Brush-type Chiral Stationary Phases Research Articles

Cross-linked polysiloxane-coated stable bond O-9-(2,6-diisopropylphenylcarbamoyl)quinine and quinidine chiral stationary phases as well as application in enantioselective cryo-HPLC.

In this work, brush-type chiral stationary phases (CSPs) with O-9-(2,6-diisopropylphenylcarbamoyl)-modified quinidine (DIPPCQD-brush/-SH) and O-9-(2,6-diisopropylphenylcarbamoyl)-modified quinine (DIPPCQN-brush/-SH) were prepared as benchmarks for comparison with new corresponding polymeric CSPs with more stable bonding chemistry. These polymeric CSPs were prepared by coating a thin poly(3-mercaptopropyl)-methylsiloxane film together with the chiral selector onto vinyl-modified silica. In a second step, immobilization of the quinine/quinidine derivatives as well as cross-linking of the polysiloxane film to the vinyl-silica is achieved by a double thiol-ene click reaction. The polymeric CSPs exhibited similar enantioselectivity as the corresponding brush phases, but showed lower chromatographic efficiencies. Chiral acidic substances were separated into enantiomers (e.g., N-protected amino acids, herbicides like dichlorprop) in accordance with an enantioselective anion-exchange process. Oxidation of residual thiol groups of the polymer DIPPCQN-CSP introduced sulfonic acid co-ligands on the silica surface, which resulted in greatly reduced retention times. Acting as immobilized counterions, they allowed to reduce the concentration of counterions in the mobile phase, which is favorable for liquid chromatography (LC)-electrospray ionization-mass spectrometry application. Ibuprofen showed a single peak under ambient column temperature. However, application of cryogenic cooling of the column enabled to achieve baseline separation at -20°C column temperature. It can be explained by an enthalpically dominated separation, which leads to an increase in separation factors when the temperature is reduced. While it is quite uncommon to work at subzero degree column temperature, this work illustrates the potential to exploit such temperature regime for optimization of LC enantiomer separations.

Evaluation of silica from different vendors as the solid support of anion-exchange chiral stationary phases by means of preferential sorption and liquid chromatography.

Chromatographic performance of a chiral stationary phase is significantly influenced by the employed solid support. Properties of the most commonly used support, silica particles, such as size and size distribution, and pore size are of utmost importance for both superficially porous particles and fully porous particles. In this work, we have focused on evaluation of fully porous particles from three different vendors as solid supports for a brush-type chiral stationary phase based on 9-O-tert-butylcarbamoyl quinidine. We have prepared corresponding stationary phases under identical experimental conditions and determined the parameters of the modified silica by physisorption measurements and scanning electron microscopy. Enantiorecognition properties of the chiral stationary phases have been studied using preferential sorption experiments. The same material was slurry-packed into chromatographic columns and the chromatographic properties have been evaluated in liquid chromatography. We show that preferential sorption can provide valuable information about the influence of the pore size and total pore volume on the interaction of analytes of different size with the chirally-modified silica surface. The data can be used to understand differences observed in chromatographic evaluation of the chiral stationary phases. The combination of preferential sorption and liquid chromatography separation can provide detailed information on new chiral stationary phases.

New Brush-Type Chiral Stationary Phases for Enantioseparation of Pharmaceutical Drugs.

The importance of chirality in drug development is unquestionable, with chiral liquid chromatography (LC) being the most adequate technique for its analysis. Among the various types of chiral stationary phases (CSPs) for LC, brush-type CSPs provide the base for interaction analysis of CSPs and enantiomers, which provide valuable results that can be applied to interaction studies of other CSP types. In order to analyze the influence of aromatic interactions in chiral recognition, we designed a set of ten new brush-type CSPs based on (S)-N-(1-aryl-propyl)-3,5-dinitrobenzamides which differ in the aromatic unit directly linked to the chiral center. Thirty diverse racemates, including several nonsteroidal anti-inflammatory drugs and 3-hydroxybenzodiazepine drugs, were used to evaluate the prepared CSPs. Chromatographic analysis showed that the three new CSPs separate enantiomers of a wide range of compounds and their chromatographic behavior is comparable to the most versatile brush-type CSP—Whelk-O1. The critical role of the nonbonding interactions in positioning of the analyte (naproxen) in the cleft of CSP-6, as well as the analysis of interactions that make enantioseparation possible, were elucidated using computational methods. Furthermore, the influence of acetic acid as a mobile phase additive, on this enantiorecognition process was corroborated by calculations.

Unusual van Deemter plots of optical isomers on a chiral brush-type liquid chromatography column

Unusual dynamic behavior of the enantiomers of 9-bromo-11b-(tert-butyl)-2,3,6,11b-tetrahydrooxazolo[3′,2′:1,5]pyrrolo[3,4-b]quinoline-5,11-dione (I) was observed on a Nautilus-R column packed with silica grafted with antibiotic ristocetin. It consisted in (i) antibatic behavior of the van Deemter plots of the enantiomers and (ii) high and strongly enantiomer dependent values of the A- and B-terms of the van Deemter equation. Although rare, such a pattern has been found earlier in chiral chromatography, with all reported cases limited to brush-type chiral stationary phases. Adsorption dynamics in this system was studied by means of the moment method and the peak parking technique; hydrodynamic properties of the column were explored by using unretained tracers. It was shown that the peculiar shape of the van Deemter curves for the enantiomers of I is conditioned by imperfect packing of the stationary phase, which result in high transcolumn eddy dispersion, and by slow adsorption/desorption kinetics. It was proven that the whole void volume of the column available to an eluent is not accessible to the studied analyte because it cannot penetrate the space between neighboring grafted ligands. Its mass transfer in pores is also affected by the fact that the stagnant layer of the binary water-acetonitrile mobile phase differs in composition from the bulk liquid due to preferential adsorption of water that influences the apparent molecular diffusivity of solutes. An effect of the structures of analyte and chiral selector on the adsorption kinetics is also briefly discussed.

Optimization of the surface modification process of cross-linked polythiol-coated chiral stationary phases synthesized by a two-step thiol-ene click reaction.

A new platform technology for the preparation of stable chiral stationary phases was successfully optimized. The chiral selector tert-butylcarbamoylquinine was firstly covalently connected to the polymer poly(3-mercaptopropyl)methylsiloxane by thiol-ene click reaction. Secondly, the quinine carbamate functionalized polysiloxane conjugate was coated onto the surface of vinyl modified silica particles and cross-linked via thiol-ene click reaction. The amount of polysiloxane, chiral selector, radical initiator, reaction solvent (chloroform and methanol), reaction time, and pore size of the supporting silica particles were varied and systematically optimized in terms of achievable plate numbers while maintaining simultaneously enantioselectivity. The optimization was based on elemental analysis data, chromatographic results, and H/u-curves (Van Deemter) of the resultant chiral stationary phases. The results suggest that better chromatographic efficiency (higher plate numbers) at equal enantioselectivity can be achieved with methanol (a poor solvent for the polysiloxane that is dispersed rather than dissolved) and a lower film thickness of quinine carbamate functionalized polysiloxane. In this study, chiral stationary phases based on 100Å silica slightly outperformed 200Å silica particles (each 5μm). The optimized two step material exhibited significantly reduced mass transfer resistance compared to the one step material and equal performance as a brush-type chiral stationary phase.

Synthesis and performance of chiral ferrocene modified silica gel for mixed-mode chromatography

A brush-type chiral stationary phase, N-ferrocenyl benzoyl-(1S, 2R)-1, 2-diphenyl ethanol-bonded on the silica gel (NFcBEs) for high performance liquid chromatography (HPLC), was prepared using γ-glycidoxypropyltrimethoxysilane as coupling reagent. The structure of this novel material was characterized by infrared spectroscopy, elemental analysis and thermogravimetric analysis. Mechanism involved in the chromatographic separation is the multi-interaction including hydrophobic, π-π, hydrogen-bonding, π-charge transfer, dipole-dipole and acid-base equilibrium interactions. Based on these interactions, successful separation could be achieved among polycyclic aromatic hydrocarbons, mono-substituted benzenes, aromatic amines, quinolines, nucleosides, phenols and 5-nitroimidazoles drugs in reversed phase liquid chromatography (RPLC). Good resolutions for substituted amine isomers were also obtained with NFcBEs. Racemates of amino acids and drug carvedilol mixtures were well separated on NFcBEs in the normal phase liquid chromatography (NPLC) mode. Such stationary phase with characteristics of multi-interaction mechanism and mixed-mode separation is potential for the analysis of complex samples. The retention behaviors of R- and S-carvedilol on NFcBEs column were investigated with the assistance of quantum chemistry calculation using the density functional theory (DFT) B3LYP method.

The synthesis and characterization of novel brush-type chiral stationary phase based on terpenoid selector for resolution of chiral drugs

In the light of the chiral resolution mechanism and structures of brush-type CSP, a new chiral selector 4′-carboxyl-1′-ursolic methyl ester-3β-yl-benzoate has been prepared. Then the terpenoid chiral selector was covalently linked to 3-aminopropyl silica gel. Its structure identification data are provided by 1H NMR, MS and elementary analysis. The enantiodiscriminating capability of the brush-type CSP was evaluated by static adsorption experiment with methyl mandelate, aniline derivative of mandelic acid, benzoin and ibuprofen. Experimental results demonstrated that the chiral selector has selectivity, and the enantiomers of methyl mandelate and ibuprofen could be separated on the CSP, which indicated that the novel brush-type CSP possess a bright prospects for chiral separation potentially.

Gone in seconds: praxis, performance, and peculiarities of ultrafast chiral liquid chromatography with superficially porous particles.

A variety of brush-type chiral stationary phases (CSPs) were developed using superficially porous particles (SPPs). Given their high efficiencies and relatively low back pressures, columns containing these particles were particularly advantageous for ultrafast "chiral" separations in the 4-40 s range. Further, they were used in all mobile phase modes and with high flow rates and pressures to separate over 60 pairs of enantiomers. When operating under these conditions, both instrumentation and column packing must be modified or optimized so as not to limit separation performance and quality. Further, frictional heating results in axial thermal gradients of up to 16 °C and radial temperature gradients up to 8 °C, which can produce interesting secondary effects in enantiomeric separations. It is shown that the kinetic behavior of various CSPs can differ from one another as much as they differ from the well-studied C18 reversed phase media. Three additional interesting aspects of this work are (a) the first kinetic evidence of two different chiral recognition mechanisms, (b) a demonstration of increased efficiencies at higher flow rates for specific separations, and

Stereoselective high-performance liquid chromatography and analytical method characterization of evacetrapib using a brush-type chiral stationary phase: A challenging isomeric separation requiring a unique eluent system

Using HPLC chiral separation screening, various columns representing the polysaccharide, macrocyclic antibiotic and brush classes were assessed in multiple chromatographic modes for the separation of evacetrapib, a potential cardiovascular drug, from its enantiomer, two diastereomers and several impurities. Screening data consistently pointed to the brush-type Whelk-O 1 chiral column as most promising for this task. A systematic separation optimization process is outlined using the (S,S) Whelk-O 1 chiral column, first for the resolution of the isomers, and later including six potential impurities. A relatively complex yet rugged separation system was eventually identified that effectively resolves all compounds within a reasonable analysis time, and should serve as an adequate tool for evacetrapib bulk drug enantiopurity measurement.

Bidentate urea-based chiral selectors for enantioselective high performance liquid chromatography: Synthesis and evaluation of “Crab-like” stationary phases

A rational approach for the design and preparation of two new “Crab-like” totally synthetic, brush-type chiral stationary phases is presented. Enantiopure diamines, namely 1,2-diaminocyclohexane and 1,2-diphenyl-1,2-ethylene-diamine were treated with 3-(triethoxysilyl)propyl isocyanate, to yield reactive ureido selectors that were eventually attached to unmodified silica particles through a stable, bidentate tether, through a facile two-step one-pot procedure. A full chemical characterization of the new materials has been obtained through solid-state NMR (both 29Si and 13C CPMAS) spectroscopy. Columns packed with the two Crab-like chiral stationary phases allow for different mechanisms of separation: normal phase liquid chromatography, reversed phase liquid chromatography and polar organic mode and show a high stability at basic pH values. In particular, the Crab-like column containing the 1,2-diphenyl-1,2-ethylene-diamine selector proved a promising candidate for the resolution of a wide range of racemates (including benzodiazepines, N-derivatized amino acids, and free carboxylic acids) both in normal phase and polar organic mode. An Hmin of 9.57 at a μsf of 0.80mm/s (corresponding to 0.8mL/min) was obtained through van Deemter analysis, based on toluene, for the Crab-like column with the 1,2-diphenyl-1,2-ethylene-diamine selector (250mm×4.6mm I.D.), with a calculated reduced height equivalent to a theoretical plate (h) of only 1.91. Finally, comparative studies were performed with a polymeric commercially available P-CAP-DP column in order to evaluate enantioselectivity and resolution of the Crab-like columns.

A practical method for the quantitative assessment of non-enantioselective versus enantioselective interactions encountered in liquid chromatography on brush-type chiral stationary phase

A convenient experimental method for the quantitative assessment of enantioselective versus non-enantioselective interactions in liquid chromatography on brush-type chiral stationary phases (CSPs) is described. This procedure involves the systematic evaluation of the retention characteristics of resolved enantiomers as chiral selectands, SAs, on a set of CSPs containing chiral selectors, SOs, attached to an achiral support S with well-defined but different surface SO loading levels. The emerging body of retention data can be dissected into non-enantioselective and enantioselective increments by equations accounting separately for the two co-existing equilibrium processes, namely the “unproductive” adsorption of the enantiomers on the achiral domains of the CSP and their “productive” enantioselective association with the surface-anchored chiral SOs. The general applicability of this approach was demonstrated using a set of CSPs loaded with different densities of a quinine carbamate anion exchange-type SO, and three acidic model SAs, i.e., N-Fmoc-phenylalanine, N-[(3,5-dipropoxybenzyloxy)carbonyl]leucine and 2-(2,4-dichlorophenoxy)propanoic acid, employing nonpolar, polar organic and reversed phase mobile phase conditions. In all tested mobile phase environments, the global retention characteristics of the studied SO was found to be dominated by enantioselective SA/SO binding rather than by non-enantioselective SA/support interactions, providing evidence for chiral recognition processes primarily capitalizing on strong electrostatic intermolecular binding forces. For the investigated compounds, the polar organic mode was identified as the most favorable mobile phase scenario, producing the highest apparent enantioseparation factors, αapp, in combination with the lowest degree of non-enantioselective adsorption. It is anticipated that the described approach toward the deconvolution of non-enantioselective versus enantioselective interactions will also be applicable to other types of brush-type CSPs. Most important, the quantitative information on the inherent thermodynamics of SO–SA interactions emerging from these studies will permit an unambiguous interpretation of the changes of the chiral recognition characteristics occurring as a consequence of mobile phase modifications. We are confident that this knowledge will be helpful in the design and evaluation of new non-invasive surface and immobilization chemistries, and will also provide valuable guidance for the optimization of operation conditions for large scale preparative enantiomer separations. Finally, it should be stressed that the advanced strategy for distinguishing enantioselective from non-enantioselective contributions has general applicability to all liquid phase-based enantioseparation techniques, well beyond the realm of chromatography, such as liquid–liquid extraction and batch-binding processes.

Mechanistic investigations of cinchona alkaloid-based zwitterionic chiral stationary phases

Novel zwitterionic cinchona alkaloid-based chiral selectors (SOs) were synthesized and immobilized on silica gel. The corresponding brush-type chiral stationary phases (CSPs) were characterized as zwitterionic ion-exchange-type materials and exhibited remarkable enantioselectivity for their zwitterionic target analytes, viz. underivatized amino acids and aminosulfonic acids. We rationally designed structural modifications on the strong cation exchange (SCX) subunit of the zwitterionic SO and investigated the influence on chiral recognition power for amphoteric solutes. SOs with chiral isopropyl- or cyclohexyl-moieties in vicinity to the SCX site showed broadest application range by baseline resolving 39 out of 53 test compounds, including α-, β-, and γ-amino acids with different substitution patterns. Furthermore, we introduced two pseudoenantiomeric zwitterionic CSPs which combined the unique features of providing comparable enantioselectivities but reversed enantiomer elution orders. By application of slightly acidic polar organic mobile phases as preferred elution mode, we found that certain amounts of aprotic acetonitrile in protic methanol substantially increased enantioselectivity and resolution of amino acids in a structure-dependent manner.

Investigation of brush‐type chiral stationary phases based on O,O′‐diaroyl tartardiamide and O,O′‐bis‐(arylcarbamoyl) tartardiamide

Four chiral organosilanes based on O,O'-dibenzoyl tartardiamide, O,O'-bis-(3,5-dimethylbenzoyl) tartardiamide, O,O'-bis-(phenylcarbamoyl) tartardiamide and O,O'-bis-[(3,5-dimethylphenyl)carbamoyl] tartardiamide were synthesized and immobilized on silica to afford corresponding brush-type chiral stationary phases (CSPs) with well-defined structures. Using 54 compounds containing a wide variety of structures as analytes, the enantioselectivities of the four CSPs were evaluated under normal-phase modes. 3,5-Dimethyl substituent in the aryl group was found to significantly affect the enantioselectivity of CSPs containing aryl ester moieties. Aryl carbamate moieties in CSPs were observed more beneficial for enantioseparation than aryl ester moieties. The additional hydrogen-bond donors (NH) present in the carbamate groups contributed greatly to the enantioselectivity of CSPs, which is contrary to the results that have been found in network-polymeric CSPs.

Investigation of Chiral Self-Recognition of π-Acidic 3,5-Dinitrobenzoylleucine Derivatives

School of Chemical Sciences, University of Illinois, Urbana, IL 61801, USAReceived September 1, 2011, Accepted October 9, 2011Key Words : Enantiomer separation, Chiral self-recognition, Chiral stationary phasePioneering work in the design of brush-type chiralstationary phases (CSPs) using chiral recognition rationaleshas led to the development of a number of Pirkle-typecolumns.

New silica monolith bonded chiral (R)‐γ butyrolactone for enantioselective micro high‐performance liquid chromatography

A single low-molecular mass chiral selector namely (R)-acryloyloxy-β-β-dimethyl-γ-butyrolactone has been bonded to a modified silica-based monolith to form a new brush-type chiral stationary phase for micro-high performance liquid chromatography (HPLC) separation.

Separation of the Enantiomers of β-Blockers Using Brush Type Chiral Stationary Phase Derived from Conformationally Rigid α-Amino β-Lactam

A brush type chiral stationary phase (CSP 2) derived from <TEX>${\alpha}$</TEX>-amino <TEX>${\beta}$</TEX>-lactam was prepared for the separation of the enantiomers of <TEX>${\beta}$</TEX>-blockers. Compared to the CSP derived from <TEX>${\alpha}$</TEX>-amino phosphonate (CSP 1), in general, the conformationally rigid CSP 2 showed greater scope and much enhanced enantioselectivity for the resolution of <TEX>${\beta}$</TEX>-blockers. The effect of various salt additives on enantioseparation of <TEX>${\beta}$</TEX>-blockers in the mobile phase was investigated. The unusual effect of temperature on the chromatographic behaviors was observed on CSP 2. It also afforded appreciable increases in enantioselectivity without significantly affecting resolution, as the column temperature was reduced.

Preparation and enantioseparation of a mixed selector chiral stationary phase derived from benzoylated tartaric acid and 1,2‐diphenylethylenediamine

L-Dibenzoyl tartaric acid was mono-esterified with benzyl alcohol, and then chlorinated with SOCl(2) to give (2S,3S)-1-(benzyloxy)-4-chloro-1,4-dioxobutane-2,3-diyl dibenzoate (Selector 1). (1R,2R)-1,2-Diphenylethylenediamine was mono-functionalized with phenyl isocyanate and phenylene diisocyanate in sequence to give (1R,2R)-1,2-diphenyl-2-(3-phenylureido)ethyl 4- isocyanatophenylurea (Selector 2). Two brush-type chiral stationary phases (CSPs) of single selector were prepared by separately immobilizing selectors 1 and 2 on aminated silica gel. Selectors 1 and 2 were simultaneously immobilized on aminated silica gel to give a mixed selector CSP. The enantioseparation ability of these CSPs was studied. The CSP of selector 1 has strongest separation ability, while the enantioseparation ability of the mixed selector CSP is relatively lower.

Transition from enantioselective high performance to ultra-high performance liquid chromatography: A case study of a brush-type chiral stationary phase based on sub-5-micron to sub-2-micron silica particles

Three brush-type chiral stationary phases (CSPs) differing in the particle size of the starting silica particles have been prepared by covalent grafting of the π-acidic bis-(3,5-dinitrobenzoyl)-derivative of trans-1,2-diaminocyclohexane (DACH-DNB). Starting silica particles of 4.3, 2.6 and 1.9 micron were used to generate the final CSPs using an improved, highly reproducible synthetic methodology, that allowed to assemble and surface-graft the whole chiral selector in only two steps. The different CSPs have been packed in columns of various length and diameters, and fully characterized in terms of flow permeability, kinetic performances and enantioselectivity using a set of test solutes. Very high speed and high resolution applications together with stereodynamic HPLC examples are demonstrated on the columns with reduced particle diameters, on which separations of several enantiomeric pairs are routinely obtained with analysis times in the 15–40 s range.

Chiral Self-Selectivity in Two Model Dinitrobenzoyl-Derivatized Brush-Type Chiral Stationary Phases

A study of chiral self-discrimination in two model chiral stationary phases, N-(3,5-dinitrobenzoyl-phenylglycine and N-(3,5-dinitrobenzoyl)leucine, is described. “Brush-type” chiral interfaces are obtained by attaching these compounds onto oxidized Si(111) samples and atomic force microscopy tips by using two sample preparation procedures: a direct and a two-step surface deposition of the chiral stationary phase onto the substrate. By using AFM, the morphologies of the resulting chiral interfaces are obtained. XPS and FTIR provide information regarding the relative distribution of the compounds on the surface. We analyze the N 1s peaks using XPS to ensure that all the NH2 groups on the surface are capped. Chemical force spectrometric measurements of the chiral self-selectivity in 2-propanol demonstrate that chiral discrimination is present in both systems, but larger forces are observed for N-(3,5-dinitrobenzoyl)phenylglycine, consistent with a previous molecular dynamics study that demonstrated much weaker solvent interactions with this species.

In‐column preparation of a brush‐type chiral stationary phase using click chemistry and a silica monolith

Brush-type chiral stationary phases (CSP) have been prepared both from a silica monolith and, separately, from 10 mum porous silica beads via a process of in-column modification including attachment of the chiral selector via copper-catalyzed azide-alkyne cycloaddition. Azide functionalities were first introduced on the pore surface of each type of support by reaction with 3-(azidopropyl)trimethoxysilane, followed by immobilization of a proline-derived chiral selector containing an alkyne moiety. This functionalization reaction was carried out in dimethylformamide (DMF) in the presence of catalytic amounts of copper (I) iodide. The separation performance of these triazole linked stationary phases was demonstrated in enantioseparations of four model analytes, which afforded separation factors as high as 11.4.