Cyclodextrin-modified Micellar Electrokinetic Chromatography Method Research Articles

Analytical Quality by Design-Compliant Development of a Cyclodextrin-Modified Micellar ElectroKinetic Chromatography Method for the Determination of Trimecaine and Its Impurities.

In 2022, the International Council for Harmonisation released draft guidelines Q2(R2) and Q14, intending to specify the development and validation activities that should be carried out during the lifespan of an analytical technique addressed to assess the quality of medicinal products. In the present study, these recommendations were implemented in Capillary Electrophoresis method development for the quality control of a drug product containing trimecaine, by applying Analytical Quality by Design. According to the Analytical Target Profile, the procedure should be able to simultaneously quantify trimecaine and its four impurities, with specified analytical performances. The selected operative mode was Micellar ElectroKinetic Chromatography employing sodium dodecyl sulfate micelles supplemented with dimethyl-β-cyclodextrin, in a phosphate-borate buffer. The Knowledge Space was investigated through a screening matrix encompassing the composition of the background electrolyte and the instrumental settings. The Critical Method Attributes were identified as analysis time, efficiency, and critical resolution values. Response Surface Methodology and Monte Carlo Simulations allowed the definition of the Method Operable Design Region: 21-26 mM phosphate-borate buffer pH 9.50-9.77; 65.0 mM sodium dodecyl sulfate; 0.25-1.29% v/v n-butanol; 21-26 mM dimethyl-β-cyclodextrin; temperature, 22 °C; voltage, 23-29 kV. The method was validated and applied to ampoules drug products.

Combining excitation-emission matrix fluorescence spectroscopy, parallel factor analysis, cyclodextrin-modified micellar electrokinetic chromatography and partial least squares class-modelling for green tea characterization

In this study, an alternative analytical approach for analyzing and characterizing green tea (GT) samples is proposed, based on the combination of excitation–emission matrix (EEM) fluorescence spectroscopy and multivariate chemometric techniques. The three-dimensional spectra of 63 GT samples were recorded using a Perkin–Elmer LS55 luminescence spectrometer; emission spectra were recorded between 295 and 800 nm at excitation wavelength ranging from 200 to 290 nm, with excitation and emission slits both set at 10 nm. The excitation and emission profiles of two factors were obtained using Parallel Factor Analysis (PARAFAC) as a 3-way decomposition method. In this way, for the first time, the spectra of two main fluorophores in green teas have been found. Moreover, a cyclodextrin-modified micellar electrokinetic chromatography method was employed to quantify the most represented catechins and methylxanthines in a subset of 24 GT samples in order to obtain complementary information on the geographical origin of tea. The discrimination ability between the two types of tea has been shown by a Partial Least Squares Class-Modelling performed on the electrokinetic chromatography data, being the sensitivity and specificity of the class model built for the Japanese GT samples 98.70% and 98.68%, respectively. This comprehensive work demonstrates the capability of the combination of EEM fluorescence spectroscopy and PARAFAC model for characterizing, differentiating and analyzing GT samples.

Enantioseparation of enilconazole using cyclodextrin-modified MEKC

Enantiomeric separation of enilconazole has been successfully achieved using cyclodextrin modified micellar electrokinetic chromatography (CD-MEKC) method with HP-γ- CD as chiral selector. Two parameters which were HP-γ-CD concentration and SDS concentration have been optimized by investigating their effects on resolution and analysis time. Enantioresolution of enilconazole was achieved by the optimized CD-MEKC system containing 40 mM HP-γ- CD, 40 mM SDS and 20 mM phosphate buffer (pH 8) with resolution (Rs = 1.3) and analysis time within 9 min. Good repeatabilities were obtained in term of RSD% (n = 3) for migration time, peak area and peak height ranging from 0.07 to 6.1 %. The optimized CD-MEKC method was a simple, less solvent used, and able to perform enantiomeric separation of enilconazole with a good resolution and relatively short analysis time.

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, Rs 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) (r(2) > 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%.

Chemometrics-guided development of a cyclodextrin-modified micellar electrokinetic chromatography method with head-column field amplified sample stacking for the analysis of 5-lipoxygenase metabolites

A new sensitive method using α-cyclodextrin-modified micellar electrokinetic chromatography has been developed to separate and quantify arachidonic acid metabolites of the lipoxygenase pathways in human polymorphonuclear leukocytes, i.e. leukotriene B4, 6-trans-leukotriene B4, 6-trans-12-epi-leukotriene B4, 5(S)-hydroxy-6-trans-8,11,14-cis-eicosatetraenoic acid, 12(S)-hydroxy-6-trans-8,11,14-cis-eicosatetraenoic acid, and 15(S)-hydroxy-6-trans-8,11,14-cis-eicosatetraenoic acid. The electrophoresis system was optimized with regard to the pH, boric acid, SDS and α-cyclodextrin concentration as well as separation voltage and temperature using a three level resolution IV fractional factorial design and a five level circumscribed central composite design. The resulting optimized conditions included 80mM sodium borate buffer, pH 10.07, containing 16.6mM sodium dodecyl sulfate, and 15mM α-cyclodextrin, using a separation voltage of 12.5kV at 23°C. Sensitivity was enhanced employing head-column field amplified sample stacking which resulted in limits of quantification between 30 and 50ng/mL and limits of detection between 10 and 17ng/mL after solid phase extraction of the lipoxygenase products. The method was validated according to the recommendations of the International Conference on Harmonization and applied to the determination of the lipoxygenase metabolites in polymorphonuclear leukocytes upon stimulation with Ca2+-ionophore A23187 and arachidonic acid. Robustness was confirmed using a three level resolution IV fractional factorial design. The novel method is suitable for the analysis of various arachidonic acid metabolites produced by cells and may be used for evaluation of lipoxygenase inhibitors.

Chiral separation of econazole using micellar electrokinetic chromatography with hydroxypropyl-γ-cyclodextrin

A cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) method with hydroxypropyl-γ-cyclodextrin (HP-γ-CD) as chiral selector for the enantiomeric separation of econazole is reported. Enantioseparation of econazole was successfully achieved by the optimized CD-MEKC system containing 40 mM HP-γ-CD, 50 mM SDS and 20 mM phosphate buffer (pH 8) solution with an analysis time of less than 9 min. Calibration curves were linear for the two stereoisomers of econazole ( r 2 > 0.998). Good repeatabilities in the migration time, peak area and peak height were obtained in terms of RSD% ranging from 0.30 to 7.67%. Combination of solid-phase extraction (SPE) procedure using diol column and the CD-MEKC method was successfully applied to the determination of econazole in a formulated cream sample.

Separation of conjugated linoleic acid isomers by cyclodextrin-modified micellar electrokinetic chromatography

A cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) method was developed for separating conjugated linoleic acid (CLA) isomers. All the seven CLA isomers (9 cis,11 cis-CLA, 9 cis,11 trans-CLA, 9 trans,11 trans-CLA, 10 trans,12 cis-CLA, 11 cis,13 cis-CLA, 11 cis,13 trans-CLA and 11 trans,13 trans-CLA) were completely separated in the optimized conditions (4% (w/v) β-cyclodextrin (β-CD), 54 mM sodium dodecyl sulphate (SDS), 80 mM borate (pH 9.0), 8 M urea, 4% (v/v) ethanol, 30 kV and 15 °C). The CD-MEKC method was superior to the gas chromatographic (GC) and silver-ion high-performance liquid chromatographic (Ag +-HPLC) methods that were generally used in analyzing CLA isomers.

High-resolution method for regulatory control of Echinacea species in Nutraceuticals by CD-MEKC

One problem in the international regulatory control of Echinacea, a therapeutic Nutraceutical, is recognition of caffeoyl solutes and alkamides in different products. Cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) has been applied to Echinacea spp. in combination with pattern recognition of some caffeoyl solutes. A novel metric based on relative migration time (RMT) data has been developed in CE to address the problem of variable reported migration time. The CD-MEKC method of Gotti's group using hydroxypropyl-β-cyclodexrin (HP-β-CD; 100 mM) with sodium dodecyl sulphate (SDS; 110 mM), in a triacid background electrolyte (10 mM, pH 8) under 19 kV was adapted to identify two key hydrophilic solutes: chlorogenic acid and cichoric acid present in all commercial products. Two internal markers were taken as reference points to calculate the RMT of any target peak: RMT = t m (target)/ t m (marker). The RMT method was robust to temperature change from 20 to 40 °C, but sensitive to pH. The lateral shift and reproducibility of the target peak t m (target) were significantly reduced by this novel transformation. In the worst cases migration time variability ranged up to 12% ( n = 6); the RMT algorithm reduced this to less than 1%. In general, the RMT transformation reduced the variability of migration time data by a factor of 2–5. For systematic comparison of electrophoretic profiles for test sample and standard, a new pattern recognition algorithm permits sequential peak-by-peak comparison using specified segments of the electropherograms for comparison of test and Echinacea purpurea (root product) as a standard. This algorithm was capable of rapidly characterising the similarity of target peaks in a test sample relative to those in the reference standard. Combination of the RMT algorithm and pattern recognition in CE is expected to offer a robust approach to international regulatory characterisation and control of Nutraceuticals.

Cyclodextrin-modified micellar electrokinetic chromatography for the analysis of Esterom, a topical product consisting of hydrolyzed benzoylecgonine in propylene glycol.

Esterom, a new drug currently in human clinical trials, is a mixture of compounds in a propylene glycol vehicle. It is being evaluated as a topical treatment to aid in the relief of muscle pain and to increase range of motion. Benzoylecgonine is the major component of Esterom and there are at least nine other minor constituents, including four hydroxypropyl esters that have multiple diasteriomers. The aim of the study was to develop a capillary electrophoresis method for the simultaneous separation of the main components in Esterom, including the multiple proposed diastereomers of the esters. Due to the complex sample composition, the use of micelles and cyclodextrins as buffer modifiers was evaluated. A cyclodextrin-modified micellar electrokinetic chromatography method was able to determine 7 of the 8 UV-active Esterom components, with baseline separation of 7 of the 10 diastereomers of the hydroxypropyl esters.

Determination of cinnamaldehyde, cinnamic acid, paeoniflorin, glycyrrhizin and [6]-gingerol in the traditional Chinese medicinal preparation Kuei-chih-tang by cyclodextrin-modified micellar electrokinetic chromatography

This study presents a cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) technique with a multi-wavelength detection method to determine specific components in Kuei-chih-tang. Kuei-chih-tang, a Chinese medicinal preparation, is composed of five crude herbs, i.e., Cinnamomi ramulus, Paeoniae radix, Glycyrrhizae radix, Zingiberis rhizoma and Zizyphi fructus. The effects of SDS and γ-cyclodextrin on the analytes' migration behavior were examined. The selected components were successfully separated within 10 min using a pH 10.0 borax-NaOH buffer containing 20m M SDS and 2m M γ-cyclodextrin. The correlation coefficients of the linear calibration graphs for the analytes exceeded 0.993. Moreover, the optimized CD-MEKC method was employed to analyze four different Kuei-chih-tang samples.