New bioanalytical microemulsion Electrokinetic chromatography method for the simultaneous determination of Trifluridine with its metabolites and Tipiracil in rat plasma: Application to pharmacokinetic studies.

A Microemulsion Electrokinetic Chromatography method coupled with diode array detector (MEEKC-DAD) was developed for the first time and found to be efficient, sensitive, and selective for the simultaneous analysis of Trifluridine (FTD), and its metabolites 5-(trifluoromethyl) uracil (FTY) and 5-carboxy-2'-deoxyuridine (5CDU), and Tipiracil (TIP) in rat plasma. Sample pre-treatment involved a simple protein precipitation from plasma using acetonitrile. The separation was achieved using a fused silica capillary (65 cm total length, 55 cm effective length and 50 µm i.d.) and a microemulsion solution consisted of 1.66% sodium dodecyl sulfate (SDS), 0.91% heptane, 6.61% 1-butanol, and 90.72% borate buffer (20 mM, pH 9.5). Electrophoretic separation was carried out at 20 °C and 20 kV. The samples were injected for 40 s at 20 mbar and detected simultaneously at 205 nm. The electrophoretic parameters indicated that the developed MEEKC-DAD method permitted complete resolution of the analytes within 13 min. The developed method was fully validated according to the FDA guidelines for bioanalytical method validation. The method was linear in the range 200-4000 ng/ml for FTD, FTY, 5CDU, and 100-1000 ng/ml for TIP. The intra/inter-day accuracy and precisions were ≤4% for all drugs. Extraction recovery and stability were also assessed and were within acceptable range. After being validated, the method was applied for the determination of the studied drugs in plasma samples collected from rats injected intraperitoneally with a combination of FTD and TIP. The results obtained were used to study the pharmacokinetics of FTD with its metabolite and TIP in rat plasma.

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 chlorophenols in water using dispersive liquid-liquid microextraction coupled with water-in-oil microemulsion electrokinetic chromatography in normal stacking mode.

The current routes to couple dispersive liquid-liquid microextraction with capillary electrophoresis are the evaporation of water immiscible extractants and the back-extraction of analytes. In this study, a new methodology for this combination using water-in-oil microemulsion electrokinetic chromatography coupled with normal stacking mode on-line sample concentration was developed to analyze chlorophenols in water samples. The analytes were extracted with tributyl phosphate and the extractant dilution (3×) was directly injected into an electrophoresis buffer (7.7 cm) containing 5% sodium dodecyl sulfate, 78% 1-butanol, 2% 1-heptane, and 15% sodium acetate solution (pH 8.0). This proposed method is very simple and convenient compared to the conventional procedures. The key parameters affecting separation and concentration were systematically optimized. Under the optimized conditions, dispersive liquid-liquid microextraction contributed an enrichment factor of 45-50, and the overall sensitivity improvement was 312-418-fold. Limits of detection between 1.4 and 3.0 ng/mL and limits of quantification between 4.5 and 10.2 ng/mL were achieved. Acceptable repeatability lower than 3.0% for migration time and 9.0% for peak areas were obtained. The developed method was successfully applied for analysis of the chlorophenols in real water samples.

Sensitive enantioanalysis of ß-blockers via field-amplified sample injection combined with water removal in microemulsion electrokinetic chromatography.

In this study, an on-line sample preconcentration technique, field-amplified sample injection combined with water removal by electroosmotic flow (EOF) pump, was applied to realize a highly sensitive chiral analysis of ß-blocker enantiomers by MEEKC. The introduction of a water plug in capillary before the electrokinetic injection provided the effective preconcentration of chiral compounds. And then the water was moving out of the column from the injection end under the effect of the EOF, which avoided dilution of the stacked ß-blocker enantiomers concentration suffering from the presence of water in separation buffer. Moreover, the addition of H3 PO4 and methanol in the sample solution greatly improved the enhancement efficiency further. Under optimized conditions, more than 2700-fold enhancement in sensitivity was obtained for each enantiomer of bupranolol (BU), alprenolol (AL), and propranolol (PRO) via electrokinetic injection. LODs were 0.10, 0.10, 0.12, 0.11, 0.02, and 0.02 ng/mL for S-BU, R-BU, S-AL, R-AL, S-PRO, and R-PRO, respectively. Eventually, the proposed method was successfully applied to the determination of BU, AL, and PRO in serum samples with good recoveries ranging from 93.4 to 98.2%.

Determination of amino acids by microemulsion electrokinetic chromatography laser induced fluorescence method.

In this study, detection of 20 FITC-derivatized amino acids using an MEEKC-LIF was demonstrated. In order to achieve good separation for hydrophobic amino acids, the MEEKC method was employed and detection limits were obtained in the range of 0.32-2.2 nM, which is comparable to previous reports on amino acid analyses. Furthermore, a significant reduction in the reaction time from 1 h for conventional derivatization to 3 min for the microwave-assisted derivatization was observed and achieved, as opposed to the traditional pretreatment of real sample due to its complexity prior to the analysis of amino acid content. Finally, this microwave-assisted derivatization MEEKC-LIF method successfully determined amino acids in beverage, food, and biological samples (rat brain) with good recovery.

Quantification of the level of fat-soluble vitamins in feed based on the novel microemulsion electrokinetic chromatography (MEEKC) method.

BACKGROUND: Simultaneous quantification of liposoluble vitamins is not a new area of interest, since these compounds co-determine the nutritional quality of food and feed, a field widely explored in the human and animal diet. However, the development of appropriate methods is still a matter of concern, especially when the vitamin composition is highly complex, as is the case with feed designated for laboratory animals, representing a higher health and microbiological status. RESULTS: A method combining microemulsion electrokinetic chromatography (MEEKC) with liquid-liquid extraction was developed for the determination of four fat-soluble vitamins in animal feed. A separation medium consisting of 25 mmol L⁻¹ phosphate buffer (pH 2.5), 2-propanol, 1-butanol, sodium dodecyl sulfate and octane allowed the simultaneous determination of vitamins A, D, E and K within a reasonable time of 25 min. The polarity of the separation voltage was reversed in view of the strongly suppressed electro-osmotic flow, and the applied voltage was set at 12 kV. The fat-soluble vitamins were separated in the order of decreasing hydrophobicity. CONCLUSION: It was proved that the proposed MEEKC method was sufficiently specific and sensitive for screening fat-soluble vitamins in animal feed samples after their sterilization.

Separation and determination of phenolic compounds using novel deep eutectic solvent-in-water microemulsion electrokinetic chromatography.

BACKGROUND: Microemulsion electrokinetic chromatography (MEEKC) is a mode of capillary electrophoresis with a wide range of applications in which microemulsion is utilized as background electrolyte to achieve the separation of analytes. Microemulsions are composed of oil droplets, aqueous buffer, surfactant, and co-surfactant. Currently, conventional organic reagents act as the most commonly used oil phase in microemulsions, which are unfriendly to the environment. Recently, deep eutectic solvent (DES) has become a new type of eco-friendly solvent due to its non-toxicity. Therefore, it is of great value to establish a new MEEKC method by replacing conventional organic reagents as the oil phase with DES. RESULTS: The novel DES/W MEEKC method was established for phenolic compounds in Senecio scandens samples. Single-factor experiments and response surface methodology were performed to systematically optimize the crucial parameters for the method, including the type and content of the oil phase, surfactant content, concentration of borax buffer, and pH of the background solution. Under the optimized conditions, satisfactory regression curves were established for all standard analytes with correlation coefficients ≥0.9990. The method featured high sensitivity and favorable accuracy, with the instrumental detection limit in the range of 0.22-1.04 µg/mL, and intraday and interday precision for migration time expressed as relative standard deviations of 0.18-0.82% and 1.25-2.50%, respectively. The DES/W MEEKC method was successfully applied to Senecio scandens with good recoveries of 87.72-106.99%. In conclusion, the newly established DES/W MEEKC method is highly efficient, green and environmentally friendly. SIGNIFICANCE: DES is considered a green and efficient solvent. The DES/W MEEKC method is highly efficient and environmentally friendly. Actually, the method provides a novel and effective analytical tool for the simultaneous separation and determination of multiple phenolic compounds, especially in complex plant matrices. In the future, the DES/W MEEKC method still has the prospect of being widely used in the separation of other complex phytochemicals.

Microstructure of microemulsion modified with ionic liquids in microemulsion electrokinetic chromatography and analysis of seven corticosteroids.

In this work, the influences of ionic liquid (IL) as a modifier on microemulsion microstructure and separation performance in MEEKC were investigated. Experimental results showed that synergetic effect between IL 1-butyl-3-methylimidazolium tetrafluoro-borate (BmimBF4 ) and surfactant SDS gave a decreased CMC. With increment of IL in microemulsion, negative ζ potential of the microdroplets reduced gradually. The influence of IL on the dimensions of microdroplet was complicated. At BmimBF4 less than 8 mM, IL made microemulsion droplet smaller in size. While at BmimBF4 more than 10 mM, the size increased and reached to a maximum value at 12 mM, where the microdroplets were larger than that without IL. After that, the micreodroplet size decreased again. Relative fluorescence intensity of the first vibration band of pyrene to the third one (I1 /I3 ) enhanced as IL was added to microemulsion, which indicated that this addition increased environmental polarity in the inner core of microdroplets. Prednisone, hydrocortisone, prednisolone, hydrocortisone acetate, cortisone acetate, prednisolone acetate, and triamcinolone acetonide were analyzed with MEEKC modified with IL to evaluate the separation performance. Cortisone acetate and prednisolone acetate could not be separated at all in typical microemulsion. The seven analytes could be separated by the addition of 10 mM BmimBF4 into the microemulsion system. The method has been used for analysis of corticosteroids in cosmetic samples with simple extraction; the recoveries for seven analytes were between 86 and 114%. This method provides accuracy, reproducibility, pretreatment simplicity, and could be applied to the quality control of cosmetics.

Analysis of iodinated amino acids by microemulsion electrokinetic chromatography using aliphatic amine as a cosurfactant.

A novel microemulsion electrokinetic chromatography (MEEKC) was established for the separation and determination of iodinated amino acids using n-butylamine as a novel cosurfactant. By optimizing the type of oil phase, the type and concentration of surfactant, the concentration of cosurfactant and the type and concentration of buffer in the microemulsion system, the optimal conditions for the separation of organic iodines were determined to be 0.5% ethyl acetate, 0.6% SDS, 1.2% n-butylamine and 10 mM sodium borate. The efficient and rapid separation of the five analytes (3-iodo-L-tyrosine (MIT), 3, 5-Diiodo-L-tyrosine (DIT), 3, 5-Diiodo-L-thyronine (T2), 3, 3', 5-Triiodo-L-thyronine (T3) and L-Thyroxine (T4)) was achieved under the optimal conditions. The reliability of the method was verified by measuring the precision, LOD, LOQ and recovery. The practicality of the MEEKC method was demonstrated by applying it to the determination of two iodotyrosines, MIT and DIT, in kelp. The analytical method established in this experiment will provide a reference for the study of iodotyrosines in other natural plants and food.

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.

Versatile analytical methodology for evaluation of drug-like properties of potentially multi-targeting anticancer metallodrugs.

Using inductively coupled plasma mass spectrometry (in combination with ultrafiltration) and microemulsion electrokinetic chromatography, the drug properties of two new, potentially multi-targeting Ru(III) and Pt(IV) compounds, containing biologically active ligands, were evaluated. The ruthenium complex with bexarotene was shown to bind to albumin faster than to transferrin and exhibits much the same (to albumin) binding profile in human serum. The Pt(IV)-lonidamine complex interacts with albumin relatively slowly but possesses high stability and lipophilicity (log P 1.62), which makes it possible the cellular uptake in a free (of proteins) form. Although both examined compounds display a moderate solubility (below 10-4 M), this stands compatible with their nanomolar cytotoxic activities. The Ru(III) compound, whose active moiety is a complexed anion, is deemed promising to be loaded on nanoscale anion-exchangers with the aim of controlled delivery.

After enjoying curry: urine metabolism analysis of curcuminoids by microemulsion electrokinetic chromatography with laser-induced native fluorescence detection.

Considering pH-dependent fluorescence of curcuminoids, a microemulsion electrokinetic chromatographic (MEEKC) method was developed under acidic conditions for their separation and detection using laser-induced native fluorescence (LINF), so as to solve the analysis of urine metabolism for curcuminoids. The microemulsion composition was optimized by response surface methodology (RSM), and the effects of buffer pH and organic modifiers were systematically investigated. The optimal buffer for the separation of curcuminoids was chosen as follows: 2.8% (v/v) ethyl acetate, 80 mM SDS and 2.8% (v/v) n-butanol to form microemulsion, 28% (v/v) ethanol as organic modifier, and 20 mM phosphoric acid as electrolyte at pH 3.0. Under these conditions, four curcuminoids including curcumin, demethoxy curcumin (DMC), bisdemethoxy curcumin (BDMC) and demethyl curcumin (DEC) could be well separated within 18 min, and the detection limits (LOD, based on S/N=3) were calculated to be 71, 60, 22, and 147 pg mL-1, respectively. Combined with solid-phase extraction (SPE), the developed MEEKC-LINF method has been successfully applied to continuously monitor the curcuminoids and related metabolites in human urine collected from a healthy volunteer after oral administration of curry, testifying that this method has potential for evaluating the pharmacological activity of curcuminoids.

On-line concentration of triazine herbicides in microemulsion electrokinetic chromatography by electrokinetic injection assisted micelle to cyclodextrin stacking.

A fast, simple, environmentally friendly and sensitive on-line concentration method using microemulsion system as background solution (BGS) was developed for the trace detection of propazine, atrazine, simazine in food samples. The electrokinetic injection assisted micelle to cyclodextrin stacking (MCDS) was designed for the enrichment of target compounds. The factors affected enrichment performance, such as the kind of CDs, the amount of CDs, the concentration of methanol in BGS, the concentration of micelle in sample matrix, the concentration of phosphoric acid in BGS and the sample injection time were optimized. The optimized electrophoretic condition was obtained as following: 50 mM α-CD, 20 mM SDS in sample matrix., 80 mM PA and 20% MeOH (v/v) in BGS, sample solution by electrokinetic injection at -10 kV for 80 s. Under the optimized conditions described above, the linear range was 0.1-20 ug/mL with a good linear relationship with a correlation coefficient (r) ≥ 0.9985. The SEFs for the propazine, atrazine, simazine were found to be 123, 85 and 62 respectively. The proposed MCDS-MEEKC method provided an efficient method for trace analysis of triazine herbicides in honey and dendrobium officinale samples.

Determination of the retention factor of ionizable compounds in microemulsion electrokinetic chromatography.

Determination of the retention factor of ionized compounds in microemulsion electrokinetic chromatography requires two mobility measurements at the same pH: one in the presence of the microemulsion and another in plain buffer. However, it has been observed that in some cases subtracting one mobility from another determined in a different medium leads to negative retention factors, which makes no sense from a chemical point of view. This indicates that there is some error in the process which has a direct impact when retention factors are used for further applications. Here, we evaluate how the components of the microemulsion confer different properties to the buffer medium, particularly varying the viscosity parameter (which is inversely related to mobility). Whereas sodium dodecyl sulfate, the surfactant used in the microemulsion, has little effect on the medium viscosity (only an increase of 5%-6%), the presence of 1-butanol, used as a stabilizer, increases it by around 30%. Meanwhile, heptane, which is used as an oil, provokes a slight decrease. Consequently, the mobilities obtained in the microemulsion system are shifted to higher values (less negative mobilities) compared to mobilities obtained in the aqueous buffer, and so one cannot be directly subtracted from the other. Since the microemulsion-buffer medium cannot be directly reproduced, we propose a correction that takes into account the variation of viscosities. This is determined from the electrophoretic mobility of the benzoate ion. As this ion does not interact with the microemulsion, the ratio of its mobilities (measured in plain buffer and microemulsion) is equivalent to the ratio of viscosities, and can be used as the correction factor for other measurements. Thus, mobilities in buffer and microemulsion media are placed on the same scale, overcoming the errors in retention factor determination.

Micro-matrix solid-phase dispersion coupled with MEEKC for quantitative analysis of lignans in Schisandrae Chinensis Fructus using molecular sieve TS-1 as a sorbent.

A simple and effective method was developed for determining lignans in Schisandrae Chinensis Fructus by using a micro-matrix solid phase dispersion (MSPD) technique coupled with microemulsion electrokinetic chromatography (MEEKC). Molecular sieve, TS-1, was applied as a solid supporting material in micro MSPD extraction for the first time. Parameters that affect extraction efficiency, such as type of dispersant, mass ratio of the sample to the dispersant, grinding time, elution solvent and volume were optimized. The optimal extraction conditions involve dispersing 25mg of powdered Schisandrae samples with 50mg of TS-1 by a mortar and pestle. A grinding time of 150s was adopted. The blend was then transferred to a solid-phase extraction cartridge and the target analytes were eluted with 500µL of methanol. Moreover, several parameters affecting MEEKC separation were studied, including the type of oil, SDS concentration, type and concentration of cosurfactant, and concentration of organic modifier. A satisfactory linearity (R>0.9998) was obtained, and the calculated limits of quantitation were less than 2.77µg/mL. Finally, the micro MSPD-MEEKC method was successfully applied to the analysis of lignans in complex Schisandrae fructus samples.

Highly sensitive analysis of flavonoids by zwitterionic microemulsion electrokinetic chromatography coupled with light-emitting diode-induced fluorescence detection.

A rapid zwitterionic microemulsion electrokinetic chromatography (ZI-MEEKC) approach coupled with light-emitting-diode-induced fluorescence (LED-IF, 480nm) detection was proposed for the analysis of flavonoids. In the optimization process, we systematically investigated the separation conditions, including the surfactants, cosurfactants, pH, buffers and fluorescence parameters. It was found that the baseline separation of the seven flavonoids was obtained in less than 5min with a running buffer consisting of 92.9% (v/v) 5mM sodium borate, 0.6% (w/v) ZI surfactant, 0.5% (w/v) ethyl acetate and 6.0% (w/v) 1-butanol. High sensitivity was obtained by the application of LED-IF detection. The limits of detection for seven flavonoids were in the range of 3.30×10(-8) to 2.15×10(-6)molL(-1) without derivatization. Ultimately, the detection method was successfully applied to the analysis of flavonoids in hawthorn plant and food products with satisfactory results.

Microemulsion electrokinetic chromatography for analysis of phthalates in soft drinks.

Microemulsion electrokinetic chromatography (MEEKC) is proposed for analysis of di-n-butyl phthalate (DBP) and di-(2-ethylhexyl) phthalate (DEHP) in soft drinks. However, the instability of microemulsion is a critical issue. In this research, a novel material, Pluronic® F-127, which has the properties of polymer and surfactant, was added for stabilizing the microemulsion in the MEEKC system. Our data demonstrate that the presence of Pluronic® F-127 (0.05-0.30%) also helps enhance resolution of highly hydrophobic compounds, DBP and DEHP. The electrokinetic injection of sodium dodecyl sulphate (SDS) including sample (-10 kV, 20 s) was introduced in this MEEKC system and this yielded about 25-fold sensitivity enhancement compared with hydrodynamic injection (1 psi, 10 s). During method validation, calibration curves were linear (r≥0.99), within a range of 75-500 ng/mL for DBP and 150-1000 ng/mL for DEHP. As the precision and accuracy assays, absolute values of relative standard deviation (RSD) and relative error (RE) in intraday (n=3) and interday (n=5) observations were less than 4.93%. This method was further applied for analyzing six commercial soft drinks and one was found containing 453.67 ng/mL of DEHP. This method is considered feasible for serving as a tool for analysis of highly hydrophobic molecules.

Study on Quantitative Structure-Retention Relationships of Microemulsion Electrokinetic ChromatographyUsing Phospholipids as Surfactant / 分析化学

Some new types of microemulsion using phospholipids as the main surfactant were prepared for electrokinetic chromatography and the quantitative structure-retention relationship of neutral solutes in these microemulsion electrokinetic chromatography (MEEKC) systems was studied by solvation parameters model.By using dynamic coating capillary, and with dimethyl sulfoxide (DMSO) and dodecyl benzene as the marker of electroosmotic flow and microemulsion droplets, a total of 17 kinds of stable microemulsions containing soybean phospholipids or other surfactants were prepared and the linear salvation energy relationship equations were developed for these MEEKC systems with 26 small neutral compounds.The coefficients of linear solvation energy relationship (LSER) equations were used to evaluate the similarity of two MEEKC systems.Results indicated that LSER characteristics of phospholipids-MEEKC systems were similar to those of other microemulsion systems.The volume and hydrogen bond basicity of solutes were mostly contributed to the retention in MEEKC.The different types and concentration of oil phase had no evident influence on the retention.

On_line Preconcentration for Sensitive Analysis of PAHs in Cosmetics by Reversed Microemulsion Electrokinetic Chromatography / 分析化学

A sensitive analytical method based on reversed microemulsion electrokinetic chromatography ( MEEKC) combined with on_line preconcentration technique was developed for the determination of polycyclic aromatic hydrocarbons ( PAHs ) in cosmetics. For six lipophilic PAHs analytes which are difficult to be separated under conventional conditions, three stacking techniques including large volume sample stacking ( LVSS) , dynamic pH junction and sweeping ( LVSS_DypH_sweep ) were combined to realize the efficient preconcentration and separation. Under the optimum conditions, including the microemulsion buffer with the composition of 2. 4%(w/w)SDS_0. 6% (w/w) octane_6. 6% (w/w)n_butyl alcohol_20 mmol/L NaH2PO4 ( pH 2 . 2 ) , HCB injection time of 20 s ( 16 kPa ) and sample injection time of 80 s ( 16 kPa ) , good enrichment effect was reached with the enrichment factors ranged from 25 to 80 , and the PAHs were analyzed successfully within 27 min. The developed method was used to analyze the PAHs in cosmetics. The recoveries ranged from 90 . 6% to 95 . 9%. The RSD values ( n=5 ) were less than 5 . 1%.