Rapid Capillary Zone Electrophoresis Research Articles

STUDIES OF THE CAPILLARY ELECTROPHORESIS BEHAVIOR OF SCOPOLAMINE-RELATED ANTICHOLINERGICS AND ITS APPLICATIONS

A simple and rapid capillary zone electrophoresis (CZE) method was proposed for the separation and quantitative analysis of atropine sulfate (AS), anisodamine hydrobromide (AH) and scopolamine hydrobromide (SH). The parameters affecting the migration behavior of these drugs were studied in detail, including the background electrolyte acidity and its concentration. In the optimized conditions, the method was used to determine the AS, AH and SH in various samples (Tablets, Injections and China herbal medicines of Hyoscyamus niger). The relative standard deviations of the migration time and the peak areas were 0.45–1.32% and 2.1–4.58%, respectively. The recovery of the three drugs ranges from 94.3 to 102.4%.

Quantification of Reduced and Oxidized Glutathione in Whole Blood Samples by Capillary Electrophoresis

In the past few years, there has been increasing interest in the measurement of thiols, glutathione in particular, as indicators of oxidative stress. The reduced glutathione/oxidized glutathione ratio (GSH/GSSG) is used to evaluate oxidative stress status in biological systems, and alterations of this ratio have been demonstrated in aging, cancer, HIV replication, and cardiovascular diseases (1)(2)(3)(4)(5). Surprisingly, glutathione is usually studied in plasma, which contains only 0.5% of the blood content, whereas erythrocytes contain 99.5%. We were interested in direct measurement of GSH and GSSG in whole blood samples to evaluate overall glutathione status. Numerous methods are available for measuring GSH and GSSG (6), but few of these are suitable for direct analysis in routine use. HPLC is the most widely used, but it requires a long sample preparation with pre- or postcolumn derivatization (7)(8) or a special apparatus for electrochemical detection (1)(9). Alternative methods based on GSH conjugation with a chromophore or a fluorophore have been used, but they need GSSG calculation from total glutathione measured after reduction of GSSG (10)(11). Capillary zone electrophoresis (CZE) coupled with laser-induced fluorescence has been proposed recently (12). The separation of GSH and GSSG by CZE with direct ultraviolet detection in tissues and mitochondria (13)(14) and by micellar electrokinetic capillary electrophoresis in plasma (15) has also been reported. The aim of the present work was to develop and validate a rapid CZE method suitable for a direct measurement of GSH and GSSG in whole blood. Moreover, we describe the distribution of blood GSH and GSSG concentrations in a healthy adult population and in elderly subjects. All chemicals used were of analytical-reagent grade. bis-Tris, boric acid, GSH, and GSSG were obtained from Sigma Chemical, and metaphosphoric acid was …

Determination of trimebutine maleate in rat plasma and tissues by using capillary zone electrophoresis.

A simple and rapid capillary zone electrophoresis method was developed for the determination of trimebutine maleate in rat plasma and tissues. Rat plasma and tissue homogenates were mixed with acetonitrile containing internal standard, ephedrine hydrochloride, and then centrifuged. The supernatant was dried under a stream of nitrogen, and the residue was reconstituted in methanol-water (1:1). The electrophoresis was performed in uncoated capillary with 30 mmol/L phosphate buffer of pH 6.0 as the separation electrolyte. The applied voltage was 10 kV and the UV detection was set at 214 nm. The peak height ratio vs concentration in plasma or homogenates was linear over the range of 5-500 ng/mL and the limit of quantitation was 5 ng/mL. The intra- and inter-day precision was RSD < 14% and <15%. The accuracy was relative error (RE) within +/- 14%. This method was applied to studying the pharmacokinetics and tissue distribution after a single dose of trimebutine maleate was administrated to the rats. The T(max), AUC, C(max) and t(1/2) were 30 min, 7.8 x 10(2) (ng/mL) min, 39 ng/mL and 1.7 x 10(2) min. The drug distribution was found in a decreasing order of liver, kidney, spleen, lung and heart.

Capillary Zone Electrophoresis for the Diagnosis of Congenital Hemoglobinopathies

The analysis of human hemoglobins (Hbs) is of medical importance in a number of congenital defects. The hemoglobinopathies are grouped into defective variants of Hb, such as Hb S and >600 other variants, and thalassemias characterized by abnormal expression of the genes for normal globin chains (1)(2)(3). Alkaline electrophoresis, performed historically on cellulose acetate and currently on agarose, combined with citrate agar electrophoresis at acidic pH is still widely used (4)(5)(6)(7), but high-performance cation-exchange chromatography (HPCEC) offers superior resolution, speed, and automation (8)(9)(10). Capillary electrophoresis uses numerous separation principles and shares with HPLC the advantages of high resolution and automation, with on-line detection and direct quantification (11)(12)(13)(14)(15). Capillary isoelectric focusing on coated capillaries can be used to study Hb (16)(17)(18)(19)(20)(21), but is slower than HPCEC (21). The first reported assays based on capillary zone electrophoresis (CZE) gave poor resolution of Hbs or were not quantitative (22)(23)(24). The aim of the present work was to evaluate a rapid CZE assay with dynamic coating of the fused-silica at alkaline pH to detect abnormal Hbs and to quantify Hbs for the diagnosis of thalassemias, and at acidic pH to confirm the identity of Hb variants. Reagents were obtained from Analis. The alkaline kit A2 contained a hemolyzer, an initiator consisting of a polycation, and an arginine buffer (pH 8.8) containing a polyanion. The acidic kit A1 has been described elsewhere for Hb A1c measurement (25). Adult and newborn samples were collected in EDTA-containing tubes at the maternity unit of the Hopital Saint-Antoine (AP-HP, Paris, France) and received in the laboratory for Hb analysis. Whole blood samples …

Separations of inorganic anions based on their complexations with α-cyclodextrin by capillary zone electrophoresis with contactless conductivity detection

Capillary zone electrophoresis (CZE) separations of inorganic anions based on their host–guest complex equilibria with α-cyclodextrin (α-CD) were investigated. α-CD employed as a host was found to influence selectively the effective mobilities most of the studied anions (chloride, bromide, iodide, sulfate, nitrite, nitrate, fluoride, phosphate). Its complexing ability combined with a low pH of the carrier electrolyte solution provided working conditions suitable for rapid CZE separations of the anions. For example, iodide and chloride present in the injected sample in a concentration ratio of ca. 1:2·10 3 could be separated in less than 100 s and seven of the above anions (α-CD failed to resolve chloride and bromide) were baseline-resolved in less than 120 s. A high overall selectivity of the carrier electrolytes combining the host–guest equilibria with a low pH in the analysis of highly complex samples is illustrated by CZE separations of inorganic anions present in milk. Here, no interferences to the separations of inorganic anions due to co-migrations of organic acids present in milk were observed also when a 200 nl volume of a diluted (1:10) milk sample was loaded onto the column. A contactless conductivity detector used for the detection of anions in this work proved very reliable. Under our separating conditions it provided for the studied anions concentration limits of detection in the range of 0.5–1.4 μmol/l (a 200-nl sample injection volume) when the separations were carried out in a 300 μm I.D. capillary tube made of polytetrafluoroethylene.

Influence of organic solvents in the capillary zone electrophoresis of polycyclic aromatic hydrocarbon metabolites

A rapid capillary zone electrophoresis (CZE) method was developed using a 3-(cyclohexylamino)-1-propanesulfonic acid (CAPS) buffer for the compound-class separation of nine polycyclic aromatic hydrocarbon (PAH) metabolites including tetrols, benzo[ a]pyrene diols, and hydroxyl aromatics. In addition, the effects of two organic solvents (methanol and acetonitrile) on the electroosmotic flow and electrophoretic mobilities of the analytes were studied. Separation of nine PAH metabolites was ultimately achieved within 24 min with a 100 m M CAPS buffer with 40% (v/v) MeOH at an apparent pH of 10.4. In addition, it was possible to partially resolve the parent PAH compounds from the metabolites with the previous buffer system. The simple CZE methods developed would be useful for the rapid separation and characterization of several important biomarkers.

Rapid capillary zone electrophoresis in isoelectric histidine buffer: high resolution of the poly-T tract allelic variants in intron 8 of the CFTR gene

The poly-T tract in intron 8 of the cystic fibrosis conductance transmembrane regulator (CFTR) gene exists in three variants, 5T, 7T, and 9T. The 7T and 9T variants generate a predominantly normal transcript, whereas the 5T variant engenders an anomalous product. The analysis of the poly-T tract is assuming increasing relevance, both to assess the implication of the CFTR gene in congenital bilateral absence of the vas deferens and to evaluate genotype-phenotype correlation in cystic fibrosis. Mapping of the poly-T tract has been performed by cumbersome and time-consuming methodologies. Capillary zone electrophoresis, combined with laser-induced fluorescence detection, was introduced for a rapid separation of the poly-T tract amplified products. As separation buffer, we adopted 200 mmol/L histidine (pH = pI = 7.6), and the capillary was filled with 10% polyacrylamide, allowing separations in less than 10 min. Capillary zone electrophoresis results were in perfect agreement with dot-blot analysis.

Separation of synthetic food colourants by capillary zone electrophoresis in a hydrodynamically closed separation compartment

Separation conditions enabling the complete resolution of eleven permitted synthetic food colourants and some of their subspecies by capillary zone electrophoresis (CZE) were determined. Those conditions involve combining a suitable pH of the carrier electrolyte (pH 6.8) with host-guest complexation effects of β-cyclodextrin. A 300 μm I.D. capillary tube made of fluorinated ethylene-propylene copolymer in a hydrodynamically closed separation compartment was used for the CZE separations. The capillary could accommodate 90-nl sample injection volumes, thus providing limits of detection for the dyes of 11–300 ppb using a photometric detector operating at a wavelength of 254 nm. R.S.D.s of 0.4–3.0% were typical for the determinations of the dyes present in samples at 16 ppm concentrations. Erythrosine, exhibiting residual adsorption, gave more-scattered results under identical working conditions (R.S.D. of ca. 9.0%). The utility of this rapid CZE procedure (migration times of the dyes were 2.5–10.5 min) is illustrated for several practical samples, including soft drink concentrate and liqueur and monitoring of the stability of aqueous solutions of indigo carmine.

Purity control of different bee venom melittin preparations by capillary zone electrophoresis.

A rapid capillary zone electrophoresis method was developed for the determination of melittin during its purification from bee venom by column chromatography. Low-pH buffer was selected to decrease the adsorption of the analytes on capillary wall. The average relative standard deviations of the migration time of melittin and of the peak area were 1.2 and 2.3%, respectively.

Determination of sorbic acid in food products by capillary zone electrophoresis in a hydrodynamically closed separation compartment

A simple, selective and rapid capillary zone electrophoresis method for the determination of sorbic acid in food products is described. The determination was carried out in a hydrodynamically closed separation compartment at pH 5.2 and for the samples taken into this study (soft drinks, wine, fruit and juice concentrates, margarine and marmalade) only simple sample preparation procedures were needed. A very good reproducibility in the migration time of the analyte (R.S.D. = 0.61%) was achieved and as could be expected no negative effect of matrix constituents on this performance parameter was detected. The recoveries of sorbic acid in the spiked samples ranged from 98-102% and 2-3% R.S.D.s in parallel determinations of the acid were typical.

Coupled fused silica capillaries for rapid capillary zone electrophoresis of proteins

AbstractA novel two‐dimensional electrophoretic system for the control of electroosmosis in capillary zone electrophoresis has been developed and evaluated for rapid separations of proteins. The system comprises uncoated and polyether‐coated fused silica capillaries coupled in series. An equation relating the average electroosmotic flow velocity in the coupled capillaries to the intrinsic electroosmotic velocities of the connected segments and their corresponding lengths has been derived and verified experimentally. This approach has the advantage of enabling the electroosmotic flow to be tuned independently of the applied voltage. As a consequence, rapid protein analysis at relatively low field strength was achieved without sacrificing the high separation efficiencies obtained with surface‐modified capillaries.