose tris-(4-chloro-3-methylphenyl carbamate). On all the investigated chiral columns, the retention and the enantioseparation were influenced by the nature and the concentrations of the mobile phase components and additives, and also the temperature. Experiments were performed in the temperature range 10-50°C. Thermodynamic parameters were calculated from plots of lnα vs 1/T. On these polysaccharide-based chiral columns, both enthalpy-driven separations and entropy-controlled enantioseparations were observed. The latter was advantageous with regard to the shorter retention and greater selectivity at high temperature. The sequence of elution of the stereoisomers was determined in all cases.

High-performance liquid chromatography enantioseparation of polyhalogenated 4,4'-bipyridines on polysaccharide-based chiral stationary phases under multimodal elution.

An investigation on the high-performance liquid chromatography enantioseparation of 12 polyhalogenated 4,4'-bipyridines on polysaccharide-based chiral stationary phases is described. The overall study was directed toward the generation of efficient separations in order to obtain pure atropisomers that will serve as ligands for building homochiral metal organic frameworks. Four coated columns--namely, Lux Cellulose-1, Lux Cellulose-2, Lux Cellulose-4, and Lux Amylose-2--and two immobilized columns--namely, Chiralpak IC and IA--were used under normal, polar organic, and reversed-phase elution modes. Moreover, Chiralcel OJ was considered under normal-phase and polar organic conditions. The effect of the chiral selector and mobile phase composition on the enantioseparation, the enantiomer elution order and the beneficial effect of nonstandard solvents were studied. The effect of water in the mobile phase on the enantioselectivity and retention was investigated and retention profiles typical of hydrophilic interaction liquid chromatography were observed. Interesting phenomena of solvent-induced enantiomer elution order reversal occurred under normal-phase mode. All the considered 4,4'-bipyridines were enantioseparated at the multimilligram level.

Three versus five micrometer chlorinated polysaccharide-based packings in chiral capillary electrochromatography: performance evaluation.

In this study, a test set of 44 nonacidic compounds was analyzed on four 3 µm chlorinated polysaccharide-based chiral stationary phases with cellulose tris (3-chloro-4-methylphenylcarbamate) (Lux Cellulose-2®; LC2), amylose tris (5-chloro-2-methylphenylcarbamate) (Lux Amylose-2®, LA2), cellulose tris (4-chloro-3-methylphenylcarbamate) (Lux Cellulose-4®; LC4) and cellulose tris (3,5-dichlorophenylcarbamate) (Sepapak-5®, Sp5) as selectors. The analysis times, retention factors, efficiencies and enantioselectivities were compared with the results obtained on their 5 µm analogs. All 3 µm packings, except for LA2, individually separated more compounds than their 5 µm analogs. When the cumulative success rates on the 3 and 5 µm packings were considered, it was observed that they were similar for both particle sizes; the combination of three or four 5 µm columns separated one compound more from the considered test set than that of the same number of 3 µm columns. Furthermore, it was observed that the 3 and 5 µm packings showed some complementarity. Only four compounds were not separated on any of the columns, while the use of only either the 3 or 5 µm columns resulted in 10 and nine not-separated compounds, respectively. The analyses on 5 µm LC2 and Sp5 were faster than on their 3 µm analogs. For LC4 the 3 µm packing showed the shortest analysis times and diverse analysis times for both particle sizes were obtained on LA2. Furthermore, three out of four 3 µm packings, that is, LC2, LC4, and Sp5, were found to be more efficient than their 5 µm analogs.

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.

Optimization of the HPLC enantioseparation of 3,3'-dibromo-5,5'-disubstituted-4,4'-bipyridines using immobilized polysaccharide-based chiral stationary phases.

The HPLC enantioseparation of nine atropisomeric 3,3',5,5'-tetrasubstituted-4,4'-bipyridines was performed in normal and polar organic (PO) phase modes using two immobilized polysaccharide-based chiral columns, namely, Chiralpak IA and Chiralpak IC. The separation of all racemic analytes, the effect of the chiral selector, and mobile phase (MP) composition on enantioseparation and the enantiomer elution order (EEO) were studied. The beneficial effect of nonstandard solvents, such as tetrahydrofuran (THF), dichloromethane (DCM), and methyl t-butyl ether on enantioseparation was investigated. All selected 4,4'-bipyridines were successfully enantioseparated on Chiralpak IA under normal or PO MPs with separation factors from 1.14 to 1.70 and resolutions from 1.3 to 6.5. Two bipyridines were enantioseparated at the multimilligram level on Chiralpak IA. Differently, Chiralpak IC was less versatile toward the considered class of compounds and only five bipyridines out of nine could be efficiently separated. In particular, on these columns, the ternary mixture n-heptane/THF/DCM (90:5:5) as MP had a positive effect on enantioseparation. An interesting phenomenon of reversal of the EEO depending on the composition of the MP for the 3,3'-dibromo-5,5'-bis-(E)-phenylethenyl-4,4'-bipyridine along with an exceptional enantioseparation for the 3,3'-dibromo-5,5'-bis-ferrocenylethynyl-4,4'-bipyridine (α = 8.33, R(s) = 30.6) were observed on Chiralpak IC.

Unravelling dispersion forces in liquid-phase enantioseparation. Part I: Impact of ferrocenyl versus phenyl groups.

BACKGROUND: Highly ordered chiral secondary structures as well as multiple (tunable) recognition sites are the keys to success of polysaccharide carbamate-based chiral selectors in enantioseparation science. Hydrogen bonds (HBs), dipole-dipole, and π-π interactions are classically considered the most frequent noncovalent interactions underlying enantioselective recognition with these chiral selectors. Very recently, halogen, chalcogen and π-hole bonds were also identified as interactions working in polysaccharide carbamate-based selectors to promote enantiomer distinction. On the contrary, the function of dispersion interactions in this field was not explored so far. RESULTS: The enantioseparation of chiral ferrocenes featuring chiral axis or chiral plane as stereogenic elements was performed by comparing five polysaccharide carbamate-based chiral columns, with the aim to identify enantioseparation outcomes that could be reasonably determined by dispersion forces, making available a reliable experimental data set for future theoretical studies to confirm the heuristic hypothesis. The effects of mobile phase polarity and temperature on the enantioseparation were considered, and potential recognition sites on analytes and selectors were evaluated by electrostatic potential (V) analysis and molecular dynamics (MD). In this first part, the enantioseparation of 3,3'-dibromo-5,5'-bis-ferrocenylethynyl-4,4'-bipyridine bearing two ferrocenylethynyl units linked to an axially chiral core was performed and compared to that of the analyte featuring the same structural motif with two phenyl groups in place of the ferrocenyl moieties. The results of this study showed the superiority of the ferrocenyl compared to the phenyl group, as a structural element favouring enantiodifferentiation. SIGNIFICANCE AND NOVELTY: Even if dispersion (London) forces have been envisaged acting in liquid-phase enantioseparations, focused studies to explore possible contributions of dispersion forces with polysaccharide carbamate-based selectors are practically missing. This study allowed us to collect experimental information that support the involvement of dispersion forces as contributors to liquid-phase enantioseparation, paving the way to a new picture in this field.

Separation of planar chiral ferrocenes by capillary electrokinetic chromatography and liquid chromatography.

Capillary electrokinetic chromatography (CEKC) and liquid chromatography (LC) methods were explored for the enantiomeric separation of six unsymmetrically disubstituted ferrocene derivatives. In normal-phase mode liquid chromatography, the use of n-heptane, ethanol or isopropanol with 1% n-butylamine as mobile phase on six polysaccharide-based columns, allowed to fully separate the enantiomers of three compounds out of the six (i.e 7-chloro-N-(2-((dimethylamino)methyl)ferrocenyl)quinolin-4-amine (ferroquine) (compound 1), 1-[(1S)-(1-Aminoethyl)]-2-(diphenylphosphino)ferrocene (compound 5) and 1-[(1R)-1-(Dicyclohexylphosphino)ethyl]-2-(diphenylphosphino)ferrocene (compound 6). Among the columns used, the Lux i-Cellulose-5 was the most effective. In capillary electrokinetic chromatography, a phosphate buffer of 25 mM concentration and pH equal to 2.5 was chosen as background electrolyte, leading to cationic ferrocene derivatives. The addition of neutral cyclodextrins was undertaken first and native ß- or γ-cyclodextrins were found to resolve the enantiomers of two derivatives. Then, 15 mM of anionic cyclodextrins were added to the background electrolyte. The use of SBE-ß-CD, S-ß-CD or S-γ-CD have allowed the separation of the enantiomers for most of the ferrocene derivatives studied with high resolution values in short migration time. For instance, for 1-(R)-2-(Diphenylphosphino)ethyldi-tert-butylphosphine ferrocene (compound 2), the migration times were less than 2 minutes and the resolution value was equal to 3.52 in short-end mode with 15 mM S-ß-CD, at 25 kV and 25°C. Finally, a dual cyclodextrins system was tested using 15 mM of S-ß-CD plus 15 mM HP-γ-CD in the phosphate buffer. This system allowed the improved separation of two ferrocene derivatives with an unusual resolution value equal to 41.5 in long-end mode. Overall, CEKC showed better enantioseparating power of the six chiral ferrocenes studied than liquid chromatography.

Comprehensive two-dimensional liquid chromatographic method (Chiral × Achiral) for the simultaneous resolution of pesticides.

This work demonstrates the potential of two-dimensional liquid chromatography (2D-LC) to increase the resolution capacity of multiple pesticides in a single analysis of samples that contain both chiral and achiral compounds. The setup is based on the combination of a chiral column in the first dimension and an achiral column in the second dimension using the on-line full comprehensive mode (LC × LC). This method was optimized for the separation of 24 pesticides (17 chiral and 7 achiral). The 2D-LC system was built with an active flow splitter pump in order to easily adjust the volume of sample transferred to the second dimension and to select and optimize independently the flow rate in the first dimension. The first-dimension optimization involved the comparison of enantioresolution abilities of three different polysaccharides chiral stationary phases as well as different elution conditions, while in the second-dimension parameters like stationary phase and organic modifier were explored. Other experimental variables that influence the two-dimensional peak capacity (orthogonality, sampling frequency, shift gradients, etc.) are discussed.

Unravelling functions of halogen substituents in the enantioseparation of halogenated planar chiral ferrocenes on polysaccharide-based chiral stationary phases: experimental and electrostatic potential analyses.

Planar chiral halogenated ferrocenes have come in useful as synthetic intermediates over the years, allowing for the preparation of functionalized derivatives for catalysis, material science, optoelectronics, and medicinal chemistry. Despite their chemical interest, few halogenated planar chiral ferrocenes have been prepared in enantiopure form by asymmetric synthesis so far. Enantioselective HPLC on polysaccharide-based chiral stationary phases (CSPs) has been used for resolving planar chiral ferrocenes making both enantiomers available. However, the enantioseparation of derivatives containing halogens or alkyl groups exclusively remains rather challenging. Given this context, in this study the enantioseparation of eleven dihalogenated planar chiral ferrocenes was systematically explored by using five polysaccharide-based CSPs under multimodal elution conditions. Baseline enantioseparations were achieved for nine analytes with separation factors (α) ranging from 1.15 to 1.66. Thermodynamic quantities associated with the enantioseparations were derived from van't Hoff plots, and for 1-halo-2-(iodoethynyl)ferrocenes (1-halogen = F, Cl, Br) halogen-dependent thermodynamic profiles were identified on a cellulose tris(3,5-dimethylphenylcarbamate)-based column. The impact of CSP structure and mobile phase (MP) polarity on the enantioseparation was evaluated. In addition, with the aim to unravel the functions of halogen substituents in mechanisms and noncovalent interactions underlying selector-selectand complex formation at molecular level, local electron charge density of specific molecular regions of the interacting partners were evaluated in terms of calculated electrostatic potential (V) and related source function (SF) contributions. On this basis, the impact of halogen type and position on the enantioseparation was investigated by correlating theoretical and experimental data.

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.

Enantiomer fraction evaluation of the four stereoisomers of second-generation anticoagulant rodenticides in biological matrices with polysaccharide-based chiral selectors and liquid chromatography tandem mass spectrometry.

Numerous cases of wildlife exposure to five second-generation anticoagulant rodenticides have been reported worldwide, and residues of these chiral pesticides in biological matrices are still quantified by achiral liquid chromatography methods. However, they are a mixture of cis- and trans-diastereomers, thus a mixture of four stereoisomers. Their persistence must be evaluated in a differentiated way in the food chain of concerned predator species in order to reduce the environmental impact. This article presents an evaluation of the chiral selectivity of five polysaccharide-based chiral selectors for the four stereoisomers of bromadiolone, difenacoum, brodifacoum, flocoumafen and difethialone. Different chromatographic parameters, influencing the chiral separation, such as organic modifier (acetonitrile, methanol), percentage of formic acid and water content in the mobile phase are systematically tested for all columns. It was shown that little amount of water added to the acetonitrile mobile phase may influence the retention behaviors between reversed phase and HILIC-like modes, and consequently the enantiomer elution order of the four stereoisomers. On the contrary, reversed phase is always the observed mode for the methanol water mobile phase. A suitable combination of all these parameters is presented for each second-generation anticoagulant rodenticide with a description of the enantioresolution, the enantiomer elution order and the retention times of the respective stereoisomers. A method is validated for all stereoisomers of each second-generation anticoagulant rodenticide with chicken liver and according to an official bioanalytical guideline. As an example, the enantiomer fraction is evaluated in the liver of a raptor species (rodent predator) exposed to bromadiolone and difenacoum. The results showed that only one enantiomer of trans-bromadiolone and one enantiomer of cis-difenacoum is present in hepatic residues, although all four stereoisomers are present in bromadiolone and difenacoum rodenticide baits.