Capillary Zone Electrophoretic Method Research Articles

Determination of Phenolic Glucosides in Gentiana piasezkii by Capillary Zone Electrophoresis

A capillary zone electrophoretic method has been developed for the quantitative analysis of five phenolic glucosides, 6′-O-vanilloylarbutin (VA), 7-O-feruloylorientin (FE), lutonarin (LN), isoorient (IO) and luteolin (LL), in Gentiana piasezkii with UV detection at 270 nm. 7-O-β-D-glucosyl-coumarin was selected as the internal standard. The applied voltage was 15 kV and the capillary temperature was kept constant at 25 °C. The effect of pH, the concentration of methanol and boric acid on migration were studied systematically. Optimum separation was achieved with 200 mM boric acid buffer at pH 9.50 containing 10% (v/v) methanol. Regression equations revealed good linear relationship between the peak area ratio of each compound and internal standard and its concentration. The correlation coefficients were 0.9975, 0.9997, 0.9998, 0.9998 and 0.9988 for VA, FE, LN, IO and LL, respectively. The relative standard deviations of migration time and the peak area ratio of each analyte and internal standard were <1.78% and 4.93%, respectively. The contents of the five compounds in Gentiana piasezkii were successfully determined with satisfactory repeatability and recovery.

Sulfur speciation by capillary zone electrophoresis: Determination of dithionite and its decomposition products sulfite, sulfate and thiosulfate in commercial bleaching agents

In this paper, a capillary zone electrophoretic (CZE) method was developed for the separation of the sulfur species dithionite (S 2O 4 2−), sulfite (SO 3 2−), sulfate (SO 4 2−) and thiosulfate (S 2O 3 2−). A carrier electrolyte (pH 7.0) containing 1.5 mmol L −1 pyromellitic (PM) acid, 10 mmol L −1 Tris(hydroxymethyl)-aminomethane (Tris), 0.5 mmol L −1 diethylenetriamine (DETA) and 0.1% (v/v) formaldehyde (as stabilizer for S 2O 4 2− and SO 3 2−) allowed the determination of the sulfur anions after 9 min CZE separation with indirect UV detection at 214 nm. The addition of 0.1% (v/v) formaldehyde to the sample solution stabilizes dithionite and sulfite as HOCH 2SO 2 − and HOCH 2SO 3 − anions. The procedure was applied for the determination of dithionite and its decomposition products sulfite, sulfate and thiosulfate in commercial formulations of bleaching agents. Dithionite was found to be the major component of the commercial formulations in concentrations between 30.80 and 33.30% (w/w). As anticipated, sulfite, sulfate and thiosulfate were found to be present as decomposition or by-products in the commercial formulations at concentrations of 14.30–14.80, 5.20–5.70 and 0.30–0.40% (w/w), respectively. The results were found to be in good agreement with those of polarographic and spectrophotometric determinations.

Comparison of shikimic acid determination by capillary zone electrophoresis with direct and indirect detection with liquid chromatography for varietal differentiation of red wines

Two capillary zone electrophoretic (CZE) methods for determination of shikimic acid in Chilean red wine were developed and compared with a HPLC method. Both electrophoretic methods were carried out by using a reversed electroosmotic flow induced by trimethyl(tetradecyl)ammoniumbromide (TTAB) with indirect detection at 260 nm using p-aminobenzoic acid as a UV-absorbing co-ion or by direct detection at 213 nm. In both cases, the separation was carried out in a 50 μm I.D. uncoated capillary with an effective length of 48 cm, a negative power supply of 30 kV, using a buffer based on bis[2-hydroxyethyl]imino-tris[hydroxymethyl]methane (Bis-Tris), pH 7.0 or 7.5 and hydrodynamic injection. The chromatographic separations were carried out on a C-18 reversed phase column followed by a sulfonyl-styrene-divinylbenzene (S-DVB) ion exclusion column at 70 °C with H 2SO 4 0.02 M as isocratic mobile phase and a flow rate of 0.5 mL min −1. The three methods allowed the quantification of shikimic acid with quantification limits between 1.0 and 12.0 mg L −1 and precision between 7.3 and 10.1%, however, only the concentrations obtained by CZE with direct detection were statistically similar to those of HPLC. This parameter was evaluated as analytical tool to verify varietal authenticity of red wines. In all cases, the Cabernet Sauvignon wines presented higher concentrations of shikimic acid, compared with Merlot or Carmenère wines.

Separation and Determination of Six Active Components in Two Myricaria Plants by Capillary Chromatography

A capillary zone electrophoretic (CZE) method for determination of six active components of two Myricaria plants has been developed for the first time. The analytes were completely separated within 15 min. The electrophoresis buffer was 25 mmol L−1 sodium borate containing 15% (v/v) acetonitrile (pH 10.20). The correlation coefficients of the calibration curves for the six analytes were 0.9998 or 0.9999 over the concentration ranges examined. Recoveries of the six constituents ranged from 96.3 to 106.8%. The method, combined with a relatively simple extraction procedure, was successfully used for analysis of two Myricaria plants and the assay results were satisfactory.

Micellar electrokinetic chromatographic and capillary zone electrophoretic methods for screening urinary biomarkers of human disorders: a critical review of the state-of-the-art.

Human urine plays a central role in clinical diagnostic being one of the most-frequently used body fluid for detection of biological markers. Samples from patients with different diseases display patterns of biomarkers that differ significantly from those obtained from healthy subjects. The availability of fast, reproducible, and easy-to-apply analytical techniques that would allow identification of a large number of these analytes is thus highly desiderable since they may provide detailed information about the progression of a pathological process. From among the variety of methods so far applied for the determination of urinary metabolites, capillary electrophoresis, both in the capillary zone electrophoresis (CZE) and micellar electrokinetic chromatography (MEKC) modes, represents a robust and reliable analytical tool widely used in this area. The aim of the present article is to focus the interest of the reader on recent applications of MEKC and CZE in the field of urinary biomarkers and to discuss advantages and/or limitations of each mode.

Rapid simultaneous determination of alkylxanthines by CZE and its application in analysis of pharmaceuticals and food samples

Capillary electrophoresis (CE) offers the possibility of fast, cheap and reproducible separations for pharmaceutical preparations. Alkylxanthines make up a family of compounds that are used in the treatment and prevention of bronchi asthma and chronic pulmonary disease. The group of analysed compounds include caffeine, dyphylline, theophylline, theobromine and enprofylline. This paper shows a simple capillary zone electrophoretic (CZE) method for separation of this group of xanthines. Using 20 mM borate buffer at pH 9.4 as running buffer at 55 °C it was possible to complete a total separation of a sample in 2 min. Limits of detection in the range 1.9–2.5 mg l −1 were achieved with %R.S.D. of 0.06–0.22% ( n = 5). The technique is applied to a range of samples containing the analytes, including tablets and chocolate. Reproducibility (%R.S.D.) of the chocolate analysis technique by CZE was less than 4.5%.

Optimization and validation of a capillary zone electrophoretic method for the simultaneous analysis of four atypical antipsychotics

A capillary zone electrophoretic method has been developed and optimized for separation of four atypical antipsychotics (AAPs): clothiapine (cT), clozapine (cZ), olanzapine (O), and quetiapine (Q). A three-level full-factorial design was applied to study the effect of the pH and molarity of the running buffer on separation. Combination of the studied parameters permitted the separation of the four AAPs, which was best carried out using 80 mM sodium phosphate buffer (pH 3.5). The same system can also be applied for the quantitative determination of these compounds. The method was then validated regarding linearity, precision, and accuracy. Especially, the possibility of simultaneous quantification and identification of the active ingredient in the finished product is very attractive.

Experimental Design Optimization of a Capillary Zone Electrophoresis Method for the Screening of Several Diuretics and ACE Inhibitors

Experimental design methodologies are applied to the development of a capillary zone electrophoretic method for the separation of the angiotensin-converting enzyme inhibitor enalapril and its derivative enalaprilat and the diuretics xipamide and hydrochlorothiazide. The effects of pH, buffer concentration, proportion of boric acid in the mixed boric acid-potassium dihydrogen phosphate background electrolyte, temperature, applied voltage, and percentage of organic modifier are studied. Critical factors are identified in a screening design (a 2(6-2) fractional factorial design), and afterwards, optimal conditions for the separation are reached by means of an optimization design (a 2(2) + 2 x 2 + k central composite design). The studied response is the resolution between peaks. The four studied compounds can be separated in less than 3.5 min using an electrolyte of 20mM boric acid-potassium dihydrogen phosphate (75:25, v/v) with 5% MeOH adjusted to pH 8.0 with KOH, at a potential of 30 kV. The detection wavelength and temperature are 206 nm and 35 degrees C, respectively.

Validation of a Capillary Zone Electrophoresis Method for Determination of Rimantadine Hydrochloride in Rimantadin100 Tablets and the Method Application to Dissolution Test Monitoring

A capillary zone electrophoretic method with indirect UV‐detection for determination of rimantadine, an antiviral drug against influenza A, in tablets was validated. Instrumental precision, the method precision, accuracy, calibration curve linearity, selectivity, robustness, and time stability of the sample and the standard were tested. The method was also applied to monitor dissolution tests of the tablets. The possibility of addition of an internal standard for improvement of the method precision was discussed.

Direct and fast capillary zone electrophoretic method for the determination of Gleevec and its main metabolite in human urine

A capillary zone electrophoretic (CZE) method was investigated for the determination of Gleevec and its main metabolite ( N-demethylated piperazine derivative) in human urine using a fused-silica capillary (75 μm I.D.×60 cm total length, 10 cm effective length). The separation was performed with an hydrodynamic injection time of 10 s (0.5 p.s.i.) a voltage of −25 kV, a capillary temperature of 25 °C and a 100 m M phosphoric acid adjusted to pH 2 with the addition of triethanolamine. Under these conditions, the analysis takes about 5 min. A linear response over the 0.4–30.0 mg l −1 concentration range was investigated for two compounds. A dilution of the sample was the only step necessary before the electrophoresis analysis. Detection limits of 0.1 mg l −1 for Gleevec and its metabolite ( S/ N=3) were obtained. The developed method is easy, rapid and sensitive and has been applied to determine Gleevec and its main metabolite in clinical urine samples.

Determination of Cathepsin D Activity in MCF-7 Cells by Capillary Zone Electrophoresis with On-Column Sample Stacking

A rapid and simple capillary zone electrophoretic (CZE) method has been developed for analysis of cathepsin D activity in MCF-7 cells. Enzyme active suspension was separated and incubated with hemoglobin as a substrate. Specific peptides were then separated and analyzed by capillary zone electrophoresis with on-column sample stacking. The CZE conditions, including the organic solvents used, their concentrations, and pH and separation voltage, were optimized. The results showed that dilution of the sample with acetonitrile was compensated by using large-volume injection in the stacking procedure. The limit of determination of the method was dramatically improved. This relatively rapid and simple means of determination of cathepsin D activity was successfully used to assay the expression of cathepsin D in cultured MCF-7 cells, to evaluate the estrogenic activity of hormone-disrupting chemicals present in the environment.

Quality control of commercial tablets containing the novel antipsychotic quetiapine

Quetiapine (bis [2-(2-[4-(dibenzo[b,f][1,4]thiazepin-11-yl]ethoxy)ethanol]fumarate) is the most recent agent introduced on the drug market for the treatment of psychotic disorders. Two different analytical methods for the quality control of quetiapine in commercial formulations have been developed and compared: a spectrophotometric method and a capillary zone electrophoretic (CZE) method. The spectrophotometric assay was carried out measuring the absorbance at a wavelength of 246 nm. The CZE method used an uncoated fused-silica capillary and a pH 2.5, 50 mM phosphate buffer as the background electrolyte. The detection wavelength was 205 nm, the separation voltage was 15 kV, and a complete electrophoretic run lasts less than 2.5 min. Extraction of quetiapine from the commercial tablets consisted of a simple one-step treatment with a pH 2.5, 50 mM phosphate buffer. Linearity was observed in the 5–25 μg ml −1 concentration range of quetiapine for the spectrophotometric method, and in the 5–50 μg ml −1 concentration range for the electrophoretic method. Both methods gave satisfactory results in terms of repeatability and intermediate precision (RSD<1.9%). Also accuracy values were very good for both methods, the recovery being between 98.2 and 100.5%.

Simultaneous determination of hydrochlorothiazide and several angiotensin-II-receptor antagonists by capillary electrophoresis

We have investigated the capability of the capillary zone electrophoretic (CZE) and micellar electrokinetic capillary chromatographic (MEKC) methods to simultaneously separate hydrochlorothiazide and six angiotensin-II-receptor antagonists (ARA-IIs): candesartan, eprosartan mesylate, irbesartan, losartan potassium, telmisartan, and valsartan. The CZE and MEKC methods are suitable for the qualitative and quantitative determination of combined HCT/ARA-IIs in pharmaceutical formulations. Depending on the ARA-II, at least one of the two methods can be used for each combination. The two methods have been validated in terms of their linearity of response, reproducibility, and accuracy.

Simultaneous analysis of metabisulfite and sulfate by CE with indirect UV detection. Application to and validation for a pharmaceutical formulation

Metabisulfite is used as an antioxidant agent in a number of pharmaceutical formulations. In order to quantify simultaneously both metabisulfite and its oxidation product (sulfate), a capillary zone electrophoretic (CZE) method with indirect UV detection was developed. Best results were achieved with a background electrolyte (BGE) constituted of 15 mM pyromellitic acid, 15 mM tris-(hydroxymethyl)-aminomethane and 0.2 mM tetradecyltrimethylammonium bromide at pH 8.3 and an applied electrical field of 123 V/cm in a 32.5 cm fused silica capillary. Indirect UV detection was performed at a wavelength of 225 nm. In order to validate this method, an internal standard (IS), namely ammonium formate, was used. Moreover, due to the high chloride concentration in the pharmaceutical formulation, conductivity was adjusted by adding sodium chloride into standard solutions to prevent matrix effect. Linearity and accuracy were successfully tested in a concentration range of 33.3–250 μg/ml for sodium metabisulfite and of 50–375 μg/ml for sodium sulfate. Method precision was determined on six samples each day. Thereby, relative standard deviations (R.S.D.) of 6% and 12–13% were obtained for intra-day and inter-day precision, respectively. Considering the instability of metabisulfite and its use as an antioxidant agent and not as an active principle, the method was accepted and used for routine analyses.

Capillary zone electrophoretic assay of biologically active thioacridine derivatives

AbstractA capillary zone electrophoretic method was developed for the analytical evaluation of newly synthesized biologically active thioacridine derivatives. Based on background electrolyte pH optimization, the experimental conditions ensuring satisfactory resolution of the studied set of thioacridine derivatives and their accompanying synthetic impurities were found. Optimum background electrolyte consists of 30 mM citric acid and 30 mM β‐alanine (pH 3.74). Calibration data confirmed linear response for all compounds of interest in the concentration range 1–500 μg/mL, sufficiently high sensitivity, and limits of detection not exceeding 0.16 μg/mL. The described assay proved to be suitable for rapid identification and purity evaluation of studied thioacridine derivatives. The qualitative and quantitative results have been compared with those obtained using conventional liquid chromatography. The differences were not generally significant.

Simultaneous separation of 11 protease and reverse transcriptase inhibitors for human immunodeficiency virus therapy by co-electroosmotic capillary zone electrophoresis.

In the present investigation, a co-electroosmotic capillary zone electrophoretic method is shown for the simultaneous separation of protease inhibitors and reverse transcriptase inhibitors, which are used as antiretroviral therapy drags against the human immunodeficiency virus (HIV). The investigated drugs carry basic amino groups, thus the electrophoretic system takes advantage of an acidic buffer electrolyte. In order to establish a strong cathodic electroosmotic flow (EOF), a poly-anionic surfactant is added to the background electrolyte. Thus, fast migration times due to a co-directional migration of analytes and EOF (co-electroosmotic CE) are obtained. The developed separation system exhibits good selectivities for the investigated compounds and sufficient sensitivity to monitor drug levels in the low ppm range in HIV positive patients who are treated by highly active anitiretroviral therapy.

Capillary zone electrophoresis of syringyl and guaiacyl monomers resulting from lignin thioacidolysis.

A capillary zone electrophoretic method has been developed for the quantitative determination of syringyl (S) and guaiacyl (G) monomers resulting from lignin thioacidolysis. The effects on the separation of altering a number of parameters have been investigated, resulting in an efficient and rapid separation. Analysis times were about 10 min instead of 50 min for the conventional GC methods, with no requirement for a derivatisation step prior to analysis. Relative standard deviations ranged between 8 and 12% for the absolute determination of S and G monomers, and between 1.4 and 3.5% for the S/G ratio.

Determination of dissociation constants of low molecular weight organic acids by capillary zone electrophoresis and indirect UV detection

A capillary zone electrophoretic (CZE) method has been developed to determine dissociation constants (pKa) for anionic, non-UV absorbing species in the range 3.3–4.3. The procedure was evaluated by determining the pKa for two well-characterised low molecular weight organic acids: formic and lactic acids. The results correspond well with literature values (in parentheses): formic acid 3.71 (3.75) and lactic acid 3.84 (3.85). The pKa of two polyhydroxycarboxylic acids were also determined, namely 3.64 for gluconic acid and 3.87 for isosaccharinic acid (only poorly described in the literature).

Optimization and validation of a capillary zone electrophoretic method for the analysis of several angiotensin-II-receptor antagonists.

We optimized a capillary zone electrophoretic method for separation of six angiotensin-II-receptor antagonists (ARA-IIs): candesartan, eprosartan, irbesartan, losartan potassium, telmisartan, and valsartan. A three-level, full-factorial design was applied to study the effect of the pH and molarity of the running buffer on separation. Combination of the studied parameters permitted the separation of the six ARA-IIs, which was best carried out using 60 mM sodium phosphate buffer (pH 2.5). The same system can also be applied for the quantitative determination of these compounds, but only for the more soluble ones. Some parameters (linearity, precision and accuracy) were validated.

Capillary zone electrophoretic determination of tosufloxacin and trovafloxacin in urine

A simple and rapid capillary zone electrophoretic method with UV detection has been developed for determination of tosufloxacin and trovafloxacin. The separation was performed in fused-silica capillaries (57 cm length × 75μm i.d.); the running buffer was 35mm borate + 35mm phosphate buffer solution, pH 8.6, containing 6% (v/v) acetonitrile. The applied potential was 15 kV, the temperature 30°C, and detection was at 262 nm. Piromidic acid was used as the internal standard. Response was linearly dependent on concentration in the range 1.0–120.0 μg mL−1 and the detection limit was 0.2 μg mL−1 for both compounds. The analysis was highly reproducible (RSD between 3.41 and 1.25%). The method was applied to the determination of tosufloxacin and trovafloxacin in human and rat urine. The method was validated by using HPLC as a reference method. Recovery was between 96.8 and 102%.