New cyclodextrin derivatives as chiral selectors in capillary electrophoresis

The separation of three pairs of enantiomeric herbicides has been successfully achieved by capillary electrophoresis at two different pH values in the presence of cyclodextrin derivatives previously synthesized in our laboratory. Two of these derivatives constitute a new class of receptor, the hemispherodextrins, in which a trehalose capping moiety is bonded to β-cyclodextrin. Because of their peculiar structure hemispherodextrins have very promising characteristics and the low receptor concentration required to achieve separation deserves particular interest.     

Comparing cyclodextrin derivatives as chiral selectors for enantiomeric separation in capillary electrophoresis

A total of 26 different cyclodextrin (CD) derivatives with different functional groups and degrees of substitution were tested against 35 basic pharmaceutical compounds in an effort to investigate their effectiveness as chiral selectors for enantiomeric separation in capillary electrophoresis (CE). Testing was performed under the same conditions using a low pH buffer (25 mM phosphate buffer at pH approximately 2.5). Five CD derivatives, namely, highly sulfated-beta-CD, highly sulfated-beta-CD, hydroxypropyl-beta-CD (degree of substitution approximately 1), heptakis-(2,6-O-dimethyl)-beta-CD, and heptakis(2,3,6-O-trimethyl)-beta-CD were identified to be most effective for enantiomeric separations and have a wide range of enantiomeric selectivity towards the model compounds. Over 90% of the model compounds were enantiomerically resolved with the five identified CD derivatives, at a minimum resolution of 0.5. An additional 20 compounds were also tested to demonstrate the validity of the identified CD derivatives. The five CD derivatives were recommended as the starting chiral selectors in developing enantiomeric separation methods by CE.     

New amphiphilic aminosaccharide derivatives as chiral selectors in capillary electrophoresis

Two amphiphilic aminosaccharide derivatives were investigated as chiral selector additives in capillary electrophoresis. Each substance has a glucosamine backbone carrying three hydrocarbon chains as the hydrophobic region and three carboxylic groups as the hydrophilic region, which is an artificial biologically active compound. Using each compound as a chiral selector, the optical resolution of dansylated amino acids or new quinolone antibacterial agents (NQs) was observed. Increasing the concentration of the chiral selector or the ionic strength of running solution led to successful optical resolution. In consideration of the chemical structure of each selector and the migration behavior of the enantiomers, the resolution seemed to be based on micellar electrokinetic chromatography mode. Both selectors differed in their enantioselectivity for dansylated amino acids or NQs although the chemical structures were similar.     

New single isomer negatively charged β‐cyclodextrin derivatives as chiral selectors in capillary electrophoresis

To improve resolution power of chiral selector and enantiomeric peak efficiency in CE, single isomer negatively charged β‐CD derivatives, mono(6‐deoxy‐6‐sulfoethylthio)‐β‐CD (SET‐β‐CD) bearing one negative charge and mono[6‐deoxy‐6‐(6‐sulfooxy‐5,5‐bis‐sulfooxymethyl)hexylthio]‐β‐CD (SMHT‐β‐CD) carrying three negative charges, were synthesized. The structure of these two β‐CD derivatives was confirmed by ¹H NMR and MS. SET‐β‐CD and SMHT‐β‐CD successfully resolved the enantiomers of several basic model compounds. SMHT‐β‐CD provided for a significantly greater enantioseparation than SET‐β‐CD at lower concentrations. This appears to be due to the higher binding affinity of SMHT‐β‐CD to the model compounds and the wider separation window resulting from an increased countercurrent mobility of the selector. Overall, the new chiral selectors provided enantioseparations with high peak efficiency while avoiding peak distortion due to polydispersive and electrodispersive effects. The information obtained from an apparent binding constant study suggested that the enantioseparation of the model compounds followed the predictions of charged resolving agent migration model and that the observed degree of enantioseparation difference were due to the magnitude of differences in both enantiomer‐chiral selector binding affinities (ΔK) and the mobilities of the complexed enantiomers (Δμ_c).     

Ligand exchange chiral separations by cyclodextrin derivatives in capillary electrophoresis

A beta-cyclodextrin substituted by an imidazole-bound histamine (CDmh) was successfully used to separate underivatised tryptophan racemate in capillary electrophoresis in the presence of copper(II) ion by the ligand exchange mechanism. To the best of our knowledge, this is the first example of a cyclodextrin derivative behaving as the first coordination sphere ligand in the complex added to the background electrolyte (BGE).     

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.     

Selected applications of cyclodextrin selectors in capillary electrophoresis.

Through the use of alpha-, beta-, gamma- and heptakis(2,6-di-O-methyl)-beta-cyclodextrin as stereospecific selectors or electrolyte modifiers, both in capillary zone electrophoresis and isotachophoresis, selected model isomeric compounds (including optical isomers) were resolved. Soluble alkylhydroxyalkylcellulose derivatives were further added to the cyclodextrin-modified background electrolytes under study. Their presence was found to be essential, as demonstrated by improvements in both enantioselectivity and separation efficiency. The results obtained in both electrophoretic modes, under optimized conditions, are compared and discussed.     

Enantioselective determination by capillary electrophoresis with cyclodextrins as chiral selectors

This review surveys the separation of enantiomers by capillary electrophoresis using cyclodextrins as chiral selector. Cyclodextrins or their derivatives have been widely employed for the direct chiral resolution of a wide number of enantiomers, mainly of pharmaceutical interest, selected examples are reported in the tables. For method optimisation, several parameters influencing the enantioresolution, e.g., cyclodextrin type and concentration, buffer pH and composition, presence of organic solvents or complexing additives in the buffer were considered and discussed. Finally, selected applications to real samples such as pharmaceutical formulations, biological and medical samples are also discussed.     

Hydroxyethylammonium monosubstituted cyclodextrin as chiral selector for capillary electrophoresis

A novel cationic cyclodextrin, mono-6^A-(2-hydroxyethyl-1-ammonium)-6^A-β-cyclodextrin chloride (HEtAMCD) has been successfully synthesized and applied as chiral selector in capillary electrophoresis. The NMR study revealed this chiral selector has three recognition sites: β-CD, ammonium cation and hydroxy group in the sidearm to contribute three corresponding driving forces including inclusion complexation, electrostatic interaction and hydrogen bonding. The effect of buffer pH and HEtAMCD concentration (2.5–10 mM) on enantioselectivity, chiral resolution as well as effective mobility of analytes was investigated. This elegantly designed CD exhibits outstanding enantioselectivities toward the studied hydroxyl acids and ampholytic racemates in CE with the aid of extra hydrogen bonding. Under optimum pH 6.0, chiral resolutions over 5 can be readily obtained for hydroxy acids with CD concentration below 5 mM. The comparison study between HEtAMCD and our earlier reported ammonium CDs indicates the hydroxyethylammonium group of HEtAMCD significantly increased the enantioselective capability.     

Enantiomer separation of drugs by capillary electrophoresis using proteins as chiral selectors

The separation of drug enantiomers using proteins as the chiral selectors in capillary electrophoresis (CE) is considered in this review. The proteins used include albumins such as bovine serum albumin, human serum albumin and serum albumins from other species, glycoproteins such as alpha1-acid glycoprotein, crude ovomucoid, ovoglycoprotein, avidin and riboflavin binding protein, enzymes such as fungal cellulase, cellobiohydrolase I, pepsin and lysozyme and other proteins such as casein, human serum transferrin and ovotransferrin. Protein-based CE is carried out in two modes: in one proteins are immobilized or adsorbed within the capillary, or protein-immobilized silica gels are packed into the capillary (affinity capillary electrochromatography mode), and in the other proteins are dissolved in the running buffer (affinity CE mode). Furthermore, the advantages and limitations of the two modes and the factors affecting the chiral separations of various drugs by protein-based CE are discussed.     

New derivatives of cyclodextrins as chiral selectors for the capillary electrophoretic separation of dichlorprop enantiomers

-, β- and γ-cyclodextrins (CDs), as well as some of their chemical derivatives, have been tested as chiral resolving agents for the capillary zone electrophoretic resolution of the racemic herbicide dichlorprop, (±)-2-(2,4-dichlorophenoxy)propionic acid, of which only the (+)-isomer is herbicidally active. The complexation constants of the herbicide enantiomers with the cyclodextrin host molecules have been calculated from the electrophoretic migration time data at variable cyclodextrin concentration. The experimental results showed that several of the investigated CDs allowed dichlorprop enantiomer resolution. In particular, a newly synthesised ethylcarbonate derivative of β-CD showed the best enantiomer resolution properties among the tested compounds, while the remaining ones showed inferior or no performances at all. The calculated inclusion constants allowed identification of the best conditions for enantioresolution, and an explanation of the different complexation properties of the investigated compounds has been proposed on the basis of molecular modeling.     

Enantiomeric separation of glycyl dipeptides by capillary electrophoresis with cyclodextrins as chiral selectors

Uncharged cyclodextrins were tested as chiral selectors for the enantiomeric separation of 13 glycyl dipeptides with capillary electrophoresis. Initial experiments were performed on 10 mmol/L of a cyclodextrin in 0.1 mol/L phosphoric acid -0.088 mol/L triethanolamine. Some of the resolved dipeptides were nonaromatic, which is noteworthy since, to our knowledge, no examples of the separation of small, nonaromatic molecules have been published. Mobility difference plots for Gly-DL-Leu and Gly-DL-Phe with heptakis(2,6-di-O-methyl)-beta-cyclodextrin showed relatively flat profiles in a large concentration range, which is an advantage for the development of robust quantitative analytical methods. The use of a background electrolyte (BGE) solution with pH 3.0 gave irreproducible results for two of the dipeptides, the acidic Gly-DL-Asp and Gly-DL-Glu; this pH is not advisable for the development of robust methods for these two peptides. The need for purer chiral selectors was demonstrated by comparing different batches of heptakis(2,6-di-Omethyl)-beta-cyclodextrin from the same supplier. A BGE consisting of malonic acid and triethanolamine was introduced to give better buffer capacity than the original BGE at pH 3.0.     

Enantioseparation of phenothiazines in cyclodextrin-modified capillary zone electrophoresis using sulfated cyclodextrins as chiral selectors

In this study, enantioseparations of five phenothiazines, including promethazine, ethopropazine, trimeprazine, methotrimeprazine, and thioridazine, in CD-modified CZE using dual CD systems consisting of randomly sulfate-substituted CD (MI-S-beta-CD) and a neutral CD as chiral selectors in a citrate buffer (100 mM) at pH 3.0 were investigated. The results indicate that MI-S-beta-CD is an excellent chiral selector for enantioseparation of ethopropazine. The enantiomers of promethazine can also be baseline-resolved with MI-S-beta-CD at concentrations in the range of 0.5-1.0% w/v. On the other hand, thioridazine and trimeprazine interact strongly with neutral CDs. As a result, the enantioselectivity of these two phenothiazines is remarkably and synergistically enhanced with increasing the concentration of neutral CDs in the presence of MI-S-beta-CD and simultaneous enantioseparations of these phenothiazines, except for methotrimeprazine, could favorably be achieved with the use of dual CD systems. Moreover, by varying the concentration of beta-CD or gamma-CD at a fixed concentration of MI-S-beta-CD (0.75% w/v) reversal of the enantiomer migration order of promethazine occurred. This may be attributable to the opposite effects of charged and neutral CDs on the mobility of the enantiomers of promethazine.     

A comparison of phosphated and sulfated beta-cyclodextrins as chiral selectors for capillary electrophoresis

The enantioseparation capabilities of three different functionalized beta-cyclodextrins, two sulfated beta-cyclodextrins with 4 and 15 nominal degrees of substitution and a phosphated beta-cyclodextrin with 8 degrees of substitution, were compared. While anodic detection was used with both sulfated cyclodextrins, the phosphated cyclodextrin required cathodic detection suggesting either lower ionization of the phosphated cyclodextrin or generally lower affinity of the analytes for the phosphated cyclodextrin. The effects of several experimental parameters were evaluated with respect to enantioseparation. The degrees of substitution of the cyclodextrin, pH of the background electrolyte as well as the concentration of the functionalized beta-cyclodextrin, each had a significant influence on the successful enantiomeric separation of the chiral drugs investigated.     

Polyacrylamides as hydrophilic selectors in non-aqueous capillary electrophoresis

Polyacrylamides (PAAms) were investigated as hydrophilic selectors in non-aqueous capillary electrophoresis (CE). Separation of 10 substituted benzoates and unsubstituted benzoate as model samples was greatly improved by the addition of PAAms in acetonitrile-CE. The migration behavior indicates that the carbonyl moiety of PAAms works as a good hydrogen-accepting site toward hydrogen-donating analytes such as 4-hydroxybenzoate anion (4OH-BA) in acetonitrile. PAAms also serve as electron-accepting agents with its amide proton interacting with the dissociated carboxyl groups of the benzoates. The ion-dipole interaction is useful to control the migration behavior of benzoates without hydrogen-donating substituents. The overall mode of the interaction is similar to that of polyethylene glycol (PEG) 20000 reported previously, but the complex formation constant of poly(N-tert.-butyl)acrylamide (PBAAm) with 40H-BA estimated here was 130-fold larger than that of PEG 20000. This would be ascribed to the strong basicity of the carbonyl oxygen atoms of PBAAm as compared with the ether oxygen atoms of PEG. Furthermore, a copolymer of (N-tert.-butyl)acrylamide-acrylamide [70:30 (in feed)] exhibited a complex formation constant of about fourfold larger toward 4OH-BA than PBAAm, most probably due to decrease in steric hindrance from the tert.-butyl groups. Adrenaline and its six precursors have been separated successfully using the PAAms.     

Chiral separation of hydroxyflavanones in cyclodextrin-modified capillary zone electrophoresis using sulfated cyclodextrins as chiral selectors

Chiral separations of three hydroxyflavanone aglycones, including 2'-, 3'-, and 4'-hydroxyflavanone, in capillary zone electrophoresis (CZE) using randomly sulfate-substituted beta-cyclodextrin (S-beta-CD) or dual cyclodextrin (CD) systems consisting of S-beta-CD and a neutral CD at low pH were investigated. The results indicate that S-beta-CD is an excellent chiral selector for enantioseparation of 2'-hydroxyflavanone and is a good chiral selector for 3'-hydroxyflavanone. Depending on the concentration of S-beta-CD ranging from 2.0 to 0.75% (w/v), the enantioresolution values were 10.5-19.5 and 1.8-3.4 for 2'- and 3'-hydroxyflavanone, respectively. The enantiomers of 4'-hydroxyflavanone could be effectively separated with S-beta-CD at a concentration of 2.0% (w/v) within 20 min. The enantioselectivity and enantioresolution follow the order 2'-hydroxyflavanone>3'-hydroxyflavanone>4'-hydroxyflavanone. Alternatively, with the addition of sodium dodecyl sulfate (SDS) monomers at low concentrations in the electrophoretic system, enantioselectivity of these hydroxyflavanone aglycones could be enhanced with dual CD systems. In this case, SDS monomer acted as a complexing agent probably first with S-beta-CD and then subsequently with the analytes for increasing the effective electrophoretic mobility of the analytes towards the anode and as a selectivity controller for affecting the selectivity of hydroxyflavanones. Better enantioseparation between 2'-hydroxyflavanone and 3'-hydroxyflavanone could be achieved with a dual CD system consisting of S-beta-CD and gamma-CD than that with S-beta-CD and beta-CD. In addition, possible chiral recognition mechanisms of hydroxyflavanones are discussed.     

Enantioseparation of novel psychoactive chiral amines and their mixture by capillary electrophoresis using cyclodextrins as chiral selectors

The prevalence of new psychoactive substances (NPS) has been increasing during the last decade as well as their constant growth of availability across the whole world. Regardless of the potential health hazard, NPS (often racemic compounds) are frequently sought after and abused for their psychoactive effects that may differ for individual enantiomers. In this work, capillary electrophoresis was used for the chiral separation of a mixture of eleven psychoactive chiral amines using β-cyclodextrin and carboxymethyl-β-cyclodextrin as chiral selectors at various concentrations. Chiral separation was successful for all the analytes studied. A mixture of these analytes was subsequently analyzed under optimal conditions, i.e., when using 20 mmol/L carboxymethyl-β-cyclodextrin in 50 mmol/L sodium phosphate buffer, pH 2.5. In this case, chiral separation occurred in nine out of eleven analytes. To our best knowledge, we achieved enantioseparations of seven analyzed compounds by CE for the first time.