Micellar Electrokinetic Chromatography System Research Articles

Sensitivity Enhancement for Direct Injection Capillary Electrophoresis to Determine Morphine in Human Serum via In-capillary Derivatization.

Rapid and simple micellar electrokinetic chromatography (MEKC) with in-capillary derivatization and fluorescence detection has been developed to determine morphine in human serum. The sample was introduced into a background electrolyte (BGE) containing potassium ferricyanide, whereas morphine was oxidized into highly fluorescent product, pseudomorphine. Different parameters for derivatization and subsequent separation were systematically investigated for the analysis of morphine in serum. Efficient performance of the developed MEKC system was carried out in a single run using BGE made up of 70 mM sodium tetraborate decahydrate (pH 10.5), 0.30 mM potassium ferrricyanide, 80 mM sodium dodecyl sulfate, and applied voltage of 9 kV. The combination of MEKC with in-capillary derivatization of morphine was successfully achieved with a high degree of sensitivity. The validation of the method showed good linearity between areas of morphine and the corresponding concentrations over the range of 5-5000 ng/mL. Excellent accuracy and precision were obtained at all concentration levels. The mean recoveries of morphine were ranging from 83.86 to 94.45%. The validated MEKC method successfully permitted determination of morphine in clinical samples after a single oral dose of controlled release morphine sulfate tablets.

Moment Analysis of Mass Transfer Kinetics in Micellar Electrokinetic Chromatography Systems.

Moment equations were developed for quantitatively studying the separation characteristics of micellar electrokinetic chromatography (MEKC). They explain how the first absolute and second central moments of elution peaks are correlated with some fundamental parameters of the partition equilibrium and mass transfer kinetics in MEKC systems. In order to discuss the influence of the mass transfer kinetics on peak broadening, the moment equations were used to analyze the separation behavior in MEKC systems. Separation conditions were chosen on the basis of practical MEKC experiments previously conducted. It was quantitatively clarified that both the solute permeation at the interfacial boundary of surfactant micelles and axial diffusion of solute molecules in a capillary had a predominant contribution to the spreading of the elution peaks in MEKC systems. This is a preliminary study for the analytical determination of rate constants concerning solute permeation at the interface of surfactant micelles from elution peak profiles measured by MEKC. In addition, it was also indicated that the experimental conditions of MEKC systems could be controlled so that the interfacial solute permeation would have a predominant role for the band broadening. For example, the contribution of the interfacial permeation was about 33 times larger than that of the axial diffusion of solute molecules under the MEKC conditions in a previous study. This means that the rate constants could appropriately be determined for the interfacial solute permeation.

Evaluation of the suitability of chromatographic systems to predict human skin permeation of neutral compounds

Several chromatographic systems (three systems of high-performance liquid chromatography and two micellar electrokinetic chromatography systems) besides the reference octanol–water partition system are evaluated by a systematic procedure previously proposed in order to know their ability to model human skin permeation. The precision achieved when skin-water permeability coefficients are correlated against chromatographic retention factors is predicted within the framework of the solvation parameter model. It consists in estimating the contribution of error due to the biological and chromatographic data, as well as the error coming from the dissimilarity between the human skin permeation and the chromatographic systems. Both predictions and experimental tests show that all correlations are greatly affected by the considerable uncertainty of the skin permeability data and the error associated to the dissimilarity between the systems. Correlations with much better predictive abilities are achieved when the volume of the solute is used as additional variable, which illustrates the main roles of both lipophilicity and size of the solute to penetrate through the skin. In this way, the considered systems are able to give precise estimations of human skin permeability coefficients. In particular, the HPLC systems with common C18 columns provide the best performances in emulating the permeation of neutral compounds from aqueous solution through the human skin. As a result, a methodology based on easy, fast, and economical HPLC measurements in a common C18 column has been developed. After a validation based on training and test sets, the method has been applied with good results to the estimation of skin permeation of several hormones and pesticides.

Performance of chromatographic systems to model soil–water sorption

A systematic approach for evaluating the goodness of chromatographic systems to model the sorption of neutral organic compounds by soil from water is presented in this work. It is based on the examination of the three sources of error that determine the overall variance obtained when soil–water partition coefficients are correlated against chromatographic retention factors: the variance of the soil–water sorption data, the variance of the chromatographic data, and the variance attributed to the dissimilarity between the two systems. These contributions of variance are easily predicted through the characterization of the systems by the solvation parameter model. According to this method, several chromatographic systems besides the reference octanol–water partition system have been selected to test their performance in the emulation of soil–water sorption. The results from the experimental correlations agree with the predicted variances. The high-performance liquid chromatography system based on an immobilized artificial membrane and the micellar electrokinetic chromatography systems of sodium dodecylsulfate and sodium taurocholate provide the most precise correlation models. They have shown to predict well soil–water sorption coefficients of several tested herbicides. Octanol–water partitions and high-performance liquid chromatography measurements using C18 columns are less suited for the estimation of soil–water partition coefficients.

Micellar electrokinetic chromatography systems for the separation of mixtures of charged and uncharged compounds

In this study, the migration behavior of charged and uncharged analytes was investigated under different conditions. Effective mobilities - electrophoretic mobilities under the influence of micelles - of cations, anions, and neutrals were measured at neutral, basic, and acidic pH (7.5, 11, and 2.2) using background electrolytes containing different sodium dodecyl sulfate (SDS) concentrations (0-90 mM) and acetonitrile (ACN) proportions (0-75%). SDS concentration and ACN proportion were found to have a tremendous effect on the effective mobilities and migration order of the model compounds. Although the SDS micelles preferably interact with neutrals and cations, hydrophobic bonds can also occur with anions. Cations, anions, and neutrals having rather different migration behaviors, it is possible to considerably enhance the selectivity of the method by adjusting properly the SDS concentration and the ACN proportion. These observations confirm the interest of using micellar electrokinetic chromatography not only for the separation of neutral substances but also to analyze charged compounds.

Modeling Nonspecific Toxicity of Organic Compounds to the Fathead Minnow Fish by Means of Chromatographic Systems

The performance of chromatographic systems to mimic aquatic toxicity to the fathead minnow fish is evaluated taking into account the factors that contribute to the variance of biological-chromatographic correlations. These factors are the precision to measure the fathead minnow toxicity, the precision of the surrogate chromatographic system, and the error from the dissimilarity between the fathead minnow and chromatographic systems. The precisions are estimated through the characterization of the systems by the solvation parameter model. Several chromatographic systems as well as the common reference octanol-water partition system have been selected to test their ability to model the nonspecific toxicity to the fathead minnow by means of the proposed approach. Predictions and experimental tests show that the micellar electrokinetic chromatography system of sodium taurocholate and chromatographic measurements using an immobilized artificial membrane column provide the most precise estimations of this biopartitioning property. The octanol-water partition system, the conventional C18 high-performance liquid chromatography systems, and the micellar electrokinetic chromatography system of sodium dodecylsulfate show worse performances.

Estimation of Biological Properties by Means of Chromatographic Systems: Evaluation of the Factors That Contribute to the Variance of Biological−Chromatographic Correlations

The performance of chromatographic systems to emulate biological systems is evaluated in terms of the precision that can be achieved. The variance obtained when biological parameters are correlated against physicochemical ones can be decomposed in three terms: the variance of the biological data, the variance of the physicochemical data, and the variance caused by the dissimilarity between the two correlated systems (biological and physicochemical). The three terms contribute to the overall variance observed when measurements in chromatographic systems are correlated with experimental biological properties. The Abraham linear free energy relationships (LFERs) provide a very good approach to characterize biological and physicochemical systems and thus the variance of the analyzed data and the similarity/dissimilarity between them. The contribution of the three variances to the precision of the biological parameter estimated in this way is evaluated from the characterization of the biological and chromatographic systems by means of the Abraham model. The proposed method is able to estimate the goodness of chromatographic systems to predict particular biological properties. In particular, this method is illustrated by comparison of toxicity data (-log LC(50)) for the fish fathead minnow with retention data (log k) in several micellar electrokinetic chromatography (MEKC) systems and also by correlations between retention data (log k) in the sodium taurocholate (STC) MEKC system and data of several biological systems.

Using micellar electrokinetic chromatography for the highly efficient preconcentration and separation of gold nanoparticles

In this paper, we report the use of micellar electrokinetic chromatography (MEKC) for the highly efficient preconcentration and separation of gold nanoparticles (Au NPs). We used the reversed electrode polarity stacking mode (REPSM) of the MEKC system for the on-line enhancement and separation of the Au NPs. Several parameters had dramatic effects on the systems’ performance, including the concentration of sodium dodecylsulfate (SDS) surfactant, the presence of salts in the NP solution, the pH of the running electrolyte, and the temperature of the capillary. Under the optimized conditions [buffer: SDS (70 mM) and 3-cyclohexylamino-1-propanesulfonic acid (CAPS; 10 mM) at pH 10.0; applied voltage: 20 kV; operating temperature: 25 °C; additive: sodium dihydrogenphosphate (NaH 2PO 4, 10 mM); REPSM strategy for sample preconcentration], the number of theoretical plates for the 5.3- and 40.1-nm-diameter Au NPs were 3000 and (an ultrahigh) 2.1 × 10 6, respectively; in addition, the detection sensitivities toward the Au NPs were enhanced ca. 20- and 380-fold, respectively, relative to those obtained using standard MEKC analysis conditions. Furthermore, monitoring the electropherograms using diode-array detection allowed us to identify and characterize the sizes of the separated NPs from their UV–vis spectra. Our findings suggest that MEKC is a highly efficient tool for both the preconcentration and separation of NPs.

Capillary electrokinetic chromatography of insulin and related synthetic analogues

With the implementation of recombinant DNA technology in the pharmaceutical industry, some synthetic insulins have been developed in order to improve the therapy of diabetes. These analogues differ only slightly in the amino acid sequence, therefore displaying a great challenge for analytical chemistry. Within the work presented in this paper, capillary zone electrophoresis (CZE), micellar electrokinetic chromatography (MEKC) with sodium dodecylsulphate (SDS) as micelle-forming agent, and microemulsion electrokinetic chromatography (MEEKC) with microemulsions consisting of SDS, n-octane and 1-butanol were investigated for the separation of human insulin and five synthetic analogues. Best results were achieved with a solvent-modified MEKC system consisting of 100 mM sodium dodecyl sulphate and 15% acetonitrile in 10 mM borate buffer (pH 9.2). A similar system based on perfluorooctanoic acid as micelle-forming agent in ammonium acetate (pH 9.2) was successfully employed for the hyphenation with a quadrupole/time-of-flight mass spectrometer via a sheath-flow interface. In this case, detection limits at 10 mg/L could be achieved.

Rapid Estimation of Octanol–Water Partition Coefficient for Triazole Fungicides by MEKC with Sodium Deoxycholate as Surfactant

A rapid estimation of octanol–water partition coefficient (log Pow) was developed for triazole fungicides by micellar electrokinetic chromatography (MEKC). Five standard compounds with known log Pow values from 2.9 to 4.3 (cyproconazole, bromuconazole, epoxiconazole, bitertanol and difenoconazole) were used for constructing the calibration curve of the log Pow against the MEKC retention factor, log k. A linear relationship was achieved between log Pow and log k, in the MEKC system containing 40 mM sodium deoxycholate (SDC) and 4 mM borate buffer at pH 9.3, with a correlation of determination, r2 = 0.9905. The log Pow values of four test compounds of triazole fungicides (triadimenol, myclobutanil, propiconazole and penconazole) were calculated based on the log k values measured by MEKC and the slope and intercept of the calibration curve. This MEKC method can give good estimation of the log Pow of the four test compounds of triazole fungicides with the differences between the literature log Pow values and estimated log Pow from the MEKC method were from 0.15 to 0.23 log units.

Critical micelle concentration of surfactants in aqueous buffered and unbuffered systems

Critical micelle concentration (CMC) of sodium dodecyl sulphate (SDS), lithium perfluorooctanesulfonate (LPFOS), hexadecyltrimethylammonium bromide (HTAB), tetradecyltrimethylammonium bromide (TTAB), and sodium cholate (SC), surfactants commonly used as pseudostationary phases in micellar electrokinetic chromatography (MEKC), have been determined by means of three different methods: MEKC, spectrophotometry, and conductometry. Determinations have been performed in water, and also in different concentrations of phosphate buffer at pH 7.0. CMC values ranging from 8.08 (water) to 1.99 (50 mM phosphate buffer) mM for SDS, from 7.16 (water) to 2,81 (30 mM phosphate buffer) mM for LPFOS, from 3.77 (water) to 1.93 (20 mM phosphate buffer) mM for TTAB, from 0.91 (water) to ∼0.34 (20 mM phosphate buffer) for HTAB, and around 13 mM (20 mM phosphate buffer) for SC, are obtained. The effect of the electrolyte concentration on the CMC, as well as the linear relationship between the electrolyte counter-ion concentration and the CMC are discussed. This linear relationship provides an easy way for users to estimate the CMC of a MEKC system, at a given electrolyte concentration. A comparison between experimental methods, as well as a discussion about the suitability of a given method for the determination of the CMC for a given surfactant system is also provided.

Comparison of microemulsion electrokinetic chromatography and micellar electrokinetic chromatography methods for the analysis of phenolic compounds

In this study, microemulsion electrokinetic chromatography (MEEKC) and micellar electrokinetic chromatography (MEKC) were compared for their abilities to separate and detect thirteen phenolic compounds (syringic acid, p-coumaric acid, vanillic acid, caffeic acid, gallic acid, 3,4-dihydroxybenzoic acid, 4-hydroxybenzoic acid, (+)-catechin, (-)-epigallocatechin, (-)-epicatechin gallate, (-)-epigallocatechin gallate, (-)-epicatechin, and (-)-gallocatechin), and two other ingredients (caffeine and theophylline) in teas and grapes. Separation of phenolic compounds was improved by changing the SDS concentration for MEEKC, but the SDS concentration rarely affected the resolution for MEKC. Organic modifier (acetonitrile or methanol) was found to markedly influence the resolution and selectivity for both MEEKC and MEKC systems. In addition, a higher voltage and a higher column temperature improved the separation efficiency without any noticeable reduction in resolution for MEEKC whereas they caused a poor resolution for the MEKC system. Although separations with baseline resolution were achieved by the optimized MEEKC and MEKC methods, the separation selectivity resulting from the proposed MEEKC method was completely different from that of MEKC.

Micellar electrokinetic chromatography with bile salts for predicting ecotoxicity of aromatic compounds

The retention factors of several aromatic compounds were obtained by micellar electrokinetic chromatography (MEKC) using cholate, taurocholate, deoxycholate and deoxytaurocholate as micellar systems. The possibility of using these retention factors to describe and predict several ecotoxicological activities of different aromatic compounds was evaluated. Adequate correlations retention–ecotoxicity (log LC 50 in fish and daphnia, log EC 50 in green algae and daphnia, chronic values in fish and green algae, bioconcentration factor, and soil sorption coefficient) were obtained for the micellar systems studied. The predictive ability of the models obtained for these micellar systems was compared. Predicted values concur with the experimental log LC 50 in Bluegill, Rainbow trout, Fathead minnows and Daphnia Magna values for the compounds studied. The results obtained indicated the usefulness of the MEKC systems investigated for the rapid ecotoxicity assessment of aromatic compounds.

Direct coupling of micellar electrokinetic chromatography to mass spectrometry using a volatile buffer system based on perfluorooctanoic acid and ammonia.

Direct coupling of micellar electrokinetic chromatography (MEKC) to mass spectrometry (MS) without employing partial filling is considered to be a challenge. One way of solving the problem would be the use of an MS-compatible surfactant. In the present study, the applicability of a series of surfactants (sodium dodecyl sulfate (SDS), lauric acid, cholic acid and perfluorated carboxylic acids) have been investigated both in terms of separation performance and MS compatibility. It was found that a MEKC system based on perfluorooctanoic acid (PFOA) and ammonia gave excellent results. The separation performance of the suggested system is comparable to the one obtained with standard systems based on SDS and sodium borate buffer although the selectivity is different. The electrospray ionization MS signal of the analytes is not seriously suppressed even at a PFOA concentration of 100 mM. Clusters are formed but their intensities are relatively low and comparable to those obtained with acetic acid. PFOA is volatile enough to allow long-term use, 30 h of continuous use has been recorded without any signs of decreasing performance. After use residual PFOA is easily removed from the ion-source (no memory effects). Furthermore, quantitation of trace impurities is possible at 25 ppb level when employing selected ion monitoring.

On-line micellar electrokinetic chromatography–mass spectrometry: feasibility of direct introduction of non-volatile buffer and surfactant into the electrospray interface

An on-line method for the coupling of micellar electrokinetic chromatography (MEKC) and mass spectrometry (MS) is presented which allows conventional MEKC conditions to be employed without further modification. The MEKC system is coupled directly to electrospray ionization (ESI) MS using a triaxial interface. A systematic study of the influence of the surfactant concentration, the nature and concentration of buffer salts and presence of organic modifier on the interface performance indicated the feasibility of the MEKC–MS approach. Effective interfacing of MEKC was achieved with both single quadrupole and ion-trap MS instruments. Using a background electrolyte containing 20 m M sodium dodecyl sulfate (SDS) and 10 m M sodium phosphate buffer, it is demonstrated that full MEKC runs of test mixtures of mebeverine and related compounds can be monitored by ESI-MS with satisfactory sensitivity. Sub-μg/ml levels of the analytes can still be detected in full scan mode, while detection limits are in the 10–50 ng/ml range when selected ion monitoring is applied. It is shown that such sensitivity would allow full-scan MS detection of 0.1% (w/w) levels of potential impurities in mebeverine. With the ion-trap instrument successful MEKC–MS/MS experiments were carried out providing information-rich MS spectra of the related compounds. Repeated MEKC–MS analyses proved that in the course of 1 day the migration time of mebeverine remained fairly constant while the MS-signal intensity only gradually decreased to approximately 65% of its original value. Once-a-day cleaning of the first part of the ion source, which takes only 5 min, suffices to preserve an optimal interface performance for a prolonged period of time.

Characterization of polyethylene glycols and polypropylene glycols by capillary zone electrophoresis and micellar electrokinetic chromatography

Methods based on capillary zone electrophoresis (CZE) and micellar electrokinetic chromatography (MEKC) have been developed and optimized for the separation of polyethylene glycols (PEGs) and polypropylene glycols (PPGs). To provide for charge and detectability both types of polymeric compounds were derivatized with phthalic anhydride (PhAH) or 1,2,4-benzenetricarboxylic anhydride (BTA) before the separation. Derivatization with BTA yielded more complex electropherograms, due to the occurrence of different isomeric reaction products for every PEG or PPG species. Effective mobilities of the PhAH derivatives were related to the number of monomer units in the polymers in a straightforward way. The CZE method could also be used to determine the monomer-number distribution of random and block PEG–PPG copolymers. For an MEKC analysis the PEGs and PPGs were derivatized with phenyl isocyanate. Oligomers of PEGs could be separated up to molecular masses of 5000, while for the more hydrophobic PPGs oligomeric separation was only accomplished for molecular masses of up to 1500. Due to a strongly different separation mechanism for the PEG and PPG derivatives in the MEKC system, a complete group separation of the two types of polymer molecules could be obtained.

Separation of five recently commercialized selective serotonin reuptake inhibitor antidepressants by capillary electrophoresis

Several strategies were applied in order to find a simple electrophoretic method for the investigation of five selective serotonin reuptake inhibitors (SSRI) (citalopram, fluoxetine, fluvoxamine, paroxetine, and sertraline). Variation of the pH and of the ionic strength of the background electrolyte (BGE) were studied, but did not enable separation of the analytes. A micellar electrokinetic chromatography (MEKC) strategy for the simultaneous separation of the five SSRIs was developed involving a sodium dodecylsulfate (SDS) MEKC system. The electroosmotic flow (EOF) and the migration of the analytes were determined for separation buffers of several surfactant concentrations and organic modifier percentages. The most favourable MEKC system consisted of 20 mmol/L SDS in a phosphate buffer (pH 7.5) with 30% methanol; the separation was carried out using an uncoated fused-silica capillary, a separation voltage of 25 kV with currents typically less than 40 μA, and spectrophotometric detection at 200 nm. Full separation of the mixture with baseline resolution of all analytes was obtained.

Solute–solvent interactions in micellar electrokinetic chromatography: III. Characterization of the selectivity of micellar electrokinetic chromatography systems

Several micellar electrokinetic chromatography (MEKC) systems (sodium dodecyl sulfate, lithium dodecyl sulfate, lithium perfluorooctanesulfonate, sodium cholate, sodium deoxycholate, tetradecyltrimethylammonium bromide and hexadecyltrimethylammonium bromide) have been characterized by means of the solvation parameter model. It has been observed that the coefficients of the correlation equations depend strongly on the particular set of compounds analyzed. Principal component analysis has been used to characterize the 2975 compounds with available solute descriptors and to select an appropriate subset of compounds to be analyzed by MEKC. With this set of compounds, the MEKC systems have been characterized. Principal component analysis has also been used to show the similarities and differences between the properties of the surfactants characterized by MEKC.

Solute-solvent interactions in micellar electrokinetic chromatography: IV. Characterization of electroosmotic flow and micellar markers.

A wide study of the compounds and procedures mostly used to determine the electroosmotic flow (EOF) and micelle elution times has been done in seven different micellar electrokinetic chromatography (MEKC) systems. These systems are formed from mixtures of an aqueous buffer with the surfactants sodium dodecyl sulfate, lithium dodecyl sulfate, lithium perfluorooctane sulfonate, sodium cholate, sodium deoxycholate, tetradecyltrimethylammonium bromide and hexadecyltrimethylammonium bromide. The solvation parameter model has been used to evaluate the usefulness of the compounds studied as EOF or micellar markers in each of the seven MEKC systems. It is demonstrated that methanol, acetonitrile and formamide are the best EOF markers, and that dodecanophenone is the best micellar marker.

Comparison of microemulsion electrokinetic chromatography and solvent-modified micellar electrokinetic chromatography

Microemulsion electrokinetic chromatography (MEEKC) was investigated with the goal of elucidating the influence exerted by the amount of lipophilic organic phase and the amount of co-surfactant on the migration of a number of neutral test solutes. Furthermore, MEEKC was compared to solvent-modified micellar electrokinetic chromatography (MEKC). The organic solvents methanol, ethanol, 1-propanol, 1-butanol, acetonitrile, tetrahydrofuran, and 2-ethoxyethanol serving as co-surfactants in MEEKC were used in similar concentrations in MEKC systems in order to compare the selectivity and separation efficiency of the two kinds of system. The solvent-modified MEKC technique proved to be very similar to MEEKC in performance with respect to both separation selectivity and efficiency of separation.