Chiral separation of helical chromenes with chloromethyl phenylcarbamate polysaccharide‐based stationary phases

Two chloromethyl phenylcarbamate‐based chiral stationary phases, one containing an amylose‐type chiral selector (Lux Amylose 2, from Phenomenex) and the other a cellulose‐type one (Lux Cellulose‐4, from Phenomenex), were successfully used for the chiral resolution of three helical chromenes featuring a helicene‐like structure. The compound bearing a phenyl substituent on the helicene‐like structure was enantioresolved at 25°C with Lux Cellulose‐4 and a n‐hexane/1‐propanol 99:1 v/v eluent. With a n‐hexane/2‐propanol 99.8:0.2 v/v mobile phase, the same column (operated at 35°C) provided the separation of the four isomers of the compound having a hexyl residue on the helicene‐like motif and an additional asymmetric carbon. Lux Amylose‐2 was necessary for the enantioseparation of the compound having the sole hexyl residue on the helical scaffold. For the last compound a n‐hexane/2‐propanol 99.8:0.2 v/v eluent was used, and the column temperature was fixed at 5°C. The enantiomer elution order was appraised by using electronic circular dichroism and theoretical calculations. Notably, different thermodynamics of retention and enantioseparation were observed for molecules with pronounced structural similarity, that is, the enantiomer pairs of the compound containing the additional asymmetric carbon atom. Indeed, both entropically and enthalpically controlled adsorption and separation processes were observed.     

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.     

Enantioseparation of selected chiral sulfoxides in high‐performance liquid chromatography with polysaccharide‐based chiral selectors in polar organic mobile phases with emphasis on enantiomer elution order

The separation of the enantiomers of 17 chiral sulfoxides was studied on polysaccharide‐based chiral columns in polar organic mobile phases. Enantiomer elution order (EEO) was the primary objective in this study. Two of the six chiral columns, especially those based on amylose tris(3,5‐dimethylphenylcarbamate) and cellulose tris(4‐chloro‐3‐methylphenylcarbamate) (Lux Cellulose‐4) proved to be most successful in the separation of the enantiomers of the studied sulfoxides. Interesting examples of EEO reversal were observed depending on the chiral selector or the composition of the mobile phase. For instance, the R‐(+) enantiomer of lansoprazole eluted before the S‐(?) enantiomer on Lux Cellulose‐1 in both methanol or ethanol as the mobile phase, while the elution order was opposite in the same eluents on amylose tris(3,5‐dimethylphenylcarbamate) with the S‐(?) enantiomer eluting before the R‐(+) enantiomer. The R‐(+) enantiomer of omeprazole eluted first on Lux Amylose‐2 in methanol but it was second when acetonitrile was used as the mobile phase with the same chiral selector. Several other examples of reversal in EEO were observed in this study. An interesting example of the separation of four stereoisomers of phenaminophos sulfoxide containing chiral sulfur and phosphor atoms is also reported here.     

Combined use of l‐alanine tert butyl ester lactate and trimethyl‐β‐cyclodextrin for the enantiomeric separations of 2‐arylpropionic acids nonsteroidal anti‐inflammatory drugs

In this study, a new CE method, employing a binary system of trimethyl‐β‐CD (TM‐β‐CD) and a chiral amino acid ester‐based ionic liquid (AAIL), was developed for the chiral separation of seven 2‐arylpropionic acid nonsteroidal anti‐inflammatory drugs (NSAIDs). In particular, the enantioseparation of ibuprofen, ketoprofen, carprofen, indoprofen, flurbiprofen, naproxen, and fenoprofen was improved significantly by supporting the BGE with the chiral AAIL l ‐alanine tert butyl ester lactate (l ‐AlaC₄Lac). Parameters, such as concentrations of TM‐β‐CD and l ‐AlaC₄Lac, and buffer pH, were systematically examined in order to optimize the chiral separation of each NSAID. It was observed that the addition of the AAIL into the BGE improved both resolution and efficiency significantly. After optimization of separation conditions, baseline separation (R_s>1.5) of five of the analytes was achieved in less than 11 min, while the resolution of ibuprofen and flurbiprofen was approximately 1.2. The optimized enantioseparation conditions for all analytes involve a BGE of 5 mM sodium acetate/acetic acid (pH 5.0), an applied voltage of 30 kV, and a temperature of 20°C. In addition, the results obtained by computing the %‐RSD values of the EOF and the two enantiomer peaks, demonstrated excellent run‐to‐run, batch‐to‐batch, and day‐to‐day reproducibilities.     

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.     

A novel C2 symmetric chiral stationary phase with N‐[(4‐Methylphenyl)sulfonyl]‐l‐leucine as chiral side chains

In this study, a series of chiral stationary phases based on N‐[(4‐methylphenyl)sulfonyl]‐l ‐leucine amide, whose enantiorecognition property has never been studied, were synthesized. Their enantioseparation abilities were chromatographically evaluated by 67 enantiomers. The chiral stationary phase derived from N‐[(4‐methylphenyl)sulfonyl]‐l ‐leucine showed much better enantioselectivities than that based on N‐(4‐methylbenzoyl)‐l ‐leucine amide. The construction of C₂ symmetric chiral structure greatly improved the enantiorecognition performance of the stationary phase. The C₂ symmetric chiral stationary phase exhibited superior enantioresolutions to other chiral stationary phases for most of the chiral analytes, especially for the chiral analytes with C₂ symmetric structures. By comparing the enantioseparations of the enantiomers with similar structures, the importance of hydrogen bond interaction, π–π interaction, and steric hindrance on enantiorecognition was elucidated. The enantiorecognition mechanism of trans‐N,N′‐(1,2‐diphenyl‐1,2‐ethanediyl)bis‐acetamide, which had an excellent separation factor on the C₂ symmetric chiral stationary phase, was investigated by ¹H‐NMR spectroscopy and 2D ¹H‐¹H nuclear overhauser enhancement spectroscopy.     

Synthesis and application of a chiral ionic liquid functionalized β‐cyclodextrin as a chiral selector in capillary electrophoresis

A novel chiral ionic liquid functionalized β‐cyclodextrin, 6‐O‐2‐hydroxpropyltrimethylammonium‐β‐cyclodextrin tetrafluoroborate ([HPTMA‐β‐CD][BF₄]), was synthesized and used as a chiral selector in capillary electrophoresis. [HPTMA‐β‐CD][BF₄] not only increased the solubility in aqueous buffer in comparison with the parent compound, but also provided a stable reversal electroosmotic flow, and the enantioseparation of eight chiral drugs was examined in phosphate buffer containing [HPTMA‐β‐CD][BF₄] as the chiral selector. The effects of the [HPTMA‐β‐CD][BF₄] concentration and the background electrolyte pH were studied. Moreover, the chiral separation abilities of β‐CD and [HPTMA‐β‐CD][BF₄] were compared and possible mechanisms for the chiral recognition of [HPTMA‐β‐CD][BF₄] are discussed. Copyright © 2013 John Wiley & Sons, Ltd.     

Reversed‐phase liquid chromatographic determination of enantiomers of atenolol in rat plasma using derivatization with Marfey's reagent

An HPLC method was established for enantioseparation of (R,S)‐atenolol (ATE) and determination of enantiomers in rat plasma. Marfey's reagent (1‐fluoro‐2,4‐dinitrophenyl‐5‐L‐alanine amide, FDNP‐L‐Ala‐NH₂, MR) was used as chiral derivatizing reagent with detection of diastereomers at 340 nm. It was shown that the R‐isomer eluted before the S‐isomer. The method was validated for linearity, repeatability, limits of detection and limit of quantification (LOQ). Recovery of ATE at LOQ was 92.8% for (R)‐ATE and 92.6% for (S)‐ATE. Copyright © 2009 John Wiley & Sons, Ltd.     

Efficient preparative separation of 6‐(4‐aminophenyl)‐5‐methyl‐4, 5‐dihydro‐3(2H)‐pyridazinone enantiomers on polysaccharide‐based stationary phases in polar organic solvent chromatography and supercritical fluid chromatography

6‐(4‐Aminophenyl)‐5‐methyl‐4,5‐dihydro‐3(2H)‐pyridazinone is a key synthetic intermediate for cardiotonic agent levosimendan. Very few studies address the use of chiral stationary phases in chromatography for the enantioseparation of this intermediate. This study presents two efficient preparative methods for the isolation of (R)(?)‐6‐(4‐aminophenyl)‐5‐methyl‐4,5‐dihydro‐3(2H)‐pyridazinone in polar organic solvent chromatography and supercritical fluid chromatography using polysaccharide‐based chiral stationary phases and volatile organic mobile phases without additives in isocratic mode. Under optimum conditions, Chiralcel OJ column showed the best performance (α = 1.71, Rs = 5.47) in polar organic solvent chromatography, while Chiralpak AS column exhibited remarkable separations (α = 1.81 and Rs = 6.51) in supercritical fluid chromatography with an opposite enantiomer elution order. Considering the sample solubility, runtime and solvent cost, the preparations were carried out on Chiralcel OJ column and Chiralpak AS column (250 × 20 mm i.d.; 10 µm) in polar organic mode and supercritical fluid chromatography mode with methanol and CO₂/methanol as mobile phases, respectively. By utilizing the advantages of chromatographic techniques and polysaccharide‐based chiral stationary phases, this work provides two methods for the fast and economic preparation of (R)(?)‐6‐(4‐aminophenyl)‐5‐methyl‐4,5‐dihydro‐3(2H)‐pyridazinone, which are suitable for the pharmaceutical industry.     

Conformational analysis of some 1R,4S‐2‐arylidene‐p‐menthan‐3‐ones by 1H NMR spectroscopy and molecular simulation

Based on ¹H NMR spectral analysis combined with molecular simulation, conformational states of the cyclohexanone ring were studied for some 1R,4S‐2‐(4‐X‐benzylidene)‐p‐menthan‐3‐ones (X = COOCH₃ or C₆H₅) in CDCl₃ and C₆D₆. The co‐existence of chair conformers with an axial orientation of both alkyl substituents and twist‐boat forms was established for the compounds studied at room temperature (22–23° C). The substituent X does not influence appreciably the ratio of these conformers, but the fraction of twist‐boat forms increases noticeably in benzene solutions as compared with CDCl₃ solutions. Rotameric states of the isopropyl fragment were also characterised for the compounds studied. Distinctions in conformational states for the 1R,4S‐2‐arylidene‐p‐menthan‐3‐ones and (?)‐menthone were revealed and are discussed. Copyright © 2002 John Wiley & Sons, Ltd.     

Enantioselective determination of carboxyl acid amide fungicide mandipropamid in vegetables and fruits by chiral LC coupled with MS/MS

An efficient enantioselective method for the determination of mandipropamid in vegetables and fruits was presented by LC coupled with MS/MS. The mandipropamid residues in samples (potato, pepper, grape, and watermelon) were extracted with acetonitrile containing 1% acetic acid. An aliquot was cleaned up with primary and secondary amine and C₁₈ sorbent. Complete enantioseparation of mandipropamid enantiomers in <4 min was obtained on a Lux Cellulose‐2 column at 25°C using methanol with 0.1% formic acid/0.1% aqueous formic acid solution (85:15, v/v) as mobile phase. Good linearity was obtained over the concentration range of 0.5–250 μg/L for each enantiomer in the standard solution and sample matrix calibration curves. Quantification was achieved using matrix‐matched standard calibration curves. The interday mean recoveries, intraday repeatability, and inter‐day reproducibility varied from 76.4 to 97.1%, 3.4 to 9.4%, and 3.5 to 11.4%, respectively. The limits of quantification for mandipropamid enantiomers in vegetables and fruits were both 1 μg/kg. Moreover, the absolute configuration of mandipropamid enantiomers was determined by the combination of experimental and predicted electronic circular dichroism spectra, and the first eluted enantiomer was confirmed as (R)‐mandipropamid on five chiral columns.     

(1R,2R)‐1,2‐Di­phenyl‐1,2‐bis(8‐quinoline­sulfonyl­amino)ethyl­enedi­amine–acetone (1/1)

The crystal structure of the new chiral complex (1R,2R)‐1,2‐di­phenyl‐1,2‐bis(8‐quinoline­sulfonyl­amino)‐ ethyl­enedi­amine–acetone (1/1), C₃₂H₂₆N₄O₄S₂^.C₃H₆O, is reported. The conformation of the C₃₂H₂₆N₄O₄S₂ (BQSDA) mol­ecule is determined by a bifurcated N—H?N hydrogen‐bond system. The acetone of solvation is linked to the BQSDA mol­ecule by an N—H?O hydrogen bond.     

Preparative enantioseparation of loxoprofen precursor by recycling countercurrent chromatography with hydroxypropyl‐β‐cyclodextrin as a chiral selector

Recycling countercurrent chromatography was successfully applied to the resolution of 2‐(4‐bromomethylphenyl)propionic acid, a key synthetic intermediate for synthesis of nonsteroidal anti‐inflammatory drug loxoprofen, using hydroxypropyl‐β‐cyclodextrin as chiral selector. The two‐phase solvent system composed of n‐hexane/n‐butyl acetate/0.1 mol/L citrate buffer solution with pH 2.4 (8:2:10, v/v/v) was selected. Influence factors for the enantioseparation were optimized, including type of substituted β‐cyclodextrin, concentration of hydroxypropyl‐β‐cyclodextrin, separation temperature, and pH of aqueous phase. Under optimized separation conditions, 50 mg of 2‐(4‐bromomethylphenyl)propionic acid was enantioseparated using preparative recycling countercurrent chromatography. Technical details for recycling elution mode were discussed. The purities of both the S and R enantiomers were over 99.0% as determined by high‐performance liquid chromatography. The enantiomeric excess of the S and R enantiomers reached 98.0%. The recovery of the enantiomers from eluted fractions was 40.8–65.6%, yielding 16.4 mg of the S enantiomer and 10.2 mg of the R enantiomer. At the same time, we attempted to enantioseparate the anti‐inflammatory drug loxoprofen by countercurrent chromatography and high‐performance liquid chromatography using a chiral mobile phase additive. However, no successful enantioseparation was achieved so far.     

HPLC enantioseparation of β2‐homoamino acids using crown ether‐based chiral stationary phase

RP high‐performance liquid chromatographic methods were developed for the enantioseparation of eleven unusual β²‐homoamino acids. The underivatized analytes were separated on a chiral stationary phase containing (+)‐(18‐crown‐6)‐2,3,11,12‐tetracarboxylic acid as chiral selector. The effects of organic (alcoholic) and acidic modifiers, the mobile phase composition and temperature on the separation were investigated. The structures of the substituents in the α‐position of the analytes substantially influenced the retention and resolution. The elution sequence was determined in some cases: the S enantiomers eluted before the R enantiomers.     

High‐performance liquid chromatographic enantioseparation of (RS)‐bupropion using isothiocyanate‐based chiral derivatizing reagents

A high‐performance liquid chromatographic (HPLC) method for enantioseparation of bupropion was developed using two isothiocyanate‐based chiral derivatizing reagents, (S)‐1‐(1‐naphthyl) ethyl isothiocyanate, (S)‐NEIT, and (R)‐α‐methyl benzyl isothiocyanate, (R)‐MBIT. The diastereomers synthesized with (S)‐NEIT were enantioseparated by reversed‐phase HPLC using gradient elution with mobile phase containing water and acetonitrile, whereas diastereomers synthesized with (R)‐MBIT were enantioseparated using triethyl amine phosphate buffer and methanol. Derivatization conditions were optimized and the method was validated for accuracy, precision and limit of detection. The limit of detection was found to be 0.040–0.043 µg/mL for each of the diastereomers prepared with (S)‐NEIT. Copyright © 2013 John Wiley & Sons, Ltd.     

Surface‐imprinted microspheres prepared by a template‐oriented method for the chiral separation of amlodipine

The surface imprinting technique has been developed to overcome the mass‐transfer difficulty, but the utilization ratio of template molecules in the imprinting procedure still remains a challengeable task to be improved. In this work, specifically designed surface‐imprinted microspheres were prepared by a template‐oriented method for enantioseparation of amlodipine besylate. Submicron mesoporous silica microspheres were surface‐modified with double bonds, followed by polymerizing methacrylic acid to generate carboxyl modified mesoporous silica microspheres (PMAA@SiO₂). Afterwards, PMAA@SiO₂ was densely adsorbed with (S )‐amlodipine molecules to immobilize template molecules through multiple hydrogen bonding interactions. Then surface molecular imprinting was carried out by cross‐linking the carboxyl group of PMAA@SiO₂ with ethylene glycol diglycidyl ether. The surface‐imprinted microspheres showed fast binding kinetics of only 20 min for equilibrium adsorption, and the saturation adsorption capacity reached 137 mg/g. The imprinted materials displayed appreciable chiral separation ability when used as column chromatography for enantioseparation of amlodipine from amlodipine besylate, and the enantiomeric excess of (S )‐amlodipine reached 13.8% with only 2.3 cm column length by no extra chiral additives. Besides, the imprinted materials exhibited excellent reusability, and this allows the potential application for amplification production of amlodipine enantiomer.     

Binding modes identification through molecular dynamic simulations: A case study with carnosine enantiomers and the Teicoplanin A2‐2‐based chiral stationary phase

In the present study, an in silico methodology able to define the binding modes adopted by carnosine enantiomers in the setting of the chiral recognition process is described. The inter‐ and intramolecular forces involved in the enantioseparation process with the Teicoplanin A2‐2 chiral selector and carnosine as model compound are successfully identified. This approach fully rationalizes, at a molecular level, the (S) < (R) enantiomeric elution order obtained under reversed‐phase conditions. Consistent explanations were achieved by managing molecular dynamics results with advanced techniques of data analysis. As a result, the time‐dependent identification of all the interactions simultaneously occurring in the chiral selector‐enantiomeric analyte binding process was obtained. Accordingly, it was found that only (R)‐carnosine is able to engage a stabilizing charge–charge interaction through its ionized imidazole ring with the carboxylate counter‐part on the chiral selector. Instead, (S)‐carnosine establishes intramolecular contacts between its ionized functional groups, that limit its conformational freedom and impair the association with the chiral selector unit.     

In‐situ photopolymerized polyhedral oligomeric silsesquioxane‐derived monolithic capillary columns with quinidine functionality for enantioseparation by nano‐liquid chromatography

The successful fabrication of monolithic capillary columns for enantiomer separations was achieved within vinylized fused silica capillaries via fast “one‐pot” photo‐initiated free radical polymerization reaction. A mixture consisting of polyhedral oligomeric silsesquioxane, O‐[2‐(methacryloyloxy)ethylcarbamoyl]‐10,11‐dihydroquinidine was copolymerized in the presence of n‐butanol, ethylene glycol and photo‐initiator 2,2‐dimethoxy‐2‐phenylacetophenone. The morphology of the resultant polymeric hybrid inorganic‐organic material and its permeability as well as porosity can be controlled by adjusting the composition of the monomers and binary porogenic solvent. The chromatographic characteristics of the columns have been investigated. Separation factors of N‐acetyl‐phenylalanine (Ac‐Phe) and dichlorprop dropped with decrease of chiral functional monomer. Permeability was better when the macroporogen ethyleneglycol was present at higher concentrations during the polymerization. In general, the chiral compounds were well separated (dichlorprop: α = 1.53, R_s up to 4.14; Ac‐Phe: α = 1.36, R_s up to 2.69) by nano‐HPLC with an optimized enantioselective monolithic capillary column which can be prepared within a few minutes.     

Study of molecular structures for arylidene derivatives of 3R‐methylcyclohexanone by 1H NMR and molecular simulation

Conformational states of the cyclohexanone ring were established for 3R‐methyl‐6‐(4‐phenylbenzylidene)cyclohexanone and several 2,6‐bis(4‐X‐benzylidene)‐3R‐methylcyclohexanones (X = Br, OCOCH₃ and C₆H₅) by ¹H NMR spectroscopy combined with molecular simulation using the semi‐empirical methods AM1 and PM3. The first compound studied contains only one arylidene group, and exists predominantly in a chair conformation of the cyclohexanone ring with an equatorial orientation of the methyl substituent in C₆D₆ and CDCl₃ solutions at room temperature (22–23 °C). In contrast, the 2,6‐bis(arylidene) derivatives of 3R‐methylcyclohexanone preferentially adopt conformations with an axially oriented methyl group. The extent of twisting of enone fragments was also characterized for the compounds studied based on simulation results and comparison of chemical shifts for the arylidene protons of appropriate model compounds. Copyright © 2003 John Wiley & Sons, Ltd.     

Chiral capillary electrophoresis with cationic pyrrolidinium‐β‐cyclodextrin derivatives as chiral selectors

New single‐isomer, cationic β‐cyclodextrins, including mono‐6‐deoxy‐6‐pyrrolidine‐β‐cyclodextrin chloride (pyCDCl), mono‐6‐deoxy‐6‐(N‐methyl‐pyrrolidine)‐β‐cyclodextrin chloride (N‐CH₃‐pyCDCl), mono‐6‐deoxy‐6‐(N‐(2‐hydroxyethyl)‐pyrrolidine)‐β‐cyclodextrin chloride (N‐EtOH‐pyCDCl), mono‐6‐deoxy‐6‐(2‐hydroxymethyl‐pyrrolidine)‐β‐cyclodextrin chloride (2‐MeOH‐pyCDCl) were synthesized and used as chiral selectors in capillary electrophoresis for the enantioseparation of carboxylic and hydroxycarboxylic acids and dansyl amino acids. The unsubstituted pyCDCl exhibited the greatest resolving ability. Most analytes were resolved over a wide range of pH from 6.0 to 9.0 with this chiral selector. In general, increasing pH led to a decrease in resolution. The effective mobilities of all the analytes were found to decrease with increasing CD concentration. The optimal concentration for most carboxylic acids and dansyl amino acid was in the range 5–7.5 mM and >15 mM for hydroxycarboxylic acids. ¹H NMR experiments provided direct evidence of inclusion in the CD cavity.