Evaluation of the enantioselectivity of glycogen-based dual chiral selector systems towards basic drugs in capillary electrophoresis

Several chiral reagents including cyclodextrins (CDs) and derivatives, crown ethers, proteins, chiral surfactants and polymers have been involved in dual selector systems for enantioseparation of a series of chiral compounds by capillary electrophoresis (CE). In comparison to the chiral reagents above-mentioned, there is no report concerning the use of polysaccharides in dual chiral CE system. In this paper we first investigate the enantioselectivity of polysaccharide-based dual selector systems towards some chiral drugs. During our recent work, glycogen belonging to the class of branched polysaccharides has been used as a novel chiral selector in CE. In this study, three glycogen-based dual chiral CE systems have been established for enantiomeric separations of several racemic basic drugs consisting of duloxetine, cetirizine, citalopram, sulconazole, laudanosine, amlodipine, propranolol, atenolol and nefopam. These three dual systems combined glycogen (neutral polysaccharide) with chondroitin sulfate A (CSA, ionic polysaccharide), β-CD and HP-β-CD, respectively. It was found that the dual system of glycogen/CSA exhibited good enantioselective properties toward the tested drugs. More importantly, compared to the single selector systems, synergistic effect was observed when glycogen was used with CSA for most of the analytes. This indicated the enhancement of enantioseparation observed for these analytes in glycogen/CSA system might be due to some favorable interaction effects between glycogen and CSA. Moreover, in order to evaluate the stereoselectivity of glycogen/CSA, the influences of buffer pH and selector concentration on enantioseparation of the studied drugs were also investigated.     

On the use of dispersed nanoparticles modified with single layer beta-cyclodextrin as chiral selecor to enhance enantioseparation of clenbuterol with capillary electrophoresis

A new strategy for chiral separation by capillary electrophoresis employing modified-nanoparticles as chiral selector is described for clenbuterol analysis. Nanoparticles modified with β-cyclodextrin (β-CD) form a large surface area platform to serve as a pseudostationary chiral phase, which can be applied for the enhancement of the enantioseparation. The application of four kinds of nanoparticles was investigated (multi-walled nanotubes (MWNTs), polystyrene (PS), TiO₂ and Al₂O₃) modified with single layer β-CD as chiral selector in the enantioseparation of clenbuterol by capillary electrophoresis (CE). Successful clenbuterol enantioseparation could be achieved with the β-CD-modified MWNTs as chiral selector. X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) confirmed the β-CD modification of the nanoparticles. The effects of nanoparticles, surfactant, chiral selector (β-CD) and run buffer were studied in relation to the enantiomeric separation of clenbuterol. This study opens attractive perspectives for the use of modified nanoparticles for chiral separational purposes in CE.     

Study on enantiomeric separation of basic drugs by NACE in methanol‐based medium using erythromycin lactobionate as a chiral selector

A wide variety of chiral selectors have been employed in CZE, and among them macrocyclic antibiotics including glycopeptides, ansamycins, aminoglycosides and polypeptides exhibited prominent enantioselective properties toward abundant racemic compounds. Compared with CZE, the use of macrocyclic antibiotics as chiral selectors in NACE has not been reported previously. In this study, an approach to the enantioseparation of basic drugs by means of NACE with erythromycin lactobionate (EL) belonging to the group of macrolide antibiotics has been investigated. Especially different from the above four classes of antibiotics, there are no reports concerned with the use of macrolides which belong to macrocyclic antibiotics as chiral selectors in CE. In this work EL is first used as a chiral selector in NACE for the enantiomeric separations of two racemic basic drugs that possess high separability consisting of propranolol and duloxetine. Furthermore, EL possesses advantages such as high solubility and low viscosity in the solvent and very weak UV absorption. The chiral separations were achieved using Tris‐boric acid as the BGE and methanol as the organic medium. In the course of this work we observed that both migration time and enantioseparation were influenced by several parameters such as the pH and composition of the BGE, EL concentration, capillary temperature and applied voltage. Consequently, these parameters were systematically optimized in order to obtain the optimum enantioseparations.     

Evaluation of clarithromycin lactobionate as a novel chiral selector for enantiomeric separation of basic drugs in capillary electrophoresis

Nowadays, macrocyclic antibiotics are presenting an increasing number of enantioseparation applications. The macrocyclic antibiotics used as chiral selectors in capillary electrophoresis (CE) include the ansamycins and the glycopeptides. The macrolides, another important class of macrocyclic antibiotics, have been reported as a new type of chiral selectors recently. In this study, clarithromycin lactobionate (CL), belonging to the group of macrolide antibiotics, was first investigated for its potential as a novel chiral selector in CE for enantiomeric separation of several basic drugs. As observed, CL allowed excellent separation of the enantiomers of metoprolol, atenolol, propranolol, bisoprolol, esmolol, ritodrine, and amlodipine, as well as partial enantioresolution of labetalol and nefopam. In addition, CL possesses advantages such as high solubility and low viscosity in the solvent and very weak UV absorption. In the course of this study, it was found that both migration times and enantioseparation of the basic drugs were influenced by several experimental parameters, e.g. selector concentration, the composition and pH of the BGE, the type and concentration of organic modifier, and applied voltage. Thus, the effects of these factors were systematically investigated, and satisfactory enantioseparations of the studied drugs were achieved at the buffer pH range of 7.3–7.5 using 12.5 mM borax buffer with 50% v/v methanol, 60 mM CL, and 20 kV applied voltage. Moreover, comparison of the influences of the studied parameters was further investigated by means of Statistical Product and Service Solutions (SPSS) in this article.     

Investigation of enantiomeric separation of basic drugs by capillary electrophoresis using clindamycin phosphate as a novel chiral selector

A wide number of chiral selectors have been employed in CE and among them macrocyclic antibiotics exhibited excellent enantioselective properties toward plenteous racemic drugs. Different from macrocyclic antibiotics, the use of lincomycin antibiotics as chiral selectors has not been reported previously. In this study clindamycin phosphate belonging to the group of lincomycin antibiotics is first used as a novel chiral selector for the enantiomeric separations of several racemic basic drugs, which possess high separability, consisting of nefopam, citalopram, tryptophan, chlorphenamine and propranolol. Other basic drugs giving partial enantioseparation include tryptophan methyl ester, metoprolol and atenolol. Clindamycin phosphate possesses advantages such as high solubility and low viscosity in the water and very weak UV absorption. In the course of this work we observed that both migration time and enantioseparation were influenced by several parameters such as pH of the BGE, clindamycin concentration, capillary temperature, applied voltage and organic modifier. The optimum pH that was in the neutral or weak basic region but varied among drugs, a low capillary temperature and a clindamycin concentration of 60 or 80 mM are recommended as the optimum conditions for chiral separation of these drugs. Moreover, comparison of the influences of the studied parameters was further investigated by means of Statistical Product and Service Solutions in this paper.     

Di-n-amyl L-tartrate-boric acid complex chiral selector in situ synthesis and its application in chiral nonaqueous capillary electrophoresis

A chiral selector, di-n-amyl L-tartrate-boric acid complex, was in situ synthesized by the reaction of di-n-amyl L-tartrate with boric acid in a nonaqueous background electrolyte (BGE) using methanol as the medium. And a new method of chiral nonaqueous capillary electrophoresis (NACE) was developed with the complex as the chiral selector. It has been demonstrated that the chiral selector is suitable for the enantioseparation of some β-blockers and β-agonists in NACE. Some chiral analytes that could not be resolved in aqueous microemulsion electrokinetic chromatography (MEEKC) with the same chiral selector obtained baseline resolutions in the NACE system. The enantioseparation mechanism was considered to be ion-pair principle and the nonaqueous system was more favorable for the ion-pair formation which is quite useful for the chiral recognition. The addition of a proper concentration of triethylamine into the BGE to control the apparent pH (pH*) enhanced the enantiomeric discrimination. In order to achieve a good enantioseparation, the effects of di-n-amyl L-tartrate and boric acid concentration, triethylamine concentration, applied voltage, as well as capillary length were investigated. Under the optimum conditions, all of the tested chiral analytes including six β-blockers and five β-agonists were baseline resolved.     

Experimental designs to investigate capillary electrophoresis-electrospray ionization-mass spectrometry enantioseparation with the partial-filling technique

An experimental design approach is described to evaluate the main electrophoretic parameters involved in the enantioseparation of pharmaceuticals by capillary electrophoresis (CE) coupled to electrospray ionization-mass spectrometry (ESI-MS). For all experiments, the partial-filling technique was applied to avoid the chiral selector entering in the mass spectrometer ion source with a negative effect on the electrospray performance. To carry out enantioseparation, a volatile buffer constituted of 20 mM ammonium acetate at pH 4.0, and a polyvinyl alcohol-coated capillary were used. Methadone was employed as the model compound and three different cyclodextrins (CDs), namely sulfobutyl ether-beta-CD, carboxymethylated-beta-CD and hydroxypropyl-beta-CD, were selected in order to study the countercurrent process. Two different experimental designs were chosen: (i) a full-factorial design to examine the effects and significance of the investigated factors, and (ii) a central composite face-centered design to establish the mathematical model of the selected responses in function of experimental factors. The chiral selector concentration, percentage of the capillary filled with the chiral selector, and drying gas nebulization pressure were three relevant factors taken into consideration. For each CD, the methadone enantiomeric resolution, apparent selectivity, and migration time of the second enantiomer were established as responses. The latter were systematically related to experimental parameters with the help of multiple linear regression. It is noteworthy that the behaviour was different in function of the chiral selector charge. Results revealed that the nebulization pressure involved in the electrospray process and the CD concentration had a significant effect on the enantiomeric resolution, while the effect of the separation zone length was less pronounced. Finally, response surfaces were drawn from the mathematical model and experimental conditions were selected to allow a robust determination of methadone enantiomers by CE-MS.     

Fast enantioseparation of arylglycine amides by capillary electrophoresis with highly sulfated-beta-cyclodextrin as a chiral selector

Nine racemic arylglycine amides were synthesized and successfully enantioseparated by capillary electrophoresis (CE) using highly sulfated beta-cyclodextrin (HS-beta-CD) as a chiral selector. Baseline enantioseparation of the analytes was obtained around neutral pH but not in the acidic conditions that are commonly used. HS-beta-CD content, buffer pH, type and concentration, and organic modifier concentration were studied and optimized for fast and efficient separation. A chiral CE separation system composed of 1.5% (w/v) HS-beta-CD, 0 to 10% (v/v) methanol and 20 mM 3-(N-morpholino)propanesulfonic acid at pH 6.5 was shown suitable for baseline enantioseparation of the mentioned amides within 6 min, including simultaneous enantioseparation of three positional isomer series (methyl-, methoxyl or chloro-substituted). By using this system, D-enantiomers migrated ahead of the L-enantiomers and the enantiomeric resolution order of arylglycine amides was more or less parallel to the pK(a), order of the analytes.     

Evaluation of the enantioseparation capability of the novel chiral selector clindamycin phosphate towards basic drugs by micellar electrokinetic chromatography

To date, a series of chiral selectors have been utilized successfully in capillary electrophoresis (CE). Among these various chiral selectors, macrocyclic antibiotics have been demonstrated to represent powerful enantioselectivity towards many chiral compounds. Differing from macrocyclic antibiotics, the use of lincosamide antibiotics as chiral selectors has not been reported previously. In our recent work, clindamycin phosphate belonging to the group of lincosamides has been first used as a chiral selector in capillary zone electrophoresis (CZE). In this paper, a micellar electrokinetic chromatography (MEKC) method has been developed for the evaluation of enantioseparation capability of this novel chiral selector towards several racemic basic drugs. As observed during the course of this work, clindamycin phosphate allowed excellent separation of the enantiomers of nefopam, citalopram, tryptophan, chlorphenamine, propranolol and metoprolol, as well as partial enantioresolution of tryptophan methyl ester and cetirizine. In this MEKC chiral separation system, different types of anionic surfactants, organic additives and background electrolytes were tested, and satisfactory enantioseparations of basic drugs above-mentioned were achieved using sodium dodecyl sulfate (SDS) as the surfactant, isopropanol as the organic additive, and phosphate as the background electrolyte. Furthermore, both migration times and enantioseparation of the analytes were influenced by several experimental parameters such as pH of the BGE, clindamycin phosphate and SDS concentrations, phosphate and isopropanol concentrations, and applied voltage. Consequently, the effects of these factors on enantioseparations of the studied basic drugs were systematically investigated in order to evaluate the stereoselectivity of clindamycin phosphate in MEKC.     

Synthesis and application of clindamycin succinate as a novel chiral selector for capillary electrophoresis

A novel chiral selector, clindamycin succinate, was synthesized and first used as a chiral selector in capillary electrophoresis (CE). The chiral resolution ability of this kind of clindamycin derivation was studied by CE using some racemic drugs as model analytes. From the experimental results, it was found that both resolution and selectivity of the selector were dependent on the following parameters: concentration of chiral selectors, pH of the running buffer, temperature of the capillary column, applied voltage and organic modifier used. The results show that the chiral selector possesses high resolution toward some racemic drugs, including ofloxacin, chlorphenamine, tryptophan, propranolol, sotalol and metoprolol. Excellent chiral resolution of these tested drugs was achieved under the optimal conditions of 50 mM clindamycin succinate, 10% MeOH v/v, 50 mM Tris buffer, pH 4.0, at 22 kV and 20 °C within 25 min.     

Enantiomeric separation of chiral peptide nucleic acid monomers by capillary electrophoresis with charged cyclodextrins

Direct chiral separation of chiral peptide nucleic acid (PNA) monomers has been achieved for the first time by capillary electrophoresis (CE) with charged cyclodextrins as chiral selectors added to the electrophoretic buffer. Selectively modified 6-deoxy-6-N-histamino-beta-cyclodextrin and sulfobutyl ether-beta-CD were successfully used as chiral selectors for the enantiomeric separation of chiral monomers based on different aminoethylamino acids bearing thymine or adenine as nucleobases. Chiral separations were obtained at low selector concentrations (1-3 mM) with good enantioselectivity and resolution factors. Separations were optimized as a function of pH in order to exploit the effect of the electrostatic interactions between the oppositely charged selector and selectand. The method has been applied to the analysis of the enantiomeric excess of chiral monomers used for the solid phase synthesis of chiral PNA oligomers. CE chiral analysis showed that a very high enantiomeric purity was generally achieved in the synthesis of all monomers, except for histidine and aspartic acid based monomers in which ca. 10% of the "wrong" enantiomer was always present.     

Glycogen: A novel branched polysaccharide chiral selector in CE

Various chiral selectors have been employed in CE and among them linear polysaccharides exhibited powerful enantioselective properties. Different from linear polysaccharides, the use of branched polysaccharides as chiral selectors in CE has not been reported previously. In this study glycogen belonging to the class of branched polysaccharides was used as a novel chiral selector for the enantiomeric separations for the first time. Since glycogen is electrically neutral, the method is applicable to ionic compounds. Eighteen chiral compounds including 12 basic drugs and six acidic drugs have been tested to demonstrate the potential of this chiral selector. BGE and selector concentrations and buffer pH were systematically optimized in order to obtain successful chiral separations. Among the tested compounds, the enantiomers of ibuprofen, which is an acidic drug, were successfully recognized by 3.0% w/v glycogen with 90 mM Tris‐H₃PO₄ buffer (pH 7.0). The enantiomers of basic drugs such as citalopram, cetirizine and nefopam were also baseline‐resolved with 50 mM Tris‐H₃PO₄ buffer (pH 3.0) containing 3.0% glycogen. Amlodipine belonging to basic compound only gave partial enantioseparation under the above‐mentioned condition.     

Investigation of the enantioselectivity of tetramethylammonium‐lactobionate chiral ionic liquid based dual selector systems toward basic drugs in capillary electrophoresis

Recently, chiral ionic liquids (CILs) have received increasing interest in chiral separation by CE. Nevertheless, the lack of deep perception of the specific mechanism about CILs in CE for enantioseparation still perplexes a legion of researchers, despite the strenuous efforts. In this paper, a lactobionic acid based ionic liquid, tetramethylammonium‐lactobionate (TMA‐LA) was applied for the first time in CE to establish dual selector system with clindamycin phosphate (CP) for enantiomeric separation. Compared to single CP system or single TMA‐LA system significantly improved separations of seven tested rameric drugs (propranolol, nefopam, citalopram, chlorphenamine, metoprolol, bisoprolol, and esmolol) were observed in the dual selector systems. Several crucial parameters such as type and proportion of organic modifier, buffer composition and pH, and concentration of TMA‐LA/CP were systematically investigated to achieve satisfied enantioseparation. Meanwhile, molecular modeling was applied to demonstrate the chiral recognition mechanism of the TMA‐LA/CP dual‐selector separation system using the molecular docking software Autodock, which well supported the experimental results. The existence of TMA‐LA/CP complex may give rise to a higher discriminatory ability against the enantiomers, indicating the reason of improved separation in TMA‐LA/CP system. All the influence factors evaluated by means of Statistical Product and Service Solutions (SPSS) to research the influences on the chiral separation system.     

Influence of (hydroxy)alkylamino substituents on enantioseparation ability of single-isomer amino-beta-cyclodextrin derivatives in chiral capillary electrophoresis

A family of single-isomer amino-beta-cyclodextrin (amino-beta-CD) derivatives containing an amino or (hydroxy)alkylamino group in one of the primary positions has been synthesized. The steric effect and hydrogen bond forming ability of the different substituents on enantioseparation of acidic enantiomers has been studied by capillary electrophoresis (CE). Three enantiomeric model compounds (mandelic acid, cis-permethrinic acid, and cis-deltamethrinic acid) having significantly different apparent complex stability constants with beta-CD were applied in the experiments. Dependence of separation selectivity, resolution as well as mobility difference on chiral selector concentration (0.1-20 mM, pH 6.0) was investigated. Each amino-beta-CD showed higher enantioselectivity than the native beta-CD. One hydroxyalkyl group attached to the primary amino N-atom significantly increased both the enantioselectivity and the resolution compared to the primary amino-beta-CD, while two hydroxyalkyl moieties decreased them due to the predominance of steric hindrance. The value of the apparent complex stability constants obtained suited well the mobility difference model (by Wren). On the other hand, the optimum selector concentrations calculated according to the model were slightly lower than the experienced concentrations giving the maximum enantioresolution of enantiomers.     

Electrically driven microseparation methods for pesticides and metabolites: IV. Effects of the nature of fluorescent labels on the enantioseparation of pesticides and their degradation products by capillary zone electrophoresis with UV and laser-induced fluorescence detection

Three different fluorescent tags, namely 5-aminonaphthalene-1-sulfonic acid (ANSA), 7-aminonaphthalene-1,3-disulfonic acid (ANDSA), and 8-aminonaphthalene-1,3,6-trisulfonic acid (ANTS) were evaluated in the precolumn derivatization of some chiral phenoxy acid herbicides, some chiral transformation products of pyrethroid insecticides, and in their subsequent enantiomeric separation by capillary electrophoresis (CE). The electrolyte systems consisted of sodium phosphate buffers containing chiral surfactants such as octylglucoside (OG) or octylmaltoside (OM) at concentrations above the critical micellar concentration (CMC). Among the three different tags investigated, the ANDSA derivatives of the various solutes were more readily enantioseparated than the ANSA and ANTS derivatives. While the tagging with ANSA allowed the enantioseparation of a limited number of the chiral solute-ANSA derivatives investigated, the ANTS derivatization yielded derivatives that could not be enantioseparated. The polarity of the three different tags increases by increasing the number of sulfonic acid groups in the molecule in the following order: ANSA (one sulfonic acid) < ANDSA (two sulfonic acid groups) < ANTS (three sulfonic acid groups). This seems to indicate that the intermediate polarity of the ANDSA tag allowed more equitable nonpolar/polar interactions of the ANDSA-derivatized solutes with the OG or OM micelles, and consequently the enantioseparation of the solute-ANDSA derivatives. Thus, there is a solute polarity window for enantioresolution with alkylglycoside micelle by CE. Solutes of intermediate polarity that undergo more equitable nonpolar/polar interactions with the micelles exhibited chiral separations.     

Study of the enantioseparation capability of chiral dual system based on chondroitin sulfate C in CE

It has been reported that chiral dual system is able to improve the enantioseparation of enantiomers in many cases. Currently, the dual systems involved in CE chiral separation are mostly dual CDs systems, and the polysaccharides‐based chiral dual system was reported in only one paper. To the best of our knowledge, the use of chondroitin sulfate C (CSC)‐based dual system for enantiomeric separation has not been reported previously. Herein, four CSC‐based chiral dual systems, namely CSC/glycogen, CSC/chondroitin sulfate A (CSA), CSC/hydroxypropyl‐β‐CD (HP‐β‐CD), as well as CSC/β‐CD (β‐CD), were evaluated for the first time for their enantioseparation capability by CE in this paper. During the course of the work, the influences of chiral selector concentration and buffer pH values on enantioseparation in dual systems were systematically investigated. Under the optimized conditions, the dual system consisting of CSC and glycogen exhibited better separations toward nefopam, duloxetine, sulconazole, atenolol, laudanosine, and cetirizine enantiomers compared to the single CSC or glycogen system. The combination of CSC and HP‐β‐CD improved the separation of amlodipine and chlorphenamine enantiomers. However, no synergistic effect was observed in the CSC/CSA and CSC/β‐CD systems.     

Evaluation of the synergistic effect with amino acids for enantioseparation of basic drugs using capillary electrophoresis

The synergistic effect of two acidic amino acids, aspartic and glutamic acid, on the electrophoretic enantioseparation of four basic drugs was evaluated in the BGE containing a CD and at different pHs. Chlorpheniramine, hydroxyzine, propranolol and tramadol were used as the basic model drugs. However, no enantioseparations were achieved with a BGE containing sole amino acid, but the combined use of an acidic amino acid and a CD showed improved enantioseparations (synergistic effect) compared with the single CD system. The results demonstrated that at optimized pH, the electrostatic interactions of the anionic amino acids with the positively charged basic drugs could result in a decrease of the analyte migration velocity and it consequently improved the enantioseparation. The effective parameters such as the amino acid and chiral selector type and concentration, buffer pH, applied voltage, and capillary temperature were optimized. Favorable enantiomeric resolution and migration times of the model drugs were achieved with a 100 mM phosphate buffer solution (pH 3.0) containing 5.0 mM HP‐α‐CD/HP‐β‐CD and 20 mM aspartic acid with an 18 kV applied voltage at 25°C. ¹H NMR experiments were also carried out in a mixture of an analyte and CD in the absence and presence of aspartic acid. The NMR results were consistent with the results obtained by CE which showed the synergistic effect of amino acid.     

Evaluation of the enantioselectivity of carbon nanoparticles‐modified chiral separation systems using dextrin as chiral selector by capillary electrokinetic chromatography

Nanoparticles (NPs) can be used as pseudostationary phases (PSPs) in EKC, which is similar to the use of micelle additives as applied in MEKC. To date, the use of NPs to enhance enantiomeric separation by EKC with β‐CD or its derivative as chiral selector has been reported only in two papers. However, to the best of our knowledge, there has been no prior effort to use NPs for achieving enantioseparation with polysaccharides as chiral selector. This paper describes for the first time the use of carbon nanoparticles (CNPs) as PSPs to modify chiral separation system employing dextrin as chiral selector for the enantioseparations of several basic drugs in capillary EKC. Three different types of CNPs, including carbogenic nanoparticles (NPs), carboxylated single‐walled carbon nanotubes, and carboxylated multiwalled carbon nanotubes, were used as running buffer additives, respectively. The potential of the PSPs and the effects of dextrin concentration, buffer pH, and buffer concentration on the enantioseparations were evaluated. Four pairs of tested enantiomers were successfully resolved in less than 15 min with the resolution values in the range of 1.41–4.52 under optimized conditions. Compared to the buffer without NPs, the introduction of NPs into the buffer enhanced the separation of the enantiomers.     

Enantiorecognition of profens by capillary electrophoresis using a novel chiral selector eremomycin

The evaluation of a macrocyclic glycopeptide antibiotic, eremomycin, as a chiral selector in capillary electrophoresis (CE) has been performed. The stability of eremomycin in solution and capillary electrolyte, as well as its optical and electrophoretic properties have been discussed. The effect of experimental parameters influencing the enantioseparation of several profens has been studied. Excellent enantioseparation of profens has been achieved and migration order has been validated. Comparison of enantioseparations of profens in CE by using eremomycin-mediated electrolytes and in HPLC with eremomycin immobilized on silica has revealed similar trends for both methods.     

Avoparcin, a new macrocyclic antibiotic chiral run buffer additive for capillary electrophoresis.

Avoparcin, like vancomycin, teicoplanin, and ristocetin A, belongs to the family of macrocyclic glycopeptide antibiotics. These antibiotics have all been used as effective chiral selectors for capillary electrophoresis (CE), thin-layer chromatography (TLC), and high performance liquid chromatography (HPLC). The present work focuses on avoparcin, which has been shown to be an excellent chiral selector for the CE enantioseparation of many N-blocked amino acids, as well as several anti-inflammatory drugs of pharmaceutical importance. The use of avoparcin as a chiral run buffer additive in CE is discussed, as well as the effects of changing experimental parameters, like avoparcin concentration, pH, organic modifiers, etc. Comparisons of enantioseparations of some N-3,5-dinitrobenzoyl-derivatized amino acids, using either avoparcin, ristocetin A, teicoplanin, or vancomycin in the run buffer, are also made. In general, vancomycin had the longest migration times, and ristocetin A the shortest, while avoparcin was intermediate. Generally, at least one of the four chiral selectors produced an excellent separation, while a different macrocyclic antibiotic produced a poor separation. Currently, we see no way to predict which chiral run buffer additive will be best or worst for an individual solute.