Effect of fabrication strategy on the enantioseparation performance of β‐cyclodextrin‐functionalized polymethacrylate monoliths: A comparative evaluation

To evaluate the effect of the preparation strategy on the enantioseparation performance of β‐cyclodextrin‐functionalized monoliths, a series of β‐cyclodextrin‐functionalized organic polymeric monolithic columns were prepared through two‐step, single‐step, and one‐pot approaches, using the same cyclodextrin, linker–spacer, and crosslinker. Physicochemical characterization of the columns was carried out by determining the morphology, β‐cyclodextrin density, permeability, and chromatographic efficiency. For each type of monolithic column, the enantioresolution of 22 chiral compounds, including mandelic acid derivatives, nonsteroidal anti‐inflammatory drugs, N‐derivatized amino acids, and herbicides, was comparatively studied under optimum chromatographic conditions. The β‐cyclodextrin‐functionalized monolithic columns prepared through the one‐pot approach exhibited higher enantioresolution for most chiral compounds, and they have the advantage of good controllability and simple preparation. On the other hand, the enantioresolution obtained on columns prepared through the single‐step approach was quite unsatisfactory, and therefore the effect of using different linking spacers and crosslinkers was studied. A significant improvement of enantioresolution for 2‐chloro‐mandelic acid was obtained by using N ,N‐methylenebisacrylamide instead of ethylene dimethacrylate as the crosslinker in the single‐step preparation.     

Use of various β‐cyclodextrin derivatives as chiral selectors for the enantiomeric separation of ofloxacin and its five related substances by capillary electrophoresis

A capillary electrophoretic method for the enantioseparation of ofloxacin and its five related substances (potential impurities, indicated as impurities B–F) was developed using β‐cyclodextrin derivatives as chiral selectors. To our knowledge, there are no previous studies about using capillary electrophoresis for the separation of impurities B–D. Six β‐cyclodextrin derivatives including cationic (piperidine‐ and cyclohexylamine‐), neutral (dimethyl‐ and hydroxypropyl‐), and anionic (carboxymethyl‐ and sulfated‐) β‐cyclodextrin derivatives were tested and operational parameters such as buffer pH and concentration of β‐cyclodextrin derivatives were investigated. The best resolutions were all obtained with anionic β‐cyclodextrin derivatives: ofloxacin, impurities C–F could be best resolved with carboxymethyl‐β‐cyclodextrin at satisfactory resolutions of 8.27, 9.98, 5.92, 8.49 and 6.78, respectively, while for impurity B, a particularly impressive resolution value, up to 21.38, was observed using sulfated‐β‐cyclodextrin. The enhancement of enantioseparation observed for the tested analytes using anionic β‐cyclodextrin derivatives might be due to some favorable interaction between selectors and analytes. Given the fact that the selection of chiral selector depends on the structures of analytes, with the help of structural similarities and differences of the analytes, the structure–separation relationship was further discussed.     

Monolithic silica columns functionalized with substituted polyproline‐derived chiral selectors as chiral stationary phases for high‐performance liquid chromatography

In this study, two polyproline‐derived chiral selectors are bonded to monolithic silica gel columns. In spite of high chiral selector coverage, the derivatization was found to have only a slight effect on the hydrodynamics of the mobile phase through the column. The enantioseparation ability of the resulting chiral monolithic columns was evaluated with a series of structurally diverse racemic test compounds. When compared to analogous bead‐based chiral stationary phases, higher enantioseparation and broader application domain were observed for monolithic columns. Moreover, the increase in flow rate produces a minor reduction of resolution, which permits to shorten analysis time. Additionally, increased loadability defines chiral polyproline derived monoliths as adequate for preparative chromatography.     

Chiral separation of cathinone derivatives using β‐cyclodextrin‐assisted capillary electrophoresis–Comparison of four different β‐cyclodextrin derivatives used as chiral selectors

In the past decade, more than 100 different cathinone derivatives slopped over entire Europe due to their enormous popularity. Generally, these novel psychoactive substances are easily available via the internet. This fact leads to various social problems, since cathinones are substances with consciousness‐changing effects and are mainly misused for recreational matters by their consumers. Cathinones possess a chiral center including two enantiomeric forms with potentially different pharmacological behavior. This fact makes analytical method development regarding their chiral separation indispensable. In this study, a chiral capillary zone electrophoresis method for the enantioseparation of 61 cathinone and pyrovalerone derivatives was developed by means of four different β‐cyclodextrin derivatives. As chiral selectors, native β‐cyclodextrin as well as three of its derivatives namely acetyl‐β‐cyclodextrin, 2‐hydroxypropyl‐β‐cyclodextrin, and carboxymethyl‐β‐cyclodextrin were used. The cathinone and pyrovalerone derivatives were either purchased in internet stores or seized by police. As a result, overall 58 of 61 studied substances were partially or baseline separated by at least one of the four chiral selectors using 10 mM of β‐cyclodextrin derivative in a 10 mM sodium phosphate buffer (pH 2.5). Furthermore, the method was found to be suitable for simultaneous enantioseparations, for enantiomeric purity checks and to differentiate between positional isomers. Moreover, an intra‐ and an interday validation was performed successfully for each chiral selector to prove the robustness of the method.     

Comparative evaluation of a one‐pot strategy for the preparation of β‐cyclodextrin‐functionalized monoliths: Effect of the degree of amino substitution of β‐cyclodextrin on the column performance

To further evaluate the feasibility and applicability of the one‐pot strategy in monolithic column preparation, two novel β‐cyclodextrin‐functionalized organic polymeric monoliths were prepared using two β‐cyclodextrin derivatives, i.e. mono(6‐amino‐6‐deoxy)‐β‐cyclodextrin and heptakis(6‐amino‐6‐deoxy)‐β‐cyclodextrin. In this improved method, mono(6‐amino‐6‐deoxy)‐β‐cyclodextrin or heptakis(6‐amino‐6‐deoxy)‐β‐cyclodextrin reacted with glycidyl methacrylate to generate the corresponding functional monomers and were subsequently copolymerized with ethylene dimethacrylate. The polymerization conditions for both monoliths were carefully optimized to obtain satisfactory column performance with respect to column efficiency, reproducibility, permeability, and stability. The obtained poly(glycidyl methacrylate‐mono(6‐amino‐6‐deoxy)‐β‐cyclodextrin‐co‐ethylene dimethacrylate) and poly(glycidyl methacrylate‐heptakis(6‐amino‐6‐deoxy)‐β‐cyclodextrin‐co‐ethylene dimethacrylate) monoliths exhibited a uniform structure, good permeability, and mechanical stability as indicated by scanning electron microscopy and micro‐high‐performance liquid chromatography experimental results. Because of the probable existence of multi‐glycidyl methacrylate linking spacers on the poly(glycidyl methacrylate‐heptakis(6‐amino‐6‐deoxy)‐β‐cyclodextrin‐co‐ethylene dimethacrylate) monolith, the effect of the ratio of glycidyl methacrylate/heptakis(6‐amino‐6‐deoxy)‐β‐cyclodextrin was especially studied, and satisfactory reproducibility could still be achieved by strictly controlling the composition of the polymerization mixture. To investigate the effect of the degree of amino substitution of β‐cyclodextrin on column performance, a detailed comparison of the two monoliths was also carried out using series of analytes including small peptides and chiral acids. It was found that the β‐cyclodextrin‐functionalized monolith with mono‐glycidyl methacrylate linking spacers demonstrated better chiral separation performance than that with multi‐glycidyl methacrylate linking spacers.     

A novel in situ strategy for the preparation of a β‐cyclodextrin/polydopamine‐coated capillary column for capillary electrochromatography enantioseparations

Inspired by the chiral recognition ability of β‐cyclodextrin and the natural adhesive properties of polydopamine under alkaline conditions, in this study, a rapid and in situ modification strategy was developed to fabricate β‐cyclodextrin/polydopamine composite material coated‐capillary columns for open tubular capillary electrochromatography. The results of scanning electron microscopy, FTIR spectroscopy, streaming potential, and electro‐osmotic flow studies indicated that β‐cyclodextrin/polydopamine was successfully fixed on the inner wall of the capillary column. This coating can be achieved within 1 h affording a greatly reduced capillary preparation time. The performance of the β‐cyclodextrin/polydopamine‐coated capillary was validated by the analysis of seven pairs of chiral analytes, namely epinephrine, norepinephrine, isoprenaline, terbutaline, verapamil, tryptophane, carvedilol. Good enantioseparation efficiencies were achieved for all. For three consecutive runs, the relative standard deviations for the migration times of the analytes for intraday, interday, and column‐to‐column repeatability were in the range of 0.41–1.74, 1.03–4.18, and 1.66–8.24%, respectively. Moreover, the separation efficiency of the β‐cyclodextrin/polydopamine‐coated capillary column did not decrease obviously over 90 runs. The strategy should also be feasible to introduce and immobilize other chiral selectors on the inner walls surface of capillary columns.     

Comparative analysis of the full set of methylated β‐cyclodextrins as chiral selectors in capillary electrophoresis

The chiral separation ability of the full library of methylated‐β‐cyclodextrins towards pharmacologically significant racemic drugs including basic compounds was studied by chiral CE. The syntheses of all the methylated, single isomer β‐cyclodextrins were revised and optimized and the aqueous solubility of the derivatives was unambiguously established. The three most relevant commercially available methylated isomeric mixtures were also included in the screening, so a total of ten various methylated CDs were investigated. The effects of the selector concentration on the enantiorecognition properties at acidic pH were investigated. Among the dimethylated β‐cyclodextrins, the heptakis (2,6‐di‐O‐methyl)‐β‐cyclodextrin isomer (2,6‐DIMEB) resulted to be the most versatile chiral selector. Terbutaline was selected as a model compound for the in‐depth investigation of host‐guest enantiodiscrimination ability. The association constants between the two terbutaline enantiomers and 2,6‐DIMEB were determined in order to support that the enantioseparation is driven by differences is host‐guest binding. The migration order of the enantiomers was confirmed by performing spiking experiments with the pure enantiomers. 1D and 2D NMR spectroscopy was applied to the 2,3‐, and 2,6‐DIMEB/terbutaline systems to rationalize at molecular level the different enantioseparation ability of the dimethylated β‐cyclodextrin selectors.     

Polymer‐based monolithic column with incorporated chiral metal–organic framework for enantioseparation of methyl phenyl sulfoxide using nano‐liquid chromatography

A new approach to the preparation of enantioselective porous polymer monolithic columns with incorporated chiral metal–organic framework for nano‐liquid chromatography has been developed. While no enantioseparation was achieved with monolithic poly(4‐vinylpyridine‐co‐ethylene dimethacrylate) column, excellent separations of both enantiomers of (±)‐methyl phenyl sulfoxide were achieved with its counterpart prepared after admixing metal–organic framework [Zn₂(benzene dicarboxylate)(l‐lactic acid)(dmf)], which is synthesized from zinc nitrate, l ‐lactic acid, and benzene dicarboxylic acid in the polymerization mixture. These novel monolithic columns combined selectivity of the chiral framework with the excellent hydrodynamic properties of polymer monoliths, may provide a great impact on future studies in the field of chiral analysis by liquid chromatography.     

Comparison of enantioseparation of selected benzodiazepine and phenothiazine derivatives on chiral stationary phases based on β‐cyclodextrin and macrocyclic antibiotics

Enantioselective separation of some phenothiazine and benzodiazepine derivatives was studied on six different chiral stationary phases (CSPs) in HPLC. Selected CSPs, with respect to the structure of the separated compounds, were either based on β‐cyclodextrin chiral selectors – underivatized β‐cyclodextrin and hydroxypropyl ether β‐cyclodextrin, or on macrocyclic antibiotics – vancomycin, teicoplanin, teicoplanin aglycone, and ristocetin A. Measurements were carried out in a reversed‐phase separation mode. The influence of mobile phase composition on retention and enantioseparation was studied. Benzodiazepines could be enantioresolved with almost all the chiral stationary phases used, except for the vancomycin‐bonded CSP. Peak coalescence of oxazepam and lorazepam was observed if separation was carried out at laboratory temperature. Reduced temperature was required in some instances in order to avoid the on‐column racemization. Separation systems composed of teicoplanin‐bonded CSP and buffer‐methanolic or pure methanolic mobile phases were shown to be suitable even for preparative purposes due to high resolution values of the enantiomers. Enantioseparation of phenothiazine derivatives was more difficult to achieve but it was successful, at least partly, also with both types of the CSPs used (except for levomepromazine).     

(l)- or (d)-Valine tert-butylamide grafted on permethylated β-cyclodextrin derivatives as new mixed binary chiral selectors: Versatile tools for capillary gas chromatographic enantioseparation

This work deals with the synthesis of two mixed binary chiral selectors prepared by grafting (l)- or (d)-valine tert-butylamide on permethylated cyclodextrin macrocycle. The enantioselective properties of the new chiral selectors diluted in OV11 polysiloxane (35% phenyl- and 65% methylsiloxane) were investigated by means of injections of 117 racemic mixtures. The mixed chiral selectors with (l)-valine and, to a lesser extent with (d)-valine, were found to have an improved enantioselectivity toward amino acid derivatives by comparison to permethylated cyclodextrin. The enantioseparation capability of these new chiral selectors has proven to be slightly less efficient than Chirasil-l-Val (Alltech) for amino acid derivatives, but it has been extended to include terpenes, lactones, esters, aliphatic compounds and aryl alcohols.     

The study of enantioselectivity of all regioisomers of mono‐carboxymethyl‐β‐cyclodextrin used as chiral selectors in CE

This work documents the influence of the position of single carboxymethyl group on the β‐cyclodextrin skeleton on the enantioselectivity. These synthesized monosubstituted carboxymethyl cyclodextrin (CD) derivatives, native β‐cyclodextrin, and commercially available carboxymethyl‐β‐cyclodextrin with degree of substitution approximately 3 were used as additives into the BGE consisting of phosphate buffer at 20 mmol/L concentration, pH 2.5, and several biologically significant low‐molecular‐mass chiral compounds were enantioseparated by CE. The results indicate that different substituent location on β‐cyclodextrin skeleton has a significant influence on the enantioseparation of the investigated enantiomers. The enantioselectivity of 2^I‐O‐regioisomer was better than with native β‐cyclodextrin. Comparable results to native β‐cyclodextrin were obtained for 6^I‐O‐ regioisomer and the enantioselectivity of 3^I‐O‐regioisomer was even worse than with native β‐cyclodextrin. Commercially available derivative of CD provides better resolutions than the monosubstituted carboxymethyl CD derivatives for most of the investigated analytes.     

The influence of the substituent position in monocarboxymethyl‐γ‐cyclodextrins on enantioselectivity in capillary electrophoresis

Three newly synthesized chiral selectors, namely, 2^I‐O‐, 3^I‐O‐, and 6^I‐O‐carboxymethyl‐γ‐cyclodextrin, native γ‐cyclodextrin, and commercially available carboxymethylated γ‐cyclodextrin with degree of substitution of 3–6 were used as additives in a background electrolyte composed of phosphate buffer at 20 mmol/L concentration and pH 2.5. This system was used for the analysis of several biologically significant low‐molecular‐mass chiral compounds by capillary electrophoresis. The results confirmed that the position of carboxymethyl group influences the enantioseparation efficiency of all the studied analytes. The 2^I‐O‐ and 3^I‐O‐ regioisomers provide a significantly better resolution than native γ‐cyclodextrin, while the 6^I‐O‐regioisomer gives only a slightly better enantioseparation than native γ‐cyclodextrin. The application of γ‐cyclodextrin possessing higher number of carboxymethyl groups led to the best resolution for the majority of the compounds analyzed.     

A new single‐urea‐bound 3,5‐dimethylphenylcarbamoylated β‐cyclodextrin chiral stationary phase and its enhanced separation performance in normal‐phase liquid chromatography

A new single‐urea‐bound chiral stationary phase based on 3,5‐dimethylphenylcarbamoylated β‐cyclodextrin was prepared through the Staudinger reaction of mono (6^A‐azido‐6^A‐deoxy)‐per(3,5‐dimethylphenylcarbamoylated) β‐cyclodextrin and 3‐aminopropyl silica gel under CO₂ atmosphere. The new phase exhibited good enantioseparation performance for 33 analytes using normal‐phase HPLC conditions; 19 of them were baseline separated. Effects of structure of analytes, alcoholic modifiers, and acidic/basic additives on separation performances of this new cyclodextrin chiral stationary phase have been studied in detail. The results showed that the retention and resolution of acidic and basic analytes on the CSP were greatly affected by the additives. Peak symmetry for some analytes could be improved by simultaneously adding acidic and basic additives to the mobile phase. This work expands the potential applications of the cyclodextrin‐based chiral stationary phases in the normal‐phase HPLC.     

毛细管电泳手性分离中的协同效应

综述了毛细管电泳手性分离中的协同效应。介绍了毛细管电泳手性分离中双手性选择剂的应用情况。表明用CDs/CDs.CDs/crown组成的双选择剂及聚合环糊精衍生物,聚合手性胶束体系有可能改善难拆分的对映体物质的分离效果,展示了协同效应的毛细管电泳拆分复杂物质对映体中的应用前景。     

One‐step strategy for the synthesis of a derivatized cyclodextrin‐based monolithic column

Derivatized β‐cyclodextrin (β‐CD) functionalized monolithic columns were prepared by a “one‐step” strategy using click chemistry. First, the intended derivatized β‐CD monomers were synthesized by a click reaction between propargyl methacrylate and mono‐6‐azido‐β‐CD and then sulfonation or methylation was carried out. Finally, monolithic columns were prepared through a one‐step in situ copolymerization of the derivatized β‐CD monomer and ethylene glycol dimethacrylate. The sulfated β‐CD‐based monolith was successfully applied to the hydrophilic interaction liquid chromatography separation of nucleosides and small peptides, while the methylated β‐CD‐functionalized monolith was useful for the separation of nonpolar compounds and drug enantiomers in capillary reversed‐phase liquid chromatography. The structures of the monomers were characterized by Fourier transform infrared spectroscopy and mass spectrometry. The physicochemical properties and column performance of monoliths were evaluated by scanning electron microscopy and micro high performance liquid chromatography. This strategy has considerable prospects for the preparation of other derivatized CD‐functionalized methacrylate monoliths.     

Enantiomer separation of chiral pharmaceuticals by capillary electrochromatography

Enantiomer separation of chiral pharmaceuticals by capillary electrochromatography (CEC) is achieved with open-tubular capillaries (o-CEC), with packed capillaries (p-CEC) or with monolithic capillaries. In o-CEC, capillaries are coated with a thin film containing cyclodextrin derivatives, cellulose, proteins, poly-terguride or molecularly imprinted polymers as chiral selectors. In p-CEC, typical chiral HPLC stationary phases such as silica-bonded cyclodextrin or cellulose derivatives, proteins, glycoproteins, macrocyclic antibiotics, quinine-derived and 'Pirkle' selectors, polyacrylamides and molecularly imprinted polymers are used as chiral selectors. Chiral monolithic stationary phases prepared by in situ polymerization into the capillary were also developed for electrochromatographic enantiomer separation.     

Recent developments in chiral analysis of β‐blocker drugs by capillary electromigration techniques

The latest developments in chiral analysis of β‐blocker drugs by capillary electromigration techniques are reviewed in this article. Following the previous review by Aturki et al. [Electrophoresis 2011, 32, 2602–2628], this review includes the papers published during the period from January 2011 to December 2013. During this time, some novel chiral selectors were reported and applied to improve the enantioseparation of β‐blocker drugs and structurally related compounds. These chiral selectors include CDs and their derivatives, macrocyclic antibiotics, tartrate complexs, the monolithic molecularly imprinted polymer, and the polymeric surfactants. In addition, this article summarizes the methodological improvements for enhancing sensitivity in chiral analysis of β‐blockers and structurally related compounds by CE. The involved authors described the use of online sample preconcentration techniques to increase the detection sensitivity in the enantiomeric analysis of a broad range of samples.     

A facile and efficient one‐step strategy for the preparation of β‐cyclodextrin monoliths

A novel, facile, and efficient one‐step copolymerization strategy was developed for the preparation of β‐cyclodextrin (β‐CD) methacrylate monolithic columns using click chemistry. The novel mono‐(1H‐1,2,3‐triazol‐4‐ylmethyl)‐2‐methylacryl‐β‐CD monomer was synthesized by a click reaction between propargyl methacrylate and mono‐6‐azido‐β‐CD, and then monolithic columns were prepared through a one‐step in situ copolymerization of the mono‐(1H‐1,2,3‐triazol‐4‐ylmethyl)‐2‐methylacryl‐β‐CD monomer and ethylene dimethacrylate. The physicochemical properties and column performance of the fabricated monolithic columns were characterized by elemental analysis, SEM, and micro‐HPLC. Satisfactory column permeability, efficiency, and separation performance were obtained for the optimized poly(mono‐(1H‐1,2,3‐triazol‐4‐ylmethyl)‐2‐methylacryl‐β‐CD‐co‐ethylene dimethacrylate) monolithic columns. Additionally, typical hydrophilic interaction chromatography retention behavior was observed on the monoliths at high acetonitrile content in the mobile phase. Although the enantioselectivity of our monolithic columns did not meet the level of other reported β‐CD monolithic columns, this one‐step strategy based on click chemistry still provides an interesting and effective model as it offers the possibility to easily prepare related novel CD methacrylate monoliths through a one‐step copolymerization strategy.     

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.     

Characterization of complexes between phenethylamine enantiomers and β‐cyclodextrin derivatives by capillary electrophoresis—Determination of binding constants and complex mobilities

To optimize chiral separation conditions and to improve the knowledge of enantioseparation, it is important to know the binding constants K between analytes and cyclodextrins and the electrophoretic mobilities of the temporarily formed analyte‐cyclodextrin‐complexes. K values for complexes between eight phenethylamine enantiomers, namely ephedrine, pseudoephedrine, methylephedrine and norephedrine, and four different β‐cyclodextrin derivatives were determined by affinity capillary electrophoresis. The binding constants were calculated from the electrophoretic mobility values of the phenethylamine enantiomers at increasing concentrations of cyclodextrins in running buffer. Three different linear plotting methods (x ‐reciprocal, y ‐reciprocal, double reciprocal) and nonlinear regression were used for the determination of binding constants with β‐cyclodextrin, (2‐hydroxypropyl)‐β‐cyclodextrin, methyl‐β‐cyclodextrin and 6‐O‐α‐maltosyl‐β‐cyclodextrin. The cyclodextrin concentration in a 50 mM phosphate buffer pH 3.0 was varied from 0 to 12 mM. To investigate the influence of the binding constant values on the enantioseparation the observed electrophoretic selectivities were compared with the obtained K values and the calculated enantiomer‐cyclodextrin‐complex mobilities. The different electrophoretic mobilities of the temporarily formed complexes were crucial factors for the migration order and enantioseparation of ephedrine derivatives. To verify the apparent binding constants determined by capillary electrophoresis, a titration process using ephedrine enantiomers and β‐cyclodextrin was carried out. Furthermore, the isothermal titration calorimetry measurements gave information about the thermal properties of the complexes.