Microemulsion Electrokinetic Capillary Chromatography Research Articles

Development of the Microemulsion Electrokinetic Capillary Chromatography Method for the Analysis of Disperse Dyes Extracted from Polyester Fibers.

The study aimed to develop a method for the separation of dispersed dyes extracted from polyester fibers. Nine commercially available disperse dyes, which were used to dye three polyester fabrics, were tested. Extraction of dyes from 1 cm long threads was carried out in chlorobenzene at 100 °C for 6 h. The separation was performed using microemulsion electrokinetic capillary chromatography (MEEKC) with photodiode array detection. Microemulsion based on a borate buffer with an organic phase of n-octane and butanol and a mixture of surfactants, sodium dodecyl sulphate and sodium cholate, were used. The addition of isopropanol and cyclodextrins to microemulsion resulted in a notable improvement in resolution and selectivity. The content of additives was optimized by using the Doehlert experimental design. Values of the coefficient of variance obtained in the validation process, illustrating the repeatability and intermediate precision of the migration times fit in the range of 0.11–1.24% and 0.58–3.21%, respectively. The developed method was also successfully applied to the differentiation of 28 real samples—polyester threads collected from clothing. The obtained results confirmed that proposed method may be used in the discriminant analysis of polyesters dying by disperse dyes and is promisingly employable in forensic practice.

Nanoemulsion supported microemulsion electrokinetic chromatography coupled with selected preconcentration techniques as an approach for analysis of highly hydrophobic compounds

In this paper, an oil-in-water (O/W) nanoemulsion (NE) prepared by water cold dilution of an O/W microemulsion (ME) was introduced as a sample matrix in microemulsion electrokinetic capillary chromatography (MEEKC) for the highly hydrophobic compounds analysis. Several model compounds with log PO/W values in the 4.1–10.9 range, from different chemical groups, including retinol, α-tocopherol, cholecalciferol, phylloquinone, menaquinone-7, dichlorodiphenyltrichloroethane, ivermectin have been tested. As a proof of the concept of NE formation, a dynamic light scattering technique was employed to determine the size distribution profile of NE particles. Moreover, due to relatively low conductivity of the NE matrix (50–100 times lower in comparison to the separation buffer) and a negative electric charge provided to hydrophobic compounds through NE dispersed phase, NE matrices have been combined with preconcentration techniques based on electrokinetic dosing, namely field amplified sample injection (FASI) and pressure assisted electrokinetic injection (PAEKI). The detection limits for vitamin K1 and K2-MK7 in the NE matrix in combination with FASI (NE–MEEKC–FASI) as well as PAEKI (NE–MEEKC–PAEKI) were up to 42.9 and 12.1 ng mL−1, respectively. In comparison to standard hydrodynamic injection for microemulsion sample matrix NE–MEEKC–PAEKI grant 45-fold improvement in signal sensitivity. The study presents an innovative approach, as it enables the use of preconcentration techniques for highly hydrophobic compounds (log PO/W > 4), which was not previously possible for implementation in the electromigration techniques. Likewise, the use of organic solvents has been reduced by using ME as a solvent for stock solutions and diluting with water prior to the analysis. The application to real samples was investigated using a dietary supplement containing vitamin K2-MK7 obtained from the fermentation product of soybeans.

The examination of red lipsticks using microemulsion electrokinetic capillary chromatography

Lipsticks, due to their easy transfer and widespread use, can provide important evidence in crime investigations, in particular in cases of rape, murder or burglary. Discrimination between and identification of lipstick samples is a challenge because of their similar chemical composition. The main components of lipsticks – oils and waxes – are a difficult matrix to analyse. Furthermore, lipsticks contain various additional compounds, both ionic and neutral. A promising approach to the examination of lipstick traces is the use of microemulsion electrokinetic capillary chromatography (MEEKC). In order to optimize the MEEKC method, a series of mixtures of dye standards in various microemulsions (MEs) (differed in qualitative and quantitative composition) and measurement conditions were analysed. The influence of the type and pH of the buffer, the type and amount of surfactant and oil phase, as well as the effect of the addition of a second surfactant, organic solvent and cyclodextrin, were investigated. The ME with the best separation properties contained 3% SDS, 6% butanol and 0.8% n-octane (w/w) in a borate buffer at pH 10. Short-end mode enabled the separation to be reduced to less than 4 min. Ultrasound-assisted extraction (UAE) performed for 10 min at 25 °C was found to be the most effective for real lipstick samples preparation. The developed UAE-MEEKC method was verified in terms of precision (RSDtm for real samples analysed in one day was < 2.2%, and over three days was < 3.0%) and successfully used for the discrimination of six red lipsticks.

The microemulsion electrokinetic capillary chromatography combined with reversed-electrode polarity stacking mode for enriching and quantifying lignanoids and ginsenosides in TCMs preparation Shengmai injection.

An on-line large volume sample stacking with polarity switching (LVSS) method was proposed for simultaneously determining lignanoids and ginsenosides in MEEKC. The parameters including the pH value and concentration of buffer solution, SDS, organic modifier, oil phase, running voltage, and temperature as well as injection time, sample matrix, stacking voltage, and time influencing separation and stacking were systematically optimized. The method was verified by performing precision, accuracy, stability, and recovery. Its reliability was proved by separating and quantifying two lignanoids and three ginsenosides in Shengmai injectionSMI. The sensitivity of these compounds was improved by MEEKC-LVSS method for 6-11 times than conventional MEEKC. Thus, this developed on-line MEEKC-LVSS method was sensitive, practical, and reliable.

Determination of food colorants in a wide variety of food matrices by microemulsion electrokinetic capillary chromatography. Considerations on the found concentrations and regulated consumption limits

Color additives are used widely by the food industry to confer a desirable appearance. Some of the most used colorants (Tartrazine (E102), Sunset Yellow (E110), Red Allure (E129) and Blue Brilliant (E133)) were determined in this study using microemulsion electrokinetic capillary chromatography (MEEKC). Regression coefficients were greater than 0.9981; intra- and inter-day precisions, in terms of percentage RSD, were less than 7.01% and 8.55%, respectively; recoveries were between 90 and 100% in most cases. LODs and LOQs ranged from 0.24 to 1.21 mg L−1 and from 0.80 to 4.03 mg L−1, respectively. Moreover, MEEKC consumed less solvent than HPLC, making the analysis more environmentally friendly. The proposed method is suitable for the determination of colorants in a wide variety of foods. Results showed that consumers should be aware of colorants to avoid consumption exceeding recommended amounts.

Rapid Analysis of Alpinetin and Cardamonin in Alpinia katsumadai Hayata Using Short-End Injection Microemulsion Electrokinetic Capillary Chromatography Combined with Microwave-Assisted Extraction

A rapid and simple short-end injection microemulsion electrokinetic chromatography (MEEKC) method was developed for the routine analysis of alpinetin and cardamonin in Alpinia katsumadai Hayata in combination with microwave-assisted extraction (MAE). The key parameters affecting separation and extraction were investigated and optimized. The optimal MAE was performed at 800 W for 12 min with 100% ethanol as the extraction solvent, which gave a better extraction efficiency and required a significantly shorter time than conventional heat reflux extraction (HRE) and ultrasonic extraction (USE). When this detection system based on ethyl acetate oil-based MEEKC and short-end injection was applied, the analysis was completed in less than 1.5 min. In the method validation, the relative standard deviations of the migration times and peak areas of the two constituents were 0.74, 1.86% for alpinetin and 0.63, 2.05% for cardamonin, respectively. The recoveries ranged from 94.6 to 105.8%. Compared with previously reported methods, the present method offers a dramatic decrease in overall analysis time, and has the further advantages of easy operation and low running costs.

Analysis of nucleosides and nucleobases by microemulsion electrokinetic capillary chromatography coupled with field-amplified sample injection

A microemulsion electrokinetic capillary (MEEKC) chromatography method was online-coupled with field-amplified sample injection (FASI) for the analysis of nucleosides and nucleobases.

Separation of steroids using vegetable oils in microemulsion electrokinetic capillary chromatography

The steroids, hydrocortisone, androstenedione, 17-α-hydroxyprogesterone, testosterone, 17-α-methyltestosterone, and progesterone were separated with microemulsion electrokinetic chromatography (MEEKC) and detected with UV absorption. The microemulsion phases were prepared from both artificial and vegetable oils, from them the first was made of alkane and alcohol and the latter from colza, olive, linseed, and walnut oils. The electrolyte solutions were made to emulsions using sodium dodecyl sulfate and alkaline tetraborate. The solution mixtures made from ethyl acetate, sodium dodecyl sulfate, 1-butanol, acetonitrile, and sodium tetraborate were used as the reference solutions to evaluate the performance of the vegetable oil emulsions. Our study showed that the lipophilic organic phase in the microemulsion did provide resolution improvements but not selectivity changes. The results also correlate with real interactions of the steroids with the lipophilic organic microemulsion phase. The quality of the oils between the manufacturers did not have importance, which was noticed from the equal behavior of the steroids in the vegetable oil emulsions. Detection limits of the steroids in vegetable oil emulsions were at the level of 0.20-0.43μg/L. Thus, they were 2-10 times higher than the concentrations in the partial filling micellar electrokinetic chromatography (PF-MEKC), which we have obtained earlier. The repeatability (RSD%) of the electrophoretic mobilities of the steroids was between 0.50 and 3.70. The RSD% values between the inter-day separations were below 1%, but when walnut and olive oils were used the values exceeded even 10%.

Analysis of plant hormones by microemulsion electrokinetic capillary chromatography coupled with on-line large volume sample stacking

A novel method of microemulsion electrokinetic capillary chromatography (MEEKC) coupled with on-line large volume sample stacking was developed for the analysis of six plant hormones including indole-3-acetic acid, indole-3-butyric acid, indole-3-propionic acid, 1-naphthaleneacetic acid, abscisic acid and salicylic acid. Baseline separation of six plant hormones was achieved within 10 min by using the microemulsion background electrolyte containing a 97.2% (w/w) 10 mM borate buffer at pH 9.2, 1.0% (w/w) ethyl acetate as oil droplets, 0.6% (w/w) sodium dodecyl sulphate as surfactant and 1.2% (w/w) 1-butanol as cosurfactant. In addition, an on-line concentration method based on a large volume sample stacking technique and multiple wavelength detection was adopted for improving the detection sensitivity in order to determine trace level hormones in a real sample. The optimal method provided about 50-100 fold increase in detection sensitivity compared with a single MEEKC method, and the detection limits (S/N = 3) were between 0.005 and 0.02 μg mL(-1). The proposed method was simple, rapid and sensitive and could be applied to the determination of six plant hormones in spiked water samples, tobacco leaves and 1-naphthylacetic acid in leaf fertilizer. The recoveries ranged from 76.0% to 119.1%, and good reproducibilities were obtained with relative standard deviations (RSDs) less than 6.6%.

Separation of Steroids and the Determination of Estradiol Content in Transdermic Patches by Microemulsion Electrokinetic Chromatography

A novel microemulsion electrokinetic capillary chromatography (MEEKC) method has been developed which separates a range of nine steroids. A microemulsion containing ethyl acetate, butan-1-ol, sodium dodecyl sulfate, 15% (v/v) acetonitrile and 12 mmol L−1 sodium tetraborate aqueous buffer at pH 9.2 was used with direct UV detection at 200 nm. The method was validated for the determination of 17β-estradiol content, a hormone steroid, in transdermal patches. Adequate sensitivity (DL = 0.88 μg mL−1; QL = 2.65 μg mL−1) without interference from sample excipients was obtained. 17β-Estradiol migrates in approximately 5.4 min. Estrone was used as internal standard and acceptable precision (<1.2% RSD), linearity (r = 0.9996; range from 40.0 to 60.0 μg mL−1), and recovery (100.4 ± 0.9% at three concentration levels) were obtained. The principal advantage of the method is that it is rapid and avoids the need of time consuming and expensive sample pre-treatment steps.

Sample stacking microemulsion electrokinetic capillary chromatography induced by reverse migrating pseudostationary phase for the quantification of phenobarbital and its p-hydroxyphenobarbital metabolite in rat urine

For the first time, a capillary electrophoretic (CE) method with sample stacking induced by a reverse migrating pseudostationary phase (SRMP) technique has been developed and validated for sensitive determination of phenobarbital (PB) and its p-hydroxyphenobarbital (PHPB) metabolite in rat urine samples. Separation and determination were optimized on a fused-silica capillary with a total length of 50 cm (effective length 40 cm) and 75 μm ID. The microemulsion background electrolyte consisted of 0.8% (v/v) ethyl acetate, 6.6% (v/v) butan-2-ol, 1.0% (v/v) acetonitrile, 2.0% (w/v) sodium n-dodecyl sulfate (SDS), and 89.6% (v/v) of 7.5 mM ammonium formate at pH 8. When this preconcentration technique was used, the sample stacking and the separation processes took place successively with changing the voltage with an intermediate polarity switching step. For practical application, a solid-phase extraction (SPE), C(18) sorbent with n-hexane/ethyl acetate (1 : 1%, v/v) as the elution solvent was used for sample purification and concentration. The SPE method gave good extraction yields for all the analytes, with absolute recovery values of 96.9% and 99.1% for PB and PHPB, respectively. The regression equations for PB and PHPB showed excellent linearity over a concentration range of 55-1386 ng mL(-1) for PB and PHPB (r = 0.998). The developed microemulsion electrokinetic capillary chromatography (MEEKC) method for separation of the studied compounds with SRMP as the electrophoretic preconcentration technique allowed detection limits in urine samples at 16.8 ng mL(-1) for PB and PHPB which are 15-fold lower than the reported CE method in the literature. The precision results, expressed by the intra-day and inter-day relative standard deviation (RSD) values range from 3.6 to 7.1% (repeatability) and from 3.2 to 7.2% (intermediate precision) for PB and PHPB, respectively, which were in line with Food and Drug Administration (FDA) criteria.

Rapid Determination of Water-Soluble and Fat-Soluble Vitamins in Commercial Formulations by MEEKC

Microemulsion electrokinetic capillary chromatography has been successfully applied to the separation and determination of water-soluble vitamins (thiamine hydrochloride, riboflavin, niacin, pyridoxine hydrochloride, folic acid, cobalamin, ascorbic acid) and a fat-soluble vitamin (α-tocopherol acetate). The optimal microemulsion buffer contained sodium dodecylsulfate (SDS) as surfactant, butan-1-ol as the co-surfactant, ethyl acetate as the oil and pH 9.2 tetraborate buffer, modified with 15% (v/v) 2-propanol. UV detection at 214 nm gave adequate sensitivity without interference from sample excipients. Under the optimized conditions, the vitamins were baseline separated in less than 7 min. Analytical curves of peak area versus concentration presented coefficients of determination (R 2 ) > 0.99, acceptable limits of quantification between 8.40 and 16.23 μg mL−1 were obtained. Vitamin levels in liquid formulation were quantified with intra-day precision better than 0.99% RSD for migration time and 1.19% RSD for peak area ratio. Recoveries ranged between 98.7 and 101.7%. The method was considered appropriate for rapid and routine analysis.

Novel, selective sample stacking microemulsion electrokinetic capillary chromatography induced by reverse migrating pseudostationary phase for the determination of the new ultra-short acting hypnotic “HIE-124” in mice serum

Microemulsion electrokinetic capillary chromatography (MEEKC) with sample stacking induced by reverse migrating pseudostationary phase (SRMP) technique in a suppressed electro-osmotic flow (EOF) strategy was investigated for analysing the new ultra-short hypnotic HIE-124 in mice serum. The proposed method utilized fused-silica capillary with a total length of 50 cm (effective length 40 cm), applied voltages for stacking and separation were 5.0 kV for 4.30 min and subsequently 25 kV, respectively, with a sample injection of 0.5 psi for 90 s. All the runs were carried out at 25 °C and detected at 213 nm. The optimum microemulsion background electrolyte (BGE) solution consisted of 0.8% (v/v) ethyl acetate, 6.6% (v/v) butan-2-ol, 1.0% (v/v) acetonitrile, 2.0% (w/v) sodium dodecyl sulfate (SDS), and 89.6 mL with 25 mM phosphate buffer pH 8. When this preconcentration technique was used, the sample stacking and the separation processes took place successively with changing the voltage with an intermediate polarity switching step. The proposed method was validated carefully with respect to high specificity of the method, good linearity ( r = 0.9994), fair wide linear concentration range (66–1500 ng mL −1), limit of detection and quantitation were 21.6 and 65.5 ng mL −1, respectively. The mean relative standard deviation (RSD) of the results of intra- and inter-day precision and accuracy were less than 6.0%, and overall recovery higher than 95% of HIE-124 in mice serum. The developed method could be used for the trace analyses of HIE-124 in serum and was finally used for the pharmacokinetic study investigation of HIE-124 in mice serum.

Analysis of Nonsteroidal Anti-inflammatory Drugs in Water Samples Using Microemulsion Electrokinetic Capillary Chromatography Under pH-Suppressed Electroosmotic Flow with an On-Column Preconcentration Technique

Microemulsion electrokinetic capillary chromatography (MEEKC) in a suppressed electroosmotic flow (EOF) strategy was investigated for analysing a group of nonsteroidal anti-inflammatory drugs (NSAIDs) in water samples. The EOF was effectively suppressed with an acidic buffer as the aqueous phase. Four water immiscible solvents, oils (n-heptane, n-octane, ethyl acetate and di-n-butyl tartrate) and three organic solvents (methanol, 2-propanol and acetonitrile) were tested to optimise the pseudostationary phase and to obtain efficient separations. The optimum microemulsion background electrolyte (BGE) solution consisted of 0.8% (w/w) n-heptane, 6.6 (w/w) butan-1-ol, 15.0% (w/w) acetonitrile, 3.3% (w/w) sodium dodecyl sulfate (SDS), and 74.3% (w/w) of 25 mM sodium phosphate at pH 2.5. Stacking with reverse migrating pseudostationary phase (SRMP) was applied to enhance the concentration sensitivity of the NSAIDs. When this preconcentration technique was used, the sample stacking and the separation processes took place successively with the same voltage without an intermediate polarity switching step. Detection limits (LODs) in the order of 5–15 µg L−1 for the NSAIDs were obtained using SRMP for standard solutions. The developed method was validated for the analysis of NSAIDs in tap water samples by combining an off-line solid-phase extraction (SPE) step and the on-column preconcentration technique SRMP. The LODs were in the 100–230 ng L−1 range.

Sample stacking for the analysis of penicillins by microemulsion electrokinetic capillary chromatography

We present a method for determining eight penicillin antibiotics using microemulsion electrokinetic chromatography (MEEKC). We studied how the composition of the microemulsion affected separation by modifying such parameters as the surfactant or the addition of organic solvents. The best microemulsion system consisted of 0.5% ethyl acetate, 1.2% 1-butanol, 2% Brij 35, 10% 2-butanol and 86.3% 10 mM borate buffer at pH 10. We studied the suitability of this microemulsion composition for analyzing a commercial drug. To improve the sensitivity of the method, we used the stacking technique reversed electrode polarity stacking mode (REPSM), which increased the detection limits by about 40-fold.

Rapid Determination of Diterpenoids in Andrographis paniculata by Microemulsion Electrokinetic Capillary Chromatography with Short-End Injection

A rapid, simple, and reliable method has been developed for routine capillary electrophoretic analysis of two diterpenoids, andrographolide and dehydroandrographolide, in Andrographis paniculata. For this system, using ethyl acetate oil-based microemulsion electrokinetic chromatography (MEEKC) and the short-end injection technique, analysis was complete in less than 2.5 min. In method validation the relative standard deviations of migration time and peak area of the two constituents were, respectively, 0.54% and 1.70% for andrographolide and 0.45% and 2.11% for dehydroandrographolide. Regression equations revealed linear relationships (correlation coefficients 0.9994 for andrographolide and 0.9993 for dehydroandrographolide) between peak area and concentration. The effects of buffer pH, borate concentration, SDS concentration, co-surfactant type and concentration, injection time, and running potential were systematically investigated. The method can be successfully implemented in routine quality-control testing.

Development of a capillary electrophoretic method for the separation of diastereoisomers of a new human immunodeficiency virus protease inhibitor

A capillary electrophoretic (CE) method was developed for the separation of diastereoisomers of a new human immunodeficiency virus (HIV) protease inhibitor TMC114. In total 16 isomers of this drug have been synthesized (eight pairs of enantiomers). We succeeded in the separation of the eight diastereoisomers, but no enantiomers could be separated. Because of the high similarity and water-insolubility of these isomers, the separation is a real challenge. Different CE modes were tried out: capillary zone electrophoresis (CZE), nonaqueous capillary electrophoresis (NACE), micellar electrokinetic capillary chromatography (MEKC), and microemulsion electrokinetic capillary chromatography (MEEKC). Only MEEKC offered resolution of these compounds.

Electrokinetic capillary chromatography in a polar continuous-phase water-in-oil microemulsion constituted by water, sodium dodecyl sulfate, and n-pentanol.

A water-in-oil (w/o) microemulsion (ME) constituted by 15% Tris buffer, pH 8.4, in water and 85% sodium dodecyl sulfate (SDS)/n-pentanol 1:4 mixture, capable of dissolving up to 30% vegetable oils and lard, was used as background electrolyte in reverse microemulsion electrokinetic capillary chromatography (RMEEKC). Owing to the free SDS ions in the continuous phase and some degree of percolation, the ME showed a high conductivity (0.65 mS. cm(-1) at 25 degrees C) and sustained a very stable capillary current. Previous rinsing of the capillary with a quaternary ammonium salt for electroosmotic flow (EOF) reduction, a series of nonionic and anionic solutes dissolved either in the ME or in fat samples diluted with the ME (1:4 ratio), were injected. Using -20 kV, fair separations of the solutes in the migration time order singly charged anions < nonionic solutes < doubly charged anions approximately pyromellitate were obtained, salicylate (I) showing by far the shortest migration time, and phthalate (II) and pyromellitate the longest. Separation was attributed to partition between the aqueous droplets, where pyromellitate and II were assumed to be trapped, and the n-pentanol continuous phase, where the mobilitites could be also modified by association of the solutes with SDS ions. Adequate EOF markers were not found, thus the relative mobility of any solute with respect to the mobility of the droplets, mu(r), was expressed as a fraction of the mobility of I with respect to that of the droplets, being mu(r) = (t(II) - t(R)) t(I) / [(t(II) - t(I)) t(R)], where t(R), t(I), and t(II) are the migration times of the solutes I and II, respectively. The application of RMEEKC to the analysis of both hydrophilic and hydrophobic samples, including edible fats, was demonstrated.

A comparative study of microemulsion electrokinetic capillary chromatography and micellar electrokinetic capillary chromatography for the analysis of UV‐absorbing compounds in human urine

Separations of human urine by microemulsion electrokinetic chromatography (MEEKC) and micellar electrokinetic capillary chromatography (MEKC) with respect to resolution, migration times and efficiencies were optimized and compared. The optimised MEEKC and MEKC methods were simple and fast, both of which are excellent characteristics for the complex separations required in clinical and biomedical studies. However, resolution in MEKC was significantly greater than in MEEKC although migration times were 30% faster for the optimised MEEKC method. In addition, a faster analysis method (short-end injection) specifically for routine screening purposes was also investigated. With both MEEKC and MEKC modes, this provided short separations (less than 4 min for urine) with no major compromise in resolution. In conclusion, we found that MEEKC offered no real advantage over MEKC for urine analysis.

Comparison of Different Capillary Electrophoresis Modes and HPLC for the Separation of Xanthones

Nine xanthones from a Chinese traditional medicine, Securidaca inappendiculata Hassk, were separated by different capillary electrophoresis (CE) modes, including capillary zone electrophoresis (CZE), micellar electrokinetic chromatography (MEKC), microemulsion electrokinetic capillary chromatography (MEEKC), and capillary electrochromatography (CEC). The comparison of separation selectivity and efficiency for xanthones by using these CE methods indicated that different CE modes greatly varied in the separation selectivity and efficiency for xanthones; their advantages and disadvantages are discussed. Moreover, compared with traditional chromatographic methods, e.g., high performance liquid chromatography (HPLC), CE shows higher separation ability and analytical speed for these xanthones.