Nonaqueous Capillary Electrophoretic Enantioseparation of Water Insoluble Tröger's Base Derivatives Using β‐Cyclodextrin as Chiral Selector

Racemic mixtures of six Tröger's base derivatives were separated by chiral nonaqueous capillary electrophoresis. The separation protocol was optimized first for suitable solvents. Then the applicability of various salts dissolved in organic solvents and their mixtures was evaluated. As chiral selectors β‐cyclodextrin and heptakis(2,6‐di‐O‐methyl)‐β‐cyclodextrin at various concentrations were used. The best enantioselectivity for the studied analytes was obtained utilizing formamide as organic nonaqueous solvent containing a mixture of sodium citrate and tris(hydroxymethyl)aminomethane acetate as electrolytes, and β‐cyclodextrin as chiral additive. The experimental results demonstrated the feasibility of nonaqueous capillary electrophoresis for enantioseparation of Tröger's base derivatives. This technique represents a suitable alternative to more commonly used capillary electrophoresis in aqueous environment. Chirality 25:810–813, 2013. © 2013 Wiley Periodicals, Inc.     

GC Separation of Enantiomers of Alkyl Esters of 2‐Bromo Substituted Carboxylic Acids Enantiomers on 6‐TBDMS‐2,3‐di‐alkyl‐ β‐ and γ‐Cyclodextrin Stationary Phases

The gas chromatographic separation of enantiomers of 2‐Br carboxylic acid derivatives was studied on four different 6‐TBDMS‐2,3‐di‐O‐alkyl‐ β‐ and ‐γ‐CD stationary phases. The differences in thermodynamic data {ΔH and –ΔS} for the 15 structurally related racemates were evaluated. The influence of structure differences in the alkyl substituents covalently attached to the stereogenic carbon atom, as well as in the ester group of the homologous analytes, and the selectivity of modified β‐ and γ‐ cyclodextrin derivatives was studied in detail. The cyclodextrin cavity size, as well as elongation of alkyl substituents in positions 2 and 3 of 6‐TBDMS‐β‐CD, also affected their selectivity. The quality of enantiomeric separations is influenced mainly by alkyl chains of the ester group of the molecule and this appears to be independent of the CD stationary phase used. In some cases the separations occur as the result of external adsorption rather than inclusion complexations with the chiral selector. It was found that the temperature dependencies of the selectivity factor were nonlinear. Chirality 26:279–285, 2014. © 2014 Wiley Periodicals, Inc.     

Combined Use of Ionic Liquid and Hydroxypropyl‐β‐Cyclodextrin for the Enantioseparation of Ten Drugs by Capillary Electrophoresis

In the present study, hydroxypropyl‐β‐cyclodextrin and an ionic liquid (1‐ethyl‐3‐methylimidazolium‐l ‐lactate) were used as additives in capillary electrophoresis for the enantioseparation of 10 analytes, including ofloxacin, propranolol hydrochloride, dioxopromethazine hydrochloride, isoprenaline hydrochloride, chlorpheniramine maleate, liarozole, tropicamide, amlodipine benzenesulfonate, brompheniramine maleate, and homatropine methylbromide. The effects of ionic liquid concentrations, salt effect, cations, and anions of ionic liquids on enantioseparation were investigated and the results proved that there was a synergistic effect between hydroxypropyl‐β‐cyclodextrin and the ionic liquid, and the cationic part of the ionic liquid played an important role in the increased resolution. With the developed dual system, all the enantiomers of 10 analytes were well separated in resolutions of 5.35, 1.76, 1.85, 2.48, 2.88, 1.43, 5.45, 4.35, 2.76, and 2.98, respectively. In addition, the proposed method was applied to the determination of the enantiomeric purity of S‐ofloxacin after validation of the method in terms of selectivity, repeatability, linearity range, accuracy, precision, limit of detection (LOD), and limit of quality (LOQ). Chirality 25:409–414, 2013. © 2013 Wiley Periodicals, Inc.     

Separation of Selected Imidazole Enantiomers Using Dual Cyclodextrin System in Micellar Electrokinetic Chromatography

Cyclodextrin‐modified micellar electrokinetic chromatography (CD‐MEKC) method was developed for simultaneous enantioseparation of three imidazole drugs namely tioconazole, isoconazole and fenticonazole. Three easily available and inexpensive cyclodextrins namely 2‐hydroxypropyl‐β‐cyclodextrin (HP‐β‐CD), 2‐hydroxypropyl‐γ‐cyclodextrin (HP‐γ‐CD) and heptakis(2,6‐di‐O‐methyl)‐β‐cyclodextrin (DM‐β‐CD) were evaluated to discriminate the six stereoisomers of the drugs. However, none of the three CDs gave a complete enantioseparation of the drugs. Effective enantioseparation of tioconazole, isoconazole and fenticonazole was achieved using a combination of 35 mM HP‐γ‐CD and 10 mM DM‐β‐CD as chiral selectors. The best separation using both HP‐γ‐CD and DM‐β‐CD (35 mM:10 mM) as chiral selectors were accomplished in background electrolyte (BGE) containing 35 mM phosphate buffer (pH 7.0), 50 mM sodium dodecyl sulfate (SDS) and 15% (v/v) acetonitrile at 27 kV and 30 °C with all peaks resolved in less than 15 min with resolutions, R_s 1.90‐27.22 and peak efficiencies, N > 180 000. The developed method was linear over the concentration range of 25–200 mg l^‐1 (r² > 0.998) and the detection limits (S/N = 3) of the three imidazole drugs were found to be 2.7‐7.7 mg l^‐1. The CD‐MEKC method was successfully applied to the determination of the three imidazole drugs in spiked human urine sample and commercial cream formulation of tioconazole and isoconazole with good recovery (93.6‐106.2%) and good RSDs ranging from 2.30‐6.8%. Chirality 25:328–335, 2013. © 2013 Wiley Periodicals, Inc.     

Comparative Enantioseparation of Ketoprofen with Trimethylated α‐, β‐, and γ‐Cyclodextrins in Capillary Electrophoresis and Study of Related Selector–Selectand Interactions Using Nuclear Magnetic Resonance Spectroscopy

The enantiomers of ketoprofen were separated by capillary electrophoresis using the (2,3,6‐tri‐O‐methyl)‐derivatives of α‐, β‐, and γ‐cyclodextrin (CyD) as chiral selectors. The affinity pattern of the ketoprofen enantiomers toward these CyDs changed depending on their cavity size. Thus, with hexakis (2,3,6‐tri‐O‐methyl)‐α‐CyD and heptakis (2,3,6‐tri‐O‐methyl)‐β‐CyD, the R enantiomer of the drug migrated first, whereas the enantiomer migration order was reversed in the presence of octakis(2,3,6‐tri‐O‐methyl)‐γ‐CyD. The change in the migration order was rationalized on the basis of changes in the structure of the complexes between the ketoprofen enantiomers and the chiral selectors as derived from nuclear magnetic resonance spectroscopy experiments. Chirality, 25:79–88, 2013.© 2012 Wiley Periodicals, Inc.     

Separation of Enantiomers by Inclusion Gas Chromatography: On the Influence of Water in the Molecular Complexation of Methyl 2‐Chloropropanoate Enantiomers and the Modified γ‐Cyclodextrin Lipodex‐E

A profound influence of water has previously been detected in the complexation of the enantiomers of methyl 2‐chloropropanoate (MCP) and the chiral selector octakis(3‐O‐butanoyl‐2,6‐di‐O‐pentyl)‐γ‐cyclodextrin (Lipodex‐E) in NMR and sensor experiments. We therefore investigated the retention behavior of MCP enantiomers on Lipodex‐E by gas chromatography (GC) under hydrous conditions. Addition of water to the N₂ carrier gas modestly reduced the retention factors k of the enantiomers, notably for the second eluted enantiomer (S)‐MCP. This resulted in an overall decrease of enantioselectivity ‐Δ_S,R(ΔG) in the presence of water. The effect was fully reversible. Consequently, for a conditioned column in the absence of residual water, the determined thermodynamic data, i.e. Δ_S,R(ΔH) = –12.64 ± 0.08 kJ mol^‐1 and Δ_S,R(ΔS) = –28.18 ± 0.23 J K^‐1 mol^‐1, refer to a true 1:1 complexation process devoid of hydrophobic hydration. Chirality 28:124–131, 2015. © 2015 Wiley Periodicals, Inc.     

Preparative Enantioseparation of β‐Substituted‐2‐Phenylpropionic Acids by Countercurrent Chromatography With Substituted β‐Cyclodextrin as Chiral Selectors

Preparative enantioseparation of four β‐substituted‐2‐phenylpropionic acids was performed by countercurrent chromatography with substituted β‐cyclodextrin as chiral selectors. The two‐phase solvent system was composed of n‐hexane‐ethyl acetate‐0.10 mol L‐1 of phosphate buffer solution at pH 2.67 containing 0.10 mol L^‐1 of hydroxypropyl‐β‐cyclodextrin (HP‐β‐CD) or sulfobutylether‐β‐cyclodextrin (SBE‐β‐CD). The influence factors, including the type of substituted β‐cyclodextrin, composition of organic phase, concentration of chiral selector, pH value of the aqueous phase, and equilibrium temperature were optimized by enantioselective liquid–liquid extraction. Under the optimum separation conditions, 100 mg of 2‐phenylbutyric acid, 100 mg of tropic acid, and 50 mg of 2,3‐diphenylpropionic acid were successfully enantioseparated by high‐speed countercurrent chromatography, and the recovery of the (±)‐enantiomers was in the range of 90–91% for (±)‐2‐phenylbutyric acid, 91–92% for (±)‐tropic acid, 85–87% for (±)‐2,3‐diphenylpropionic acid with purity of over 97%, 96%, and 98%, respectively. The formation of 1:1 stoichiometric inclusion complex of β‐substituted‐2‐phenylpropionic acids with HP‐β‐CD was determined by UV spectrophotometry and the inclusion constants were calculated by a modified Benesi‐Hildebrand equation. The results showed that different enantioselectivities among different racemates were mainly caused by different enantiorecognition between each enantiomer and HP‐β‐CD, while it might be partially caused by different inclusion capacity between racemic solutes and HP‐β‐CD. Chirality 27:795–801, 2015. © 2015 Wiley Periodicals, Inc.     

A Validated Normal Phase LC Method for Enantiomeric Separation of Rasagiline Mesylate and Its (S)‐Enantiomer on Cellulose Derivative‐Based Chiral Stationary Phase

A simple, sensitive, and robust normal‐phase isocratic HPLC‐UV method was developed and validated for the enantiomeric separation of rasagiline mesylate and its (S)‐enantiomer. The rasagiline and its (S)‐enantiomer were resolved on a Chiralcel‐OJ‐H (4‐methylbenzoate cellulose coated on silica) column using a mobile phase consisting of n‐hexane:isopropyl alcohol:ethanol:diethyl amine (96:2:2:0.01) at a flow rate of 1.0 ml/min. The column temperature was maintained at 27 °C and elution was monitored at 215 nm. The resolution (R_s) between the enantiomers was found to be more than 2.0. The limit of detection and the limit of quantification of the (S)‐enantiomer were found to be 0.35 and 1.05 µg/ml, respectively. The developed method was validated as per ICH guidelines with respect to linearity, limit of detection and quantification, accuracy, precision, and robustness—and satisfactory results were obtained. The sample solution and mobile phase were found to be stable up to 48 h. The method is useful for routine evaluation of the quality of rasagiline mesylate in bulk drug‐manufacturing units. Chirality 25:324–327, 2013. © 2013 Wiley Periodicals, Inc.     

Enantiomeric resolution of some nonsteroidal antiinflammatory and anticoagulant drugs using β‐cyclodextrins by capillary electrophoresis

β‐cyclodextrin (CD) and its derivatives HP‐β‐CD, DM‐β‐CD, and TM‐β‐CD have been employed as chiral selectors for the separation of three nonsteroidal antiinflammatory drugs (NSAIDs) and anticoagulant at relatively low concentration (8–15 mM) by capillary zone electrophoresis (CZE). In this study, baseline separation was achieved for ibuprofen, ketoprofen, naproxen, and warfarin. It was found that the addition of 0.1% hydroxypropyl methyl cellulose (HPMC) was effective for separation. Under these conditions, the S‐(+) enantiomer eluted before R‐(−) in terms of ibuprofen; the calculated energy values obtained from the molecular modeling correlated well with the elution order. An equation for calculating the pK_a values by capillary electrophoresis was introduced, and the pK_a values of the four chiral drugs at 25°C were obtained based on the equation. The value pK_a + 0.5 is proposed to be the suitable pH of the background electrolyte for the separation of chiral compounds containing a carboxylic group. Chirality 11:56–62, 1999. © 1999 Wiley‐Liss, Inc.     

Screening Approach for Chiral Separation of β‐Aminoketones by HPLC on Various Polysaccharide‐Based Chiral Stationary Phases

Nine β‐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 β‐amino ketones. Chirality 27:332–338, 2015. © 2015 Wiley Periodicals, Inc.     

Chiral Separation of Cathinone and Amphetamine Derivatives by HPLC/UV Using Sulfated ß‐Cyclodextrin as Chiral Mobile Phase Additive

In the last years the identification of new legal and illegal highs has become a huge challenge for the police and prosecution authorities. In an analytical context, only a few analytical methods are available to identify these new substances. Moreover, many of these recreational drugs are chiral and it is supposed that the enantiomers differ in their pharmacological potency. Since nonenantioselective synthesis is easier and cheaper, they are mainly sold as racemic mixtures. The goal of this research work was to develop an inexpensive method for the chiral separation of cathinones and amphetamines. This should help to discover if the substances are sold as racemic mixtures and give further information about their quality as well as their origin. Chiral separation of a set of 6 amphetamine and 25 cathinone derivatives, mainly purchased from various Internet shops, is presented. A LiChrospher 100 RP‐18e, 250 x 4 mm, 5 µm served as the stationary phase. The chiral mobile phase consisted of methanol, water, and sulfated ß‐cyclodextrin. Measurements were performed under isocratic conditions in reversed phase mode using UV detection. Four model compounds of the two substance classes were used to optimize the mobile phase. Under final conditions (methanol:water 2.5:97.5 + 2% sulfated ß‐cyclodextrin) enantiomers of amphetamine and five derivatives were baseline separated within 23 min. In all, 17 cathinones were completely or partially chirally separated. However, as only 3 of 25 cathinones were baseline resolved, the application of this method is limited for cathinone analogs. Additionally, the results were compared with an RP‐8e column. Chirality 26:411–418, 2014. © 2014 Wiley Periodicals, Inc.     

Stereodynamics of Nitrogen Chiral Centers in aza‐β3‐Cyclodipeptides

The present work is devoted to the synthesis, conformational analysis, and stereodynamic study of aza‐β³‐cyclodipeptides. This pseudopeptidic ring shows E/Z hydrazide bond isomerism, eight‐membered ring conformation, and chirotopic nitrogen atoms, all of which are elements that are prone to modulate the ring shape. The (E,E) twist boat conformation observed in the solid state by X‐ray diffraction is also the ground conformation in solution, and emerges as the lowest in energy when using quantum chemical calculations. The relative configuration associated with ring chirality and with the two nitrogen chiral centers is governed by steric crowding and adopts the (P)S_NS_N/(M)R_NR_N combination which projects side chains in equatorial position. The nitrogen pyramidal inversion (NPI) at the two chiral centers is correlated with the ring reversal. The process is significantly hindered as was shown by VT‐NMR experiments run in C₂D₂Cl₄, which did not make it possible to determine the barrier to inversion. Finally, these findings make it conceivable to resolve enantiomers of aza‐β³‐cyclodipeptides by modulating the backbone decoration appropriately. Chirality 25:341–349, 2013. © 2013 Wiley Periodicals, Inc.     

β‐Cyclodextrin hydrogels containing naphthaleneacetic acid for pH‐sensitive release

β‐Cyclodextrin (β‐CD) hydrogel was prepared in a strong alkali condition using epichlorohydrin (EPI) as a cross‐linker, where the molar ratios of EPI to β‐CD were 8:1, 10:1, and 15:1. In order to endow a pH sensitivity to the hydrogel, naphthaleneacetic acid (NAA) was loaded in the hydrogel by taking advantage of its hydrophobic interaction with the cavities of β‐CD. The releases of blue dextran (a water‐soluble dye) from the hydrogels were promoted, as the pHs of the media increased. When the molar ratio of EPI to β‐CD was lower, the degrees of release were higher, and the pH dependency of the release became more prominent. In fact, the swelling ratio of the hydrogels having a lower molar ratio of EPI to β‐CD was higher. The higher swelling ratio would account for the higher degree of release and the marked pH sensitivity. Biotechnol. Bioeng. 2010;106: 295–302. © 2010 Wiley Periodicals, Inc.     

Geometrical principles of homomeric β‐barrels and β‐helices: Application to modeling amyloid protofilaments

Examples of homomeric β‐helices and β‐barrels have recently emerged. Here we generalize the theory for the shear number in β‐barrels to encompass β‐helices and homomeric structures. We introduce the concept of the “β‐strip,” the set of parallel or antiparallel neighboring strands, from which the whole helix can be generated giving it n‐fold rotational symmetry. In this context, the shear number is interpreted as the sum around the helix of the fixed register shift between neighboring identical β‐strips. Using this approach, we have derived relationships between helical width, pitch, angle between strand direction and helical axis, mass per length, register shift, and number of strands. The validity and unifying power of the method is demonstrated with known structures including α‐hemolysin, T4 phage spike, cylindrin, and the HET‐s(218‐289) prion. From reported dimensions measured by X‐ray fiber diffraction on amyloid fibrils, the relationships can be used to predict the register shift and the number of strands within amyloid protofilaments. This was used to construct models of transthyretin and Alzheimer β(40) amyloid protofilaments that comprise a single strip of in‐register β‐strands folded into a “β‐strip helix.” Results suggest both stabilization of an individual β‐strip helix and growth by addition of further β‐strip helices can involve the same pair of sequence segments associating with β‐sheet hydrogen bonding at the same register shift. This process would be aided by a repeat sequence. Hence, understanding how the register shift (as the distance between repeat sequences) relates to helical dimensions will be useful for nanotube design.     

Enantioseparation of Tröger's Base Derivatives by Capillary Electrophoresis Using Cyclodextrins as Chiral Selectors

The enantioseparation of seven Tröger's base derivatives (TBs) was carried out by capillary electrophoresis using α‐, β‐, and γ‐cyclodextrins as chiral selectors and phosphate at 20 mmol/l concentration, pH 2.5, as background electrolyte. The method was optimized with respect to the concentration of chosen chiral selectors (0–50 mmol/l) and the amount of organic solvent (acetonitrile, 0–25 % (v/v)) in the electrolyte. The results indicate that all the studied variables, i.e., type of chiral selector, its concentration, and the amount of the added organic solvent, have a significant impact on the enantioseparation of the studied TBs. The best results for the majority of the separated TBs were obtained utilizing β‐cyclodextrin at 5 mmol/l concentration and with various amounts of acetonitrile added ranging from 5 to 15% (v/v) in the background electrolyte. For the two smallest studied TBs, γ‐cyclodextrin with 10% (v/v) acetonitrile also provided good resolution. Chirality 25:379–383, 2013. © 2013 Wiley Periodicals, Inc.     

Establishment and Evaluation of the Novel Tetramethylammonium‐L‐Hydroxyproline Chiral Ionic Liquid Synergistic System Based on Clindamycin Phosphate for Enantioseparation by Capillary Electrophoresis

Much attention has been paid to chiral ionic liquids (ILs) in analytical chemistry, especially its application in capillary electrophoresis (CE) enantioseparation. However, the investigation of chiral ionic liquids synergistic systems based on antibiotic chiral selectors has been reported in only one article. In this work, a novel chiral ionic liquid, tetramethylammonium‐L‐hydroxyproline (TMA‐L‐Hyp), was applied for the first time in CE chiral separation to evaluate its potential synergistic effect with clindamycin phosphate (CP) as the chiral selector. As observed, significantly improved separation was obtained in this TMA‐L‐Hyp/CP synergistic system compared to TMA‐L‐Hyp or a CP single system. Several primary factors that might influence the separation were investigated, including CP concentration, TMA‐L‐Hyp concentration, buffer pH, types and concentrations of organic modifier, applied voltage, and capillary temperature. The best results were obtained with a 40 mM borax buffer (pH 7.6) containing 30 mM TMA‐L‐Hyp, 80 mM CP, and 20% (v/v) methanol, while the applied voltage and temperature were set at 20 kV and 20°C, respectively. Chirality 27:598–604, 2015. © 2015 Wiley Periodicals, Inc.     

一种新的测定蛋白激酶活性的方法──毛细管电泳测定蛋白激酶A的活性

建立了以毛细管电泳为基础的测定蛋白激酶A活性的新方法,可作为激酶测活的通用方法．此法基于底物及其磷酸化产物很容易在毛细管电泳中分开,且酶活力可用积分值计算,同时又发展了连续进样技术,能在一次电泳中同时进行10个以上的酶活性测定,新方法操作简单,灵敏度和精确性均优于常规的同位素法．     

β‐Bulges: Extensive structural analyses of β‐sheets irregularities

β‐Sheets are quite frequent in protein structures and are stabilized by regular main‐chain hydrogen bond patterns. Irregularities in β‐sheets, named β‐bulges, are distorted regions between two consecutive hydrogen bonds. They disrupt the classical alternation of side chain direction and can alter the directionality of β‐strands. They are implicated in protein‐protein interactions and are introduced to avoid β‐strand aggregation. Five different types of β‐bulges are defined. Previous studies on β‐bulges were performed on a limited number of protein structures or one specific family. These studies evoked a potential conservation during evolution. In this work, we analyze the β‐bulge distribution and conservation in terms of local backbone conformations and amino acid composition. Our dataset consists of 66 times more β‐bulges than the last systematic study (Chan et al. Protein Science 1993, 2:1574–1590). Novel amino acid preferences are underlined and local structure conformations are highlighted by the use of a structural alphabet. We observed that β‐bulges are preferably localized at the N‐ and C‐termini of β‐strands, but contrary to the earlier studies, no significant conservation of β‐bulges was observed among structural homologues. Displacement of β‐bulges along the sequence was also investigated by Molecular Dynamics simulations.