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This study establishes a method for rapid detection of clonidine and cyproheptadine in foods of animal origin. In order to obtain the best detection method, capillary zone electrophoresis (CZE), large volume sample stacking (LVSS), and sweeping-micellar electrokinetic capillary chromatography (sweeping-MEKC) were used respectively. The limits of detection (LODs) of clonidine and cyproheptadine by LVSS-CZE were 0.028 μg mL−1 and 0.034 μg mL−1, and those by sweeping-MEKC were 0.023 μg mL−1 and 0.031 μg mL−1, respectively. Compared with the CZE method, the two online pre-concentration technologies have greatly improved the detection sensitivity and achieved good enrichment results. However, compared with the sweeping-MEKC system, the LVSS system consumed a longer time and was greatly affected by the actual sample matrix. The sweeping-MEKC method was proved to be suitable for real sample analysis. Under the best sweeping-MEKC conditions, clonidine and cyproheptadine could be well separated within 8 min and good linear relationships in the range of 0.1–1.0 μg mL−1 (r 2 > 0.99) were obtained. This method was successfully applied to the determination of clonidine and cyproheptadine in animal-derived foods with the recoveries of 82.3%–90.1% and the relative standard deviations (RSDs) less than 3.11%. The sweeping-MEKC method is simple to operate and has great potential in the rapid detection of clonidine and cyproheptadine in animal-derived foods.

A sweeping micellar electrokinetic chromatography (sweeping-MEKC) method was developed for the determination of 1,7-naphthalenediol, 2,3-naphthalenediol, 1,5-naphthalenediol and 2,7-naphthalenediol in cosmetics. Several parameters affecting sweeping-MEKC method were studied systematically and the separation conditions were optimized as 20 mM NaH2PO4–110 mM SDS and 40% (v/v) MeOH (pH 2.4), with −22 kV applied voltage and UV detection at 230 nm. The sample matrix is 60 mmol L−1 NaH2PO4 and sample introduction was performed at 3 psi for 6 s. Separation of the four naphthalenediols was completed in less than 17 min. Limit of detection (LOD) and limit of quantitation (LOQ) are 0.0045∼0.0094 μg mL−1 and 0.015∼0.031 μg mL−1. Linear relationship (r 2 > 0.999) is satisfactory at the range of 0.1–10 μg mL−1. The developed method has been successfully applied to the determination of the four naphthalenediols in real cosmetic samples, with recoveries in foundation, sun cream and lotion in the range of 92.3%∼106.8% and relative standard deviation (RSD) less than 4.15%. A HPLC method described in the National Standards of the People’s Republic of China was carried out for the comparison with the proposed method. The results showed that the proposed sweeping-MEKC method has the advantages of fast, low cost with comparative sensitivity.

Ganoderma (lingzhi) is a famous herbal medicine and edible supplement in oriental countries for a long history. In this study, a simple micellar electrokinetic chromatography (MEKC) method was established for the analysis of nucleosides and bases, the major bioactive components in Ganoderma for the first time. By optimizing the borate concentration, the sodium dodecyl sulfate (SDS) concentration and the pH value of running buffer, 10 nucleosides and bases achieved an ideal separation. In real sample analysis, the developed method was successfully used to determine the 10 target analytes in 23 batches of Ganoderma samples from different regions. Results indicated that contents of 10 investigated analytes in each sample showed obvious variation. The principal components analysis (PCA) and hierarchical cluster analysis (HCA) analysis classified the samples into three groups, and the HCA tree visualized the relationships which was mainly contributed by geographical partition. The results indicated geographical origin to be an important factor that affect the accumulation of nucleosides and bases in Ganoderma. In summary, this study provides a simple and practical strategy for quality assessment and cultivation reference of Ganoderma.

This work presents an innovative silica-layer coated capillary with comparison study of the silica-layer coated capillary and the fused-silica capillary for the separation of seven phenolic acids viz. p-hydroxyphenylacetic acid (PHPA),p-coumaric acid (PCA),p-hydroxybenzoic acid (PHBA), caffeic acid (CFA), (3,4-dihydroxyphenyl)acetic acid (DHPA), gallic acid (GLA), and 2,3,4-trihydroxybenzoic acid (THBA), together with caffeine (CF), by capillary electro-chromatography (CEC) and micellar electrokinetic chromatography (MEKC), respectively. The running buffer was 25.0 mM borate at pH 9.0, with addition of 50.0 mM sodium dodecyl sulfate for the MEKC mode. The non-coated capillary could not separate all seven phenolic acids by CEC or MEKC. This was achieved using the coated capillary for both CEC and MEKC. The innovative coated capillary with CEC had plate numberN ≥ 2.0 × 104 m−1and resolutionRs ≥ 1.6 for all adjacent pairs of peaks. The capillary was also able to separate GLA and THBA which are structural isomers. Although MEKC mode provided comparable efficiency and selectivity, the reduced EOF of the coated capillary led to longer separation time. The linear calibration range of the seven phenolic acids and caffeine were different but the coefficients of determinations (r2) were all > 0.9965. The precisions of the relative migration times and peak area ratios of analyte to internal standard were 0.1–1.8% and 1.8–6.8%, respectively. There were no statistical differences in the efficiency of separation of the phenolic acids and caffeine for three coated capillaries. It was applied to the analysis of caffeine and phenolic acids in brewed tea using tyramine as the internal standard. The tea samples were diluted prior to analysis by CEC. The separation was less than 15 min. Caffeine, gallic acid andp-coumaric acid were detected and quantified. Caffeine and gallic acid contents were 10.8–15.0 and 2.6–4.8 mg g−1dry tea leaves, respectively.p-Coumaric acid was detected in only one of the samples with a content of 0.4 mg g−1. Percent recoveries of spiked diluted samples were 90 ± 9 to 106 ± 13%, respectively.Article highlightsSilica-layer coated capillary is first reported for simultaneous separation of seven phenolic acids by non-MEKC analysis.Performance between coated, and non-coated capillaries with analysis by CEC and MEKC were compared.Plate number, resolution, capillary reproducibility, and electroosmotic flow mobility are investigated.Graphical abstract

We present the results of studying and selectiing optimal conditions for the determination of caffeine, catechins and gallic acid in black tea by the method of micellar electrokinetic chromatography. The choice of analytes is determined by the fact that they form the main consumer qualities of tea and, due to their antioxidant properties, have a positive effect on human health. Optimization of the electrophoretic detemination of seven catechins, caffeine and gallic acid in black tea was carried out using the method of ex­ periment design. Optimized conditions — 25 mM phosphate buffer solution with pH 7.4, containing 30 mM SDS and 5% ethyl alcohol — provided a satisfactory resolution of all electrophoretogram peaks, the analysis time was 25 minutes, and the current in the system did not exceed 120 pA. After the assessment of the metrological characteristics of the proposed method the standard deviation of the determina­tion of analytes did not exceed 15%. The samples of black tea from Ceylon, Chinese, Assamese, Indian, Kenyan and Krasnodar regions of growth were analyzed, and an array of data on the contents of the analytes under study was formed. Discriminant analysis was used to develop a model and obtain classifica­tion functions for six tea groups from different regions of the world. Proceeding from the obtained classifi­cation functions, a scatter diagram of canonical values was constructed, which showed that the samples of Krasnodar and Chinese tea were localized from all the studied groups. Indian, Assamese and Ceylon teas formed a single area with close to the Chinese tea group. The correctness of the model was checked and revealed the total predictive force about 92%. It is shown that the content of catechins, gallic acid and caf­feine are suitable markers for the classification of black tea samples from different regions of origin.

The use of modifiers in the electrophoretic separation of multicomponent mixtures makes it possible to significantly expand the analytical capabilities of capillary electrophoresis. Modifiers of electrophoretic systems, different in nature and properties, can perform different functions: form stationary and pseudo-stationary phases, prevent the adsorption of analytes on the walls of a quartz capillary, help to reduce the limits of detection for analytes during preconcentration, etc. In this review, various versions of the application of modifiers to the electrophoretic separation and preconcentration of analytes of different nature are considered on an example of imidazolium ionic liquids, which can act as stationary and pseudo-stationary phases, and also directly participate in the online preconcentration of analytes of hydrophilic and hydrophobic nature.

A sweeping micellar electrokinetic chromatography (sweeping-MEKC) enrichment model was established for the determination of three chlorophenols (CPs) in cosmetics, namely, bithionol, pentachlorophenol (PCP), and 2,4,6-trichlorophenol (2,4,6-TCP). The optimum electrophoretic conditions were 20 mM NaH2PO4-80 mM sodium dodecyl sulfate (SDS) and 30% (v/v) acetonitrile (pH 2.3). The optimum on-line concentration conditions were as follows: sample matrix, 100 mM NaH2PO4; pressure injection at 20.67 kPa (3 psi) for 25 s. The linear range of bithionol, PCP, and 2,4,6-TCP are 0.20–4.00 μg mL−1, 0.10–2.00 μg mL−1, and 0.05–2.00 μg mL−1 respectively, with correlation coefficient (r) over 0.9972. The limits of detection (LOD) based on three times the signal-to-noise ratio (S/N = 3) are in the range of 0.0061–0.024 μg mL−1. Recoveries for the three CPs in powder and lotion samples are between 79.7 and 110.2% with relative standard deviation (RSD) of 1.38–5.54% and 92.2–121.3% with RSD of 0.72–6.09%, respectively. The proposed method can provide reference for the determination of trace CPs in different sample matrix.

Bile salts are a category of natural chiral surfactants which have ever been used as the surfactant and chiral selector for the separation of many chiral compounds by micellar electrokinetic chromatography (MEKC). In our previous works, the application of sodium cholate (SC) in the separation of four stereoisomers of palonosetron (PALO) by MEKC has been studied systematically. In this work, the parameters of other bile salts, including sodium taurocholate (STC), sodium deoxycholate (SDC), and sodium taurodeoxycholate (STDC) in the separation of PALO stereoisomers by MEKC were measured and compared with SC. It was found that all of four bile salts provide chiral recognition for both pairs of enantiomers, as well as achiral selectivity for diastereomers of different degrees. The structure of steroidal ring of bile salts has a greater impact on the separation than the structure of the side chain. The varying separation results by different bile salts were elucidated based on the measured parameters. A model to describe the contributions of the mobility difference of solutes in the aqueous phase and the selectivity of micelles to the chiral and achiral separation of stereoisomers was introduced. Additionally, a new approach to measure the mobility of micelles without enough solubility for hydrophobic markers was proposed, which is necessary for the calculation of separation parameters in MEKC. Under the guidance of derived equations, the separation by SDC and STDC was significantly improved by using lower surfactant concentrations. The complete separation of four stereoisomers was achieved in less than 3.5 min by using 4.0 mM of SDC. In addition, 30.0 mM of STC also provided the complete resolution of four stereoisomers due to the balance of different separation mechanisms. Its applicability for the analysis of a small amount of enantiomeric impurities in the presence of a high concentration of the effective ingredient was validated by a real sample.

INTRODUCTION Picfeltarraenins IA, IB and IV and acteoside are the four bioactive ingredients of Picria fel-terrae Lour. Their pharmacological effects include central inhibitory, cardiovascular, anti-inflammatory, anti-pyretic, analgesic, anti-bacterial, antioxidative and anti-tumor effects. OBJECTIVE We aimed to develop an efficient micellar electrokinetic chromatography (MEKC) method modified with mixed organic solvents for the simultaneous separation and determination of the four components in Picriae Herba and its formulations. METHODS Method optimization was carried out by investigating influences of significant factors on the separation, and this method was successfully applied for the determination of the four components in Picriae Herba and its formulations. RESULTS The optimal running buffer was composed of 20 mM sodium tetraborate, 40 mM sodium cholate, 10% (v/v) methanol and 10% (v/v) isopropanol (pH 9.76). The separation voltage was 18 kV, the temperature was 25°C and the detection wavelength was 266 nm. Under the optimal separation conditions, the baseline separation of four components was achieved in less than 14 min. The correlation coefficients of the calibration curves were 0.9984-0.9995 for the analytes. The intraday and interday precision ranged from 1.5% to 2.5% and from 1.4% to 5.0%, respectively. Recoveries of analytes varied from 96.6% to 104.1%. CONCLUSION The method was proved suitable for the determination of four components in Picriae Herba and its formulations. Good performance was obtained under optimal conditions, and the method provides an effective tool for the quality control of Picriae Herba and its formulations.

In this work, high-speed micellar electrokinetic chromatography with LIF detection was applied to study the antagonism between three intestinal bacteria, Escherichia coli (E. coli), Bacillus licheniformis (B. licheniformis) and Bacillus subtilis (B. subtilis). The fluorescent derivatization for the bacteria was performed by labeling the bacteria with FITC. In a high-speed CE device, the three bacteria could be completely separated within 4 min under the separation mode MEKC. The BGE was 1 × TBE containing 30 mM SDS and 1.5 × 10-5 g/mL polyethylene oxide. The limits of detection for E. coli, B. licheniformis and B. subtilis were 2.80 × 106 CFU/mL, 1.60 × 106 CFU/mL and 1.90 × 106 CFU/mL respectively. Lastly, the method was applied to investigate the antagonism between the three bacteria. The bacteria were mixed and cultured for 7 days. The samples were separated and determined every day to study the interaction between bacteria. The resulted showed that B. licheniformis and B. subtilis could not inhibit each other, but they could effectively inhibit the reproduce of E. coli. The method developed in this work was quick, sensitive and convenient, and it had great potential in the application of antagonism study for bacteria. This article is protected by copyright. All rights reserved.

The calcium signaling protein calmodulin regulates numerous intracellular processes. We introduce a sensitive microchip assay to separate and detect calmodulin binding proteins. The assay utilizes an optimized microchip electrophoresis protein separation platform with laser-induced fluorescence detection. Fluorescence-labeled calmodulin modified with a photoreactive diazirine crosslinker allowed selective detection of calmodulin binding proteins. We demonstrate successful in-vitro crosslinking of calmodulin with two calmodulin binding proteins, calcineurin and nitric oxide synthase. We compare the efficacy of commonly applied electrophoretic separation modes: microchip capillary zone electrophoresis, microchip micellar electrokinetic chromatography/gel electrophoresis, and nanoparticle colloidal arrays. Out of the methods tested, polydymethylsiloxane/glass chips with microchip zone electrophoresis gave the poorest separation, whereas sieving methods in which electro-osmotic flow was suppressed gave the best separation of photoproducts of calmodulin conjugated with calmodulin binding proteins.

Separation of a set of model proteins was tested on a microchip electrophoresis analytical platform capable of sample injection by two different electrokinetic mechanisms. A range of separation modes—microchip capillary zone electrophoresis , microchip micellar electrokinetic chromatography, and nanoparticle-based sieving—was tested on glass and polydimethylsiloxane/glass microchips and with silica-nanoparticle colloidal arrays. The model proteins calmodulin (18 kiloDalton), bovine serum albumin (66 kiloDalton) and concanavalin (106 kiloDalton) were labeled with Alexa Fluor 647 for laser-induced fluorescence detection. The best separation and resolution were obtained in a silica-nanoparticle colloidal array chip.

Caffeine (CA) is a common xanthine alkaloid found in tea leaves, coffee beans, and other natural plants, and is the most widely used psychotropic substance in the world. Accumulating evidence suggests that low plasma levels of CA and its metabolites may serve as reliable diagnostic markers for early Parkinson's disease (PD) patients. In this study, we demonstrated a new MEKC method for determining CA and its three main downstream metabolites, paraxanthine (PX), theobromine (TB), and theophylline (TP), in human plasma. Plasma samples were collected, and analyzed using MEKC, after SPE. The running buffer was composed of 35 mM phosphate, pH of 10.5, and 25 mM SDS. The separation voltage was 15 kV and the detection wavelength was at 210 nm. Under the optimum conditions, four distinct analytes were completely separated and detected in less than 12 min. Method limits of detection were as low as 7.5 ng/mL for CA, 5.0 ng/mL for TB, and 4.0 ng/mL for both PX and TP. The recoveries were between 88.0% and 105.9%. This method was successfully applied to 27 human plasma samples. The results indicate that the plasma concentrations of the four analytes are significantly lower in patients with early PD than in control subjects (p < 0.05). The area under curve was improved to 0.839 when CA and its three main metabolites were included, suggesting that MEKC testing of CA, TP, TB, and PX may serve as a potential method for early diagnosis of PD.

A micellar electrokinetic capillary chromatography (MEKC) method has been developed and validated for simultaneous analysis of febuxostat and its related substances in active pharmaceutical ingredients and pharmaceutical formulations. Analysis was performed in an uncoated fused silica capillary (34 cm effective length, 50 μm diameter). Method sensitivity was enhanced using an extended light path capillary, with an optical path 150 μm and the choice of 320 nm detector operat-ing wavelength due to lower background noise and higher selectivity. BGE consisted of 20 mM borate buffer pH 9.3 and 50 mM SDS and 2% acetonitrile. Analysis was performed at 30 kV separation voltage and 32 °C capillary temperature. The method was linear over the range 1.0–10.0 μg mL−1 for each impurity corresponding to reporting limits regarding the ICH Q3A guidelines. The method has proven to be selective, precise, accurate and robust. Proposed MEKC method has been successfully applied for the quantitative determination of febuxostat and purity assay of active pharmaceutical ingredient and tablet dosage forms.

Dairy cow feed contains, among other ingredients, soybeans, legumes, and clover, plants that are rich in phytoestrogens. Several publications have reported a positive influence of phytoestrogens on human health; however, several unfavorable effects have also been reported. In this work, a simple, selective, and eco-friendly method of phytoestrogen isolation based on the technique of noncovalent molecular imprinting was developed. Genistein was used as a template, and dopamine was chosen as a functional monomer. A layer of molecularly imprinted polymers was created in a microtitration well plate. The binding capability and selective properties of obtained molecularly imprinted polymers were investigated. The imprinted polymers exhibited higher binding affinity toward chosen phytoestrogen than did the nonimprinted polymers. A selectivity factor of 6.94 was calculated, confirming satisfactory selectivity of the polymeric layer. The applicability of the proposed sensing method was tested by isolation of genistein from a real sample of bovine milk and combined with micellar electrokinetic capillary chromatography with UV-visible detection.

Nucleosides and related compounds have vital physiological and pharmacological activities. In this study, on-line preconcentration using electrokinetic supercharging (EKS)-sweeping method was propo...

A simple, sensitive, and efficient method has been developed for the determination of the seven neonicotinoid insecticides commercially available (imidacloprid, thiacloprid, clothianidin, thiamethoxam, acetamiprid, nitenpyram, and dinotefuran) and the main metabolite 6-chloronicotinic acid. Micellar electrokinetic chromatography (MEKC) mode was applied, using 48.5 cm of total length capillary (50 μm i.d.) with an extended light-path capillary (150 μm). The running electrolyte consisted of 25 mM sodium tetraborate buffer (pH 9.2) containing 120 mM of sodium dodecyl sulfate and 15% of methanol (v/v). A voltage of 27 kV and a temperature of 25 °C were applied. Samples dissolved in deionized water were hydrodynamically injected at 50 mbar for 12 s, achieving the analysis in less than 12 min. Diode array detection (DAD) was performed at 220, 254, and 270 nm, depending on the analyte. Two different methodologies as sample treatments were developed; for water samples, solid-phase extraction was checked using different cartridges (C18, Oasis® HLB, Oasis® HLB Prime, and Strata-X), being the best option Oasis® HLB for preconcentration and cleanup. In the case of soil samples, a simple solid-liquid extraction was applied using a mixture of 1:3 (v/v) acetonitrile/dichloromethane. Satisfactory linearity, trueness, and precision were achieved, with detection limits in the range of 0.1-0.4 μg L-1 for river water and 1.0-2.9 μg kg-1 for soil samples. Recoveries in the range of 80-107% for all of the assayed neonicotinoids in water samples of different origin and 73-92% for soil samples were achieved.

4(5)-Methylimidazole (4(5)MEI) is a product of the Maillard reaction between sugars and amino acids, which occurs during the thermal processing of foods. This compound is also found in foods with caramel colorants additives. Due to its prevalence in foods and beverages and its potent carcinogenicity, 4(5)MEI has received federal and state regulatory agency attention. The aim of this review is to present the extraction procedures of 4(5)MEI from food matrices and the analytical methods for its determination. Liquid and gas chromatography coupled with mass spectrometry are the techniques most commonly employed to detect 4(5)MEI in food matrices. However, the analysis of 4(5)MEI is challenging due to the high polarity, water solubility, and the absence of chromophores. To overcome this, specialized sample pretreatment and extraction methods have been developed, such as solid-phase extraction and derivatization procedures, increasing the cost and the preparation time of samples. Other analytical methods for the determination of 4(5)MEI, include capillary electrophoresis, paper spray mass spectrometry, micellar electrokinetic chromatography, high-performance cation exchange chromatography, fluorescence-based immunochromatographic assay, and a fluorescent probe.

The first method by micellar electrokinetic chromatography (MEKC) for the determination of empagliflozin in tablets was developed and validated following the ICH guidelines. The separation was achieved in a fused silica capillary with 50 µm x 40 cm (inner diameter x effective length) at 28 oC, +28 kV voltage, hydrodynamic injection 4s (50 mBar), detection at 225 nm and paracetamol was the internal standard. The running electrolyte was a mixture of 20 mM tris(hydroxymethyl)aminomethane (pH 10) and 100 mM sodium dodecyl sulphate (1:1). Specificity was evaluated by the stress testing and the method was specific, with no interference of the degradation product. Linearity was observed in the range of 50 to 150 μg/mL (r=0.9999). The method showed adequate accuracy (recovery value=100.60±0.60%), precision (RSD values

Toxic organochloride molecules are widely used in industry for various purposes. With their high volatility, the direct detection of organochlorides in environmental samples is challenging. Here, a new organochloride detection mechanism using 1,5-diazabicyclo[4.3.0]non-5-ene (DBN) is introduced to simplify a sensing method with higher detection sensitivity. Three types of organochloride compounds-trichloroethylene (TCE), dichloromethane (DCM), and dichlorodiphenyltrichloroethane (DDT)—were targeted to understand DCM conjugation chemistry by using nuclear magnetic resonance (NMR) and liquid chromatography with a mass spectrometer (LC-MS). 13C-NMR spectra and LC-MS data indicated that DBN can be labeled on these organochloride compounds by chlorine–nitrogen interaction. Furthermore, to demonstrate the organochloride sensing capability, the labeling yield and limit of detection were determined by a colorimetric assay as well as micellar electrokinetic chromatography (MEKC). The interaction with DBN was most appreciable for TCE, among other organochlorides. TCE was detected at picomolar levels, which is two orders of magnitude lower than the maximum contaminant level set by the United States Environmental Protection Agency. MEKC, in conjunction with this DBN-labeling method, enables us to develop a field-deployable sensing platform for detecting toxic organochlorides with high sensitivity.

The online preconcentration technique, cyclodextrin-assisted sweeping (CD-sweeping), coupled with micellar electrokinetic chromatography (MEKC) was established to determine 13-cis-retinoic acid (13-cis-RA), all-trans-retinoic acid (all-trans-RA) and 4-oxo-13-cis-retinoic acid (4-oxo-13-cis-RA) in human plasma. A CD-sweeping buffer (45 mM borate (pH 9.2), containing 80 mM sodium dodecyl sulfate (SDS) and 22 mM hydroxypropyl β-CD (HP-β-CD) was introduced into the capillary and, then, the sample dissolved in 70 mM borate (pH 9.2): methanol = 9:1 (v/v) was injected into capillary by pressure. The separation voltage was 23 kV. Compared to the conventional cyclodextrin-micellar electrokinetic chromatography (CD-MEKC) method, the new technique achieved 224–257-fold sensitivity enrichment of analytes. The limits of detection of 13-cis-RA, all-trans-RA were 1 ng/mL, whereas that of 4-oxo-13-cis-RA was 25 ng/mL in plasma. The linear ranges of 13-cis-RA, all-trans-RA were between 15 and 1000 ng/mL, whereas that of 4-oxo-13-cis-RA was between 75 and 1500 ng/mL. The coefficient of correlation between the concentration of analytes and peak area ratio of analytes and internal standard (2, 4-dihydroxy-benzophenone) for intra-day (n = 3) and inter-day (n = 5) analyses were both greater than 0.999. The optimized experimental conditions were successfully applied to determine 13-cis-retinoic acid and its metabolites in plasma samples from a patient during the administration of 13-cis-RA for treating acne.

Dunn, Hankins, and Ghowsi observed separation in the case where ions incapillary electrophoresis migrate in the opposite direction of electroosmosis and ions travel faster than flow, eventually separating them and reaching the detector.For the separation of p-xylene and toluene, a similar form of electrokinetic chromatography called reverse direction MECC has been developed (micellar electrokinetic capillary chromatography).In reverse direction micellar electrokinetic capillary chromatography, the influence of injection time on separation is explored. Hydrostatic and diffusion injection were investigated in this study.

Pseudophase microextraction (PPME) as a simple in-line sample concentration technique in micellar electrokinetic chromatography (MEKC) is presented. In contrast to popular electric-field driven stacking techniques in MEKC such as sweeping, PPME is pressure-driven. The technique afforded up to 403-2968x improvements in peak heights for fenoprop, dichlorprop, 1- and 2-naphthol compared to typical injection. Under the same MEKC conditions, the improvements in PPME were up to 23-59x better compared to sweeping. Briefly in PPME, the entire capillary was loaded (up to 20 capillary volumes) with the sample prepared in a dilute solution of cetyltrimethylammonium bromide ([CTAB] > critical surface aggregation concentration). The CTAB formed aggregates at the inner capillary walls and these aggregates acted as a stationary chromatographic pseudophase. After clean-up via flushing the capillary with purified water, the MEKC background solution (BGS) with sodium dodecyl sulfate was then introduced by pressure from the outlet end to elute the retained analytes. The analytes concentrate at front of the BGS and the front was moved to the inlet end of the capillary prior to MEKC. Optimization strategies and current limitations in PPME-MEKC are described. The linear ranges using a 4 capillary volume sample load obtained for fenoprop, dichlorprop, 1- and 2-naphthol were between 1 and 160 ng/mL (r2s > 0.996), LOQs = 1-2.5 ng/mL and repeatability %RSDs (n=6) were 5% (intra-day) and 7% (inter-day) (using low analyte concentrations 1-5x LOQ). PPME-MEKC with simple dilution of fortified real samples (no off-line sample concentration) was also able to detect low levels of dichlorprop (10 ng/mL, limit set in Australia) and 1- and 2−naphthol (7.5-15 ng/mL) in a drinking water and natural water sample, respectively (% recovery = 84-108%). The concept of PPME may find use in other modes of capillary electrophoresis and other nano-microscale separations.

Rotenone, a naturally occurring isoflavone compound, is applied to water as a piscicide to manage undesired fish populations in lakes, ponds, rivers and in aquaculture. A rapid and sensitive method based on solid-phase extraction (SPE) combined with large-volume sample stacking with polarity switching (LVSS) and micellar electrokinetic chromatography (MEKC) has been developed for the identification and quantification of rotenone in lake water. Several experimental parameters for MEKC, LVSS and SPE were investigated to achieve the optimum conditions necessary for the analysis. The optimized conditions which included a background electrolyte containing 20 mM sodium borate and 25 mM sodium dodecyl sulfate (SDS) at pH 10 were used to successfully detect rotenone within 17 min. Using a combined SPE and LVSS approach, the rotenone signal was enhanced by 1000-fold compared with a normal capillary electrophoresis analysis, and limits of detection and quantification obtained were 3 and 10 μg L−1, respectively. The calibration curve was linear for rotenone concentration over the range of 10–100 μg L−1 and the method was highly reproducible with the relative standard deviation of the peak areas and migration times for method intra-day repeatability (n = 6) found to be 5.4% and 0.6%, respectively. Quantitative recoveries ranging from 85 to 88% were obtained in the lake water matrices. The potential of the proposed method to be used for quantitative determination of rotenone at trace level concentrations in water samples was demonstrated by analyzing lake water samples including surface (1 m) and bottom (13 m), which makes it a suitable practical method for analyzing rotenone residues in water.

Charge heterogeneity is an important critical quality attribute for the analysis of monoclonal antibodies (mAbs). For this, (imaged) capillary isoelectric focusing ((i)cIEF), ion exchange chromatography (IEC) and, recently, capillary zone electrophoresis (CZE) are the predominantly used techniques. In order to investigate which one is most suitable to answer a specific analytical question, here, the four aforementioned separation techniques were systematically evaluated using NISTmAb and Infliximab as test molecules. The performance parameters (precision, separation efficiency, linearity and sensitivity) were determined under comparable conditions. Moreover, important aspects for daily routine such as speed and ease of use were considered. Each technique has its own pros and cons. The (i)cIEF methodology is distinguished by its excellent separation efficiency. In addition, the native fluorescence mode in icIEF is a good tool to analyze small sample amounts (LOQ: 2.8 mg/l for Infliximab). Nevertheless, high performance liquid chromatography (HPLC) still has superior precision. CZE, and also micellar electrokinetic chromatography (MEKC), have emerged as further interesting alternatives. For all techniques, variations connected to the sample preparation strongly influence precision. Looking at the relative standard deviation (RSD) values of the relative peak areas, all techniques provide acceptable performance (RSD: 0.6–1.6%).

A rapid, simple, and sensitive method has been developed for the analysis of pyrethroid herbicides in fruits by using headspace in-tube microextraction (HS-ITME) coupled with reverse-flow micellar electrokinetic capillary chromatography (RF-MECC). In the newly developed method, by placing a capillary filled with background electrolyte (BGE) of RF-MECC in the HS above the sample solution, the pyrethroid herbicides were extracted into the acceptor phase in the capillary. After extraction, electrophoresis of the extracts in the capillary was carried out. The influence of some essential BGE components such as sodium dodecyl sulfate (SDS) and organic modifiers concentrations was investigated. Extraction parameters were also systematically investigated, including the extraction temperature, extraction time, salt concentration, and volume of the sample solution. Under the optimized conditions, enrichment factors for three pyrethroids were 309, 133, and 288, respectively. The proposed method provided a good linea...

Diode lasers, especially 405 nm diode laser, as excitation sources offer new opportunities for laser-induced fluorescence (LIF) detection, but the lack of available derivatization reagents limits their widespread applications. Herein, a commercial fluorescent dye, 7-(diethylamino)coumarin-3-carboxylic acid (DEAC-C) was introduced as derivatization reagent and corresponding derivatization strategy for DEAC-C labeling sulfonamides was developed for 405 nm LIF detection. After systematic optimization, three sulfonamides including hydrochlorothiazide (HCTZ), chlorothiazide (CTZ) and chlortalidone (CTD) could be efficiently labeled by DEAC-C in the presence of cyanuric chloride and triethylamine (1.5%) in acetonitrile at 50 °C for 180 min. Based on the laboratory-built capillary electrophoresis-LIF system, a robust method was then proposed for the separation of DEAC-C labeled sulfonamides by micellar electrokinetic chromatography (MEKC) using the classic borax-SDS system. Under the optimized conditions, a baseline separation of three sulfonamides was achieved within 15 min, and the detection limits were determined to be 0.24, 0.29, and 0.23 nM for HCTZ, CTZ, and CTD, respectively. Furthermore, the developed MEKC-LIF method was validated using sulfonamide standards and spiked human urine sample and successfully applied for the analysis of three sulfonamides in complex pharmaceutical and physiological samples.

Mass spectrometry has become a powerful tool in the field of bioanalysis. The combination of capillary electrophoresis with mass spectrometry (CE-MS) offers fast and high-resolution separation, sensitive and selective detection with small sample volumes, which is suitable for cell analysis. Here, we review the recent technical development of CE, MS and hyphenated strategies as well as their potential in cell analysis. In particular, we discuss the multiple MS compatible CE modes including capillary zone electrophoresis, capillary isoelectric focusing, capillary isotachophoresis, micellar electrokinetic capillary chromatography and capillary electrochromatography for achieving better separation efficiency and selectivity before MS detection. For enhancing the sensitivity of CE-MS, several online preconcentration methods including dynamic pH junction, transient isotachophoresis and solid phase extraction are also reviewed and discussed. We also summarize the existing applications of CE-MS in cell analysis including cell biomacromolecule analysis, cell metabolism and single cell analysis.

Introduction Hirsutine and hirsuteine are the main pharmacological activity ingredients of Uncaria rhynchophylla (UR), playing an important role in treating mental and cardiovascular diseases, such as Alzheimer's disease, hypertension, Parkinson's disease, potential anti-cancer activities and so on. Objective To develop a cyclodextrin-modified micellar electrokinetic capillary chromatography (CD-MEKC) method for the simultaneous separation and determination of hirsutine and hirsuteine from UR and its formulations. Methodology The optimal method was developed by investigating influences of significant factors on the separation, and this method was successfully applied for the determination of hirsutine and hirsuteine in UR and its formulations. Results The optimal background electrolyte (BGE) consisted of 40 mM sodium dihydrogen phosphate (pH 7.0), 150 mM 2,6-dimethyl-β-cyclodextrin (DM-β-CD), 3 mM mono-(6-ethylenediamine-6-deoxy)-β-cyclodextrin (ED-β-CD), and 30 mM sodium cholate (SC). Under these conditions, hirsutine and hirsuteine were successfully separated within 13 min at the separation voltage of 15 kV, temperature of 25°C and the detection wavelength of 224 nm. For the analytes, linear calibration curves were performed within the range 5.0-160.0 μg/mL. The limit of detection (LOD, S/N = 3) and the limit of quantitation (LOQ, S/N = 10) were 0.41, 1.42 μg/mL for hirsutine and 0.60, 2.17 μg/mL for hirsuteine, respectively. The recoveries of three samples were from 97.9% to 102.3%. Conclusion The method was successfully applied to the determination of hirsutine and hirsuteine in UR and its formulations. Meanwhile, it provides an effective reference of the quality control of UR and its formulations.

A stability-indicating micellar electrokinetic capillary chromatography (MEKC) method was developed and validated for the analysis of doxorubicin hydrochloride in injectable pharmaceutical dosage forms, using methotrexate as internal standard. A fused-silica capillary (50 µm i.d.; effective length, 40 cm) and a running electrolyte solution consisting of 10 mM borate buffer and 20 mM anionic surfactant SDS, at pH 9.3, were set as the best experimental conditions. Moreover, the capillary temperature was maintained at 26 ºC, while the applied voltage was +26 kV. Hydrodynamic sample injection (6 s at 50 mbar) was used, and the detection was set at 260 nm using a photodiode array detector. The method was validated for the requirements specificity, linearity, precision, accuracy, and robustness, following the International Conference on Harmonisation (ICH) guidelines. The method linearity was proven in the range of 25-125 µg/mL (r = 0.9995). Forced degradation studies were successfully conducted, evidencing the specificity and stability-indicating capability of the method. In addition, no interference of the excipients from the formulation was detected. The values of accuracy and precision were within the acceptable limits, and robustness studies were performed by a two-level full factorial design. The proposed method fulfilled all validation parameters and was shown to be suitable for quantitative analyses of doxorubicin, contributing, thus, to the establishment of new alternatives with advantages for the quality control of pharmaceutical formulations.

Phenylenediamines (PDs), which are reported to cause allergic dermatitis and possess genotoxicity and carcinogenicity, are the ingredients used in permanent hair dyes. The fluorescent derivatization strategy coupled with micellar electrokinetic chromatography (MEKC) were established to analyze four PDs, including o-phenylenediamine (OPD), m-phenylenediamine (MPD), p-phenylenediamine (PPD) and toluene-2,5-diamine (PTD). Additionally, 5-(4, 6-dichlorotriazinyl) aminofluorescein (DTAF) was used as a fluorescent reagent derived at amino groups of PDs and underwent nucleophilic substitution reaction to improve the detection sensitivity. The derivatization condition reacted at 90 °C for 10 min in alkaline conditions. The optimized separation conditions were 20 mM borate (pH 8.0) containing 10 mM Brij 35 and 35% (v/v) methanol. The limits of detection (S/N = 3) for MPD, PTD, PPD and OPD were 25, 25, 50 and 100 nM, respectively. Compared to MEKC-UV, the sensitivity enhancements were 30- to 81-fold when PDs were derived with DTAF. The high-sensitivity MEKC-LIF method was successfully established and applied to determine PDs in commercial hair colors for quality control and in real hair samples for evaluating the location of PDs in dyed hair samples, as well as in percutaneous absorption samples for evaluating the ability of PDs to penetrate skin. Keywords: 5-(4, 6-dichlorotriazinyl)aminofluorescein, Hair colors, Micellar electrokinetic chromatography, Percutaneous absorption, Phenylenediamines

Reactive Oxygen Species (ROS) are an important part of the normal cell growth cycle and play essential roles in many biological functions. Many techniques have been developed to detect and measure ...

Corynebacterium glutamicum (C. glutamicum) is a well-known workhorse for the industrial production of amino acids. Different carbon, nitrogen, and sulfur source may force the bacterium to produce specific metabolites. In this work, a method of high-speed MEKC with LIF detection was developed to rapidly analyze the amino acid metabolites released by C. glutamicum, which is fed with different culture mediums. Corynebacterium glutamicum was cultured in microbial fuel cells to monitor its metabolism process and collect its metabolites. In the CE system, a microliter-scale sample reservoir was designed and applied to perform tiny volume sample injection. With the assistance of microwave, the derivatization time for amino acids with FITC was greatly shortened to 6 min. Under the optimized condition, the eight candidate amino acids of metabolites could be rapidly separated within 2 min. The whole analysis process for real samples, from sampling to determination, could be shortened to less than 10 min. The results showed that C. glutamicum could produce additional l-lysine and l-valine as the metabolites when fed with glucose and l-methionine, respectively. The method proved that culture mediums used to feed C. glutamicum had great effect on the bacterium's metabolites.

Micellar electrokinetic chromatography (MEKC) is a hybrid of chromatography and electrophoresis. The separation mechanism is based on the interaction of analytes and pseudo-stationary phase and also the differential mobility in the electric field. Such property of MEKC can be made good use for separation of proteins and peptides, which have high dimensionality. In the past years, either in the form of single dimensional or multi-dimensional separations, MEKC has demonstrated its excellent usability in biomolecule separations. Recently, as the development in CE-MS was revisited, there are also new developments in MEKC-MS hyphenation techniques, which further points to the usefulness of MEKC in analysis of proteins and peptides. In this article, the recent advances of MEKC applied in separation and analysis of proteins were reviewed and the prospect of this technique was anticipated.

The main aim of this study was to apply capillary electrophoresis (CE) to the discrimination of red lipsticks – the first step towards developing the methodology for forensic purposes. The separation method was optimized using a mixture of eight dyes and micellar electrokinetic capillary chromatography (MEKC). The influence of the buffer pH and the concentration of SDS on the separation of dye mixtures was investigated. The best separation (highly efficient with the large resolution capacity in relatively short time) was accomplished using a buffer at pH 9 enriched with 80 mM SDS. The developed separation method was evaluated, determining the stability of sample (5 h) and buffer (10 h), the limit of detection of dyes (LOD In order to develop a versatile extraction method for compounds from red lipsticks, various types of solvents, temperatures and durations of ultrasound assisted extraction (UAE) were tested. The use of a specially designed and constructed device for repetitive sampling without contamination was suggested. The extraction method was characterized in terms of precision, obtaining very good results for the qualitative (RSD of migration time The proposed UAE/MEKC-DAD method was successfully applied to distinguish red lipstick samples with a very similar hue, achieving a high value of discrimination power (DP = 0.998). It was found that it has great potential in forensic investigations.

A facile micellar electrokinetic chromatography (MEKC) method was developed for quantification of aromatic amino acids in Cordyceps fungi. The proposed MEKC method had linear calibration curves and detection limits at the μmol/L level. Contents of aromatic amino acids in Cordyceps were found in the range from 0.004 9 % to 0.028 0% for tryptophan (Trp), 0.036 6% to 0.078 1% for tyrosine (Tyr), and 0.029 1% to 0.136 2% for phenylalanine (Phe). Levels of amino acids in cultivated Cordyceps militaris were found higher than those in natural Cordyceps sinensis. Interestingly, the ratio of Tyr to its precursor, Phe, in C. sinensis (1.20 ± 0.091, n=3) was significantly higher than that in cultivated C. militaris (0.54 ± 0.170, n=3). This is likely because the enzyme-catalyzed in vivo biotransformation of Phe to Tyr is much more effective in natural C. sinensis, a biological hybrid of larva and parasitic fungus, than in cultivated C. militaris.

A new, sensitive and simple field amplified sample injection (FASI) combined with micellar electrokinetic chromatography (MEKC) was developed for the analysis of terazosin (TER), alfuzosin (ALF), tamsulosin (TAM), doxazosin (DOX) and silodosin (SIL). The separation was achieved with Tris buffer (30 mM, pH 9.2) containing 30 mM SDS and 8% butanol, applying a voltage of 28 kV at 25 °C. Under the optimal conditions, the sensitivity of FASI-MEKC was improved 25.3–58.7 times as compared with the direct MEKC. The method was linear over concentration ranges of 50.0–2000.0 ng/mL for TER, DOX and SIL and 40.0–1200.0 ng/mL for ALF and TAM, with limits of detection of 9.04, 9.75, 5.21, 3.06 and 8.29 ng/mL in a respective order. The validated method was successfully applied for analysis of the five alpha blockers in dosage forms with good repeatability and recovery. Furthermore, the content uniformity testing of terazosin in its tablets as a model example was performed. The new method was also applied successfully to spiked human plasma which proved promising for the detection of these drugs in real samples.

An in-capillary 2,2-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid)-sweeping micellar electrokinetic chromatography-diode array detector (ABTS+-sweeping MEKC-DAD) method was developed and successfully applied to screening and quantifying antioxidative ingredients from natural products. The parameters affecting sweeping and separation were optimized including components of background electrolyte and sample matrix. Comparing with previously reported MEKC, the sensitivity enhancement factors of trace natural antioxidants obtained by this proposed method were from 17 to 167. The limit of detection was as low as 6 ng·mL−1. The results of other validation parameters including linearity, reproducibility, accuracy and recovery were satisfactory. Seven compounds including schizandrin, schisandrol B, schisantherin B, schisantherin A, schisanhenol, deoxyschizandrin, schisandrin B were identified as the main antioxidants in Schisandra chinensis. It was demonstrated that this developed in-capillary ABTS+-sweeping MEKC-DAD is simple, sensitive, reliable and rapid method for screening and quantifying trace antioxidants from natural products.

Microbial spoilage was the primary cause of food deterioration and caused huge economic losses and serious public health problems. The discovery of quorum sensing (QS) system got a new research orientation for the control of food spoilage. A fast, simple, and efficient method of solvent-terminated dispersive liquid-liquid microextraction (ST-DLLME) combined with micellar electrokinetic capillary chromatography was developed for the determination of N-acylhomoserine lactones from spoilage bacteria and aquatic product. In the procedure, the emulsion was formed by rapidly injecting a mixture of extraction solvent (carbon tetrachloride) and dispersive solvent (acetonitrile) rapidly into the aqueous sample solution. The acetonitrile was quickly injected into the emulsion as demulsification solvent, then the emulsion turned clear and became two phases. The last, the lower layer (carbon tetrachloride) as sample was collected and analyzed by capillary electrophoresis. Important parameters affecting the extraction efficiency of ST-DLLME were investigated in detail. Under the optimized conditions, the proposed method provided low limits of detection (below 5 ng.mL−1), a good linearity range of 5–1000 ng. mL−1, and good repeatability of the extractions (RSDs below 5.8%, n = 6). This proposed method was utilized to analyze the free extract of Pseudomonas aeruginosa CV12472, Aeromonas hydrophila PA01, fish and shrimp and obtained satisfactory results. The results indicated that the proposed method was successfully applied to the extraction of the routine analysis in the food field.

A simple and validated micellar electrokinetic chromatographic method is described for the determination of 5-hydroxymethylfurfural (HMF). The method was applied to fresh and processed (heated at 65 °C, 85 °C and 105 °C for 5 h and 20 h) multifloral and honeydew honeys. Optimum conditions were: 10 mmol L−1 Tris buffer consisting 110 mmol L−1 SDS and 10% (v/v) MeOH at pH 9.10, 20 kV of applied potential, 3 s of injection time at 5 × 103 N m−2, 280 nm of wavelength and 25 °C of fixed temperature. Mean electrophoretic mobility (m2 s−1 V−1) of HMF and methyl paraben (IS) were − 5.28 × 10−5 (RSD of 0.745%) and − 2.52 × 10−4 (RSD of 0.944%), respectively. The calibration curve was linear in the range of 0.68 µg mL−1 (5.40 × 10−6 mol L−1) and 4.05 µg mL−1 (3.21 × 10−5 mol L−1) with R = 0.9990 for inter-day precision. LOD and LOQ values were 0.007 µg mL−1 (5.48 × 10−8 mol L−1) and 0.023 µg mL−1 (1.83 × 10−7 mol L−1), respectively, as inter-day precision. The accuracy was very high with recovery values between 94.07 and 97.16%. The developed method was fully validated according to ICH guidelines. The results were compared with those of HPLC method adapted from AOAC method no: 980.23.

An optimized and economical capillary electrophoretic method for both off-line and on-line study of the enzyme aldehyde oxidase and its substrate phthalazine was developed. The separation of the substrate phthalazine and its metabolite 1-phthalazinone was achieved using micellar electrokinetic chromatography (MEKC) with sodium dodecyl sulphate in the background electrolyte (BGE). The BGE consists of 25 mM sodium phosphate buffer containing 50 mM sodium dodecyl sulphate at pH 7.4. A bare-fused-silica capillary with a capillary length of 40 cm, 50 μm ID and effective length of 30 cm was used to develop the capillary electrophoresis method. Improved separation conditions were elaborated and the separation method was validated based on the ICH and EMA guidelines. The limit of detection for phthalazine and 1-phthalazinone was 8 μM and 3 μM, respectively. The limit of quantification was 25 μM for phthalazine and 10 μM for 1-phthalazinone. The linearity of the detector response was checked for 1-phthalazinone at nine different concentrations in the range 10–500 μM and the determination coefficient was 0.9994. Accuracy was tested by comparing the corrected peak area of 1-phthalazinone reference solution at 20 μM and 50 μM with the corrected peak area of 20 μM and 50 μM 1-phthalazinone in the presence of human liver cytosol (HLC). Accuracy values of +5.3% and -2.5% were obtained at 20 μM and 50 μM, respectively. The on-line enzymatic reaction was successful with the application of the method of transverse diffusion of laminar flow profiles (TDLFP), which enables the mixing as well as separation of the enzyme and substrate inside the nanoliter-scale capillary. TDLFP is examined to be precise when performing 5 consecutive injections, with a relative standard deviation of 7.16% which is within the limitation of EMA standards. This miniaturized and low-cost incubation and separation method could be further introduced into industry and extended to other substrates.