Application of a novel micro-injector in the determination of indole derivatives in the rat pineal gland by capillary electrophoresis with electrochemical detection

A novel micro-injector has been fabricated for capillary electrophoresis (CE). It was successfully used for the determination of some indole derivatives for example melatonin (MT), serotonin (5-HT), tryptophan (Trp), and 5-hydroxy-tryptophane (5-HTrp) in the rat pineal gland by capillary electrophoresis with electrochemical detection (CE–EC). CE was performed in 0.20 mol L^–1 phosphate buffer (pH 8.0). The compounds investigated can be well separated and detected within 15 min. The working electrode used was a 300-μm diameter carbon electrode positioned opposite the outlet of the capillary. The relationship between peak current and analyte concentration was highly linear in the range from 0.10 to 500 μmol L^–1; detection limits (S/N = 3) were 0.03–0.13 μmol L^–1. The proposed method has been successfully used to analyze real biological samples.     

Determination of amino acids in tobacco samples by capillary electrophoresis/indirect absorbance detection with isolation of the electrolysis compartment and p-Aminobenzoic acid as a background electrolyte.

A fast, convenient and sensitive method of capillary zone electrophoresis (CZE) and indirect UV detection was proposed for the determination of 16 amino acids. p-Aminobenzoic acid (PAB) was selected as a background electrolyte (BGE). An isolated cell included a BGE buffer part and an electrode buffer one, which were jointed with a glass frit. The isolated cell can prevent PAB from the electrode reaction and improve the stability of the detection baseline. The separation conditions of amino acids were investigated, such as different BGEs, BGE concentration, buffer pH and electroosmotic flow (EOF) modifiers. Under the selected separation conditions, 14 amino acid peaks could be separated in 12 min. The detection limits of the amino acids were in the range of 1.7 - 4.5 micromol/L. The isolated cell is suitable for reagents reacting on the electrodes in capillary electrophoresis. The proposed method has been successfully applied to the determination of the amino acids in tobacco samples.     

Development of capillary electrophoresis methods for quantitative determination of taurine in vehicle system and biological media

CE methods have been developed for the determination of taurine in pharmaceutical formulation (microemulsion) and in biological media such as sweat. The CE system with end-column pulsed amperometric detection has been found to be an interesting method in comparison with UV and fluorescence detection for its simplicity and rapidity. A gold-disk electrode of 100 mm diameter was used as the working electrode. The effects of a field decoupler at the end of the capillary, separation voltage, injection and pressure times were investigated. A detection limit of 4 x 10(-5) mol/L was reached using integrated pulsed amperometric detection, a method successfully applied to taurine analysis of the biological samples such as sweat. For taurine analysis of oil-in-water microemulsion, fluorescence detector was the favored method, the detection limit of which was 4 x 10(-11) mol/L.     

Determination of inorganic anions in human saliva by zwitterionic micellar capillary electrophoresis

Capillary electrophoresis (CE) using sulfobetaine-type zwitterionic micelles as the background electrolyte (BGE) has been used to determine inorganic anions in human saliva. The zwitterionic micelles resulted in unique migration behavior for the separation of inorganic anions. They also prevented adsorption of proteins on the inner wall of the capillary. These properties of the zwitterionic micelles enabled the direct determination of inorganic anions in human saliva. Three species of inorganic anions, NO2-, NO3-, and SCN-, were found in real samples and the analysis was achieved within 3 min. Direct UV-absorption was used as the detection method and the detection limits for these anions were 2.0, 1.0, and 5.0 micromol L(-1), respectively (0.09, 0.06, and 0.30 microg mL(-1)).     

Tris(2,2'-bipyridyl)ruthenium(II)-zirconia-Nafion composite films applied as solid-state electrochemiluminescence detector for capillary electrophoresis

The major goal of this work was to develop a new solid-state electrochemiluminescence (ECL) detector suitable for capillary electrophoresis (CE). The detector was fabricated by coating a sol-gel derived zirconia (ZrO(2))-Nafion composite film on a graphite electrode, then the zirconia-Nafion modified electrode was immersed in tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy)(3) (2+)) solution to immobilize this active chemiluminescence reagent. The voltammetric and ECL behaviors of the detector were investigated and optimized in tripropylamine solution. The ratio of 53% for zirconia in the zirconia-Nafion composite provided the highest luminescence intensity of immobilized Ru(bpy)(3) (2+). The ECL can maintain its stability very well in the phosphate solution in the period of 5-90 h when the solid-state ECL detector was immersed in the solution all the time. The optimum distance of capillary outlet to the solid-state ECL detector has been found to be ca. 50-80 microm for a 75 microm capillary. The effects of ionic strength and pH of ECL solution on peak height were investigated. The CE with solid-state ECL detector system was successfully used to detect tripropylamine, lidocaine, and proline. The detection limits (S/N = 3) were 5 x 10(-9) mol.L(-1) for tripropylamine, 1 x 10(-8) mol.L(-1) for lidocaine and 5 x 10(-6) mol.L(-1) for proline, and the linear ranges were from 1.0 x 10(-8) to 1.0 x 10(-5) mol.L(-1) for tripropylamine, 5.0 x 10(-7) mol.L(-1) to 1.0 x 10(-5) mol.L(-1) for lidocaine and 1.0 x 10(-5) to 1.0 x 10(-3) mol.L(-1) for proline, respectively.     

中药菟丝子中生物活性成分的毛细管电泳-电化学检测

采用毛细管电泳-电化学检测法(CE-ECD)同时测定了菟丝子中芦丁、金丝桃甙、山柰酚、对香豆酸和槲皮素等5种主要生物活性成分的含量,考察了运行缓冲液酸度和浓度、分离电压、氧化电位和进样时间等实验参数对分离检测的影响。在最佳实验条件下,以直径300 μm的碳圆盘电极为工作电极,检测电位为+950 mV(vs. 参比电极),以50 mmol/L的硼砂缓冲溶液(pH 9.0)为运行缓冲液,上述各组分在19 min内能完全分离。芦丁、金丝桃甙、山柰酚、对香豆酸和槲皮素在两个数量级的范围内呈良好线性关系,检测下限(按S/N=3计) 分别为1.93×10-5,3.55×10-4,3.65×10-5,1.73×10-5和1.46×10-4 g/L。该法已成功地应用于菟丝子中活性成分的分离检测,结果令人满意。     

Determination of hesperidin and synephrine in Pericarpium Citri Reticulatae by capillary electrophoresis with electrochemical detection

A method based on capillary electrophoresis with electrochemical detection (CE-ED) has been developed for the determination of hesperidin (HP) and synephrine (SP) in the Chinese traditional herbal drug, Pericarpium Citri Reticulatae, the dried rind of the ripe fruits of Citrus reticulata Blanco (mandarin orange). The effects of some important factors such as the acidity and concentration of running buffer, separation voltage, and detection potential were investigated to determine the optimum conditions. The working electrode was a 300 microm diameter carbon disc electrode positioned opposite the outlet of the capillary. Both analytes could be well separated within 5 min in a 40 cm long capillary at a separation voltage of 12 kV in 50 mmol L(-1) borate buffer (pH 9.0). Excellent linearity was observed for the dependence of peak current on analyte concentration in the range from 2.5 x 10(-6) to 1.0 x 10(-3) mol L(-1) for SP and from 5.0 x 10(-6) to 1.0 x 10(-3) mol L(-1) for HP. The detection limits (S/N=3) for SP and HP were 4.96 x 10(-7) mol L(-1) and 6.54 x 10(-7) mol L(-1), respectively. This method has been successfully applied for the analysis of real samples, with satisfactory results.     

Dynamic pH junction-sweeping for on-line focusing of dipeptides in capillary electrophoresis with laser-induced fluorescence detection

In this paper, we developed a simple and effective on-line focusing technique combining dynamic pH junction and sweeping by capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection. Dynamic pH junction-sweeping is defined when the sample has a different buffer pH (dynamic pH junction condition) and is devoid of micelles (sweeping condition) relative to the background electrolyte (BGE). This hyphenated focusing mode was applied to the sensitive and selective focusing of four dipeptides: Tyr-Phe, Tyr-Leu, Trp-Gly, and Ala-Gln. Picomolar detectability of these dipeptides by CE-LIF detection was demonstrated through effective focusing of large sample volumes (up to 39% capillary length) using the dual pH junction-sweeping focusing mode. 25 mmol L(-1) sodium dihydrogen phosphate, pH 2.5 was used as the sample matrix, and 100 mmol L(-1) borate, 21 mmol L(-1) sodium dodecylsulfate (SDS), 16 mmol L(-1) Brij35, pH 9.0 as the background solution (BGS). The concentration detection limits (S/N = 3) of the four dipeptides were in the range of 1.0-5.0 pmol L(-1). The developed method has been successfully used for the determination of dipeptides in human serum samples.     

Development of a gas diffusion probe for the rapid measurement of pCO2 in aquatic samples

A capillary electrophoresis (CE) procedure with contactless conductivity detection (C(4)D) has been developed for monitoring of neutral mono- and disaccharides in drinks and foodstuffs. The separation of a mixture of seven neutral saccharides (glucose, fructose, galactose, mannose, ribose, sucrose and lactose) employed a quartz capillary, 5 μm i.d., with an effective length of 18.3 cm, and 75 mM NaOH (pH 12.8) as the background electrolyte (BGE). The limit of detection (LOD) values obtained lied within a range from 0.4 μmol L(-1) for lactose to 0.9 μmol L(-1) for ribose, with a separation time shorter than 140 s. The procedure was successfully applied to determinations of saccharides in fruit juices, Coca-Cola, milk, red and white wines, yoghurts, honey and a foodstuff additive.     

毛细管电泳-电化学检测法测定硫酸铜-维生素C反应体系中的羟基自由基和菊花的抗氧化活性

采用毛细管电泳-电化学检测方法,以对羟基苯甲酸为自由基捕捉剂,测定了硫酸铜-维生素C反应体系(pH 7.4)中生成的羟基自由基。研究了电极电位、运行液的pH值、电泳电压及进样时间对体系中反应物和产物分离的影响,得到了最优化的测定条件;讨论了体系中各反应物浓度和反应时间对产物浓度的影响。以直径为300 μm的碳圆盘电极为检测电极,工作电极电位为0.95 V(vs.SCE),目标产物3,4-二羟基苯甲酸在1.5×10-4~6.0×10-6 mol/L范围内线性关系良好,检测限（S/N=3）为1.5×10-6 mol/L。应用该方法,研究测定了不同品种菊花的抗氧化活性。     

Analysis of nephroloxic and carcinogenic aristolochic acids in Aristolochia plants by capillary electrophoresis with electrochemical detection at a carbon fiber microdisk electrode

Aristolochic acids (AAs) are the main bioactive ingredients in the most of Aristolochia plants, which are used to make dietary supplements, slimming pills and Traditional Chinese Medicines (TCMs). Excessive ingestion of AAs can lead to serious nephropathy. Therefore, quantitative analysis and quality control for the plants containing AAs is of great importance. In this paper, capillary electrophoresis (CE) with electrochemical detection (ED) at a 33 microm carbon fiber microdisk electrode (CFE) has been applied to detect AA-I and AA-II in Aristolochia plants. Under the optimum conditions: detection potential at 1.20 V, 2.0 x 10(-2) mol L(-1) phosphate buffer solution (PBS) (pH 10.0), injection time 25 s at a height of 17 cm and separation voltage at 12.5 kV, the AA-I and AA-II were baseline separated within 5 min. Low detection limits for AA-I and AA-II were 4.0 x 10(-8) mol L(-1) and 1.0 x 10(-7) mol L(-1), respectively. Wide linear ranges were from 4.0 x 10(-8) mol L(-1) to 1.9 x 10(-5) mol L(-1) and 1.0 x 10(-7) mol L(-1) to 5.0 x 10(-5) mol L(-1) for AA-I and AA-II, respectively. The proposed method has been successfully applied to analyze AAs contents in plant extracts. The results indicated that the contents of AAs in each part of Aristolochia debilis Sieb. Et Zucc. plant were different. Meanwhile, the CE-ED method was utilized for fingerprint analysis of medicine herbs. Six herbs (Radix aristolochiae, Fructus aristolochiae, Herba aristolochiae, Caulis aristolochiae manshuriensis, Caulis clematidis armandii, Caulis akebiae) were well distinguished by comparing their electropherograms obtained by CE-ED method.     

Characterization of derivatization of sialic acid with 2-aminoacridone and determination of sialic acid content in glycoproteins by capillary electrophoresis and high performance liquid chromatography--ion trap mass spectrometry.

A simple and highly sensitive capillary electrophoresis (CE) method for determining the content of N-acetylneuraminic acid (Neu5Ac) in glycoproteins was developed. Neu5Ac was derivatized with 2-aminoacridone (AMAC) by reductive amination, and the AMAC-Neu5Ac adduct could be readily separated from the other 11 AMAC-derivatized neutral and acidic monosaccharides usually present in glycoproteins by CE in a 0.3 mol/L borate buffer, pH 10.5, and detected at 260 nm. The derivatization of Neu5Ac was achieved at 55 degrees C for 4 h. AMAC-Neu5Ac was stable at 20 degrees C in the dark for at least 12 h while at room temperature it spontaneously converted into another substance with a lower electrophoretic mobility, which was identified as decarboxylated AMAC-Neu5Ac by high performance liquid chromatography - ion trap mass spectrometry (HPLC-ITMS). Concentration and mass of Neu5Ac as low as 1 micromol/L and 35 fmol could be detected. The linear correlation coefficient between the ratio of peak area to migration time of AMAC-Neu5Ac and the concentration of Neu5Ac ranging from 10 to 120 micromol/L was 0.9978 (n=8). This method was successfully applied to the analysis of sialic acid in human urinary trypsin inhibitor (hu-UTI), bovine alpha1-acid glycoprotein (alpha1-AGP) and recombinant human erythropoietin (rhu-EPO). By combination of CE and HPLC-ITMS we found that N-glycolylneuraminic acid (Neu5Gc) was present in bovine alpha1-AGP in addition to Neu5Ac, with a quantity comparable to that of the latter.     

High-throughput capillary electrophoretic method for determination of total aminothiols in plasma and urine.

Increased interest in the analysis of aminothiols in body fluids during the last years results in a request for high-throughput analytical methods for their determination. We report here a novel, high-throughput method for the determination of total concentrations of biogenous aminothiols - homocysteine, cysteine, glutathione, cysteinylglycine, gamma-glutamylcysteine, and of penicilamine, mercaptopropionylglycine, and cysteamine, three compounds used to treat disorders of aminothiol metabolism in plasma and urine. Samples were reduced with tris(carboxyethyl)phosphine and labeled with 5-(bromomethyl)fluorescein. Capillary electrophoretic separations were performed in 60 mmol/L borate - 15 mmol/L sodium dodecyl sulfate - 2-amino-2-methyl-1-propanol, pH 10.0, with laser-induced fluorescence detection. Analysis time was less than 2 min. The assay is linear (r > 0.999) up to 500 micromol/L. Reproducibilities of migration times (coefficient of variation, CV) were < 0.5%. Interassay repeatabilities (CV, n = 10) were 5.08% and 6.09% for 5 micromol/L addition of homocysteine and 0.60% and 3.78% for 100 micromol/L addition of cysteine in plasma and urine, respectively. Recovery values were within 94-106% and sensitivity was better than 0.19 micromol/L for all analyzed compounds. Results agreed well with a standard high-performance liquid chromatography (HPLC) method. The diagnostic usefulness of the method has been proven on 79 samples of cystinuric patients and 12 samples of homocystinuric patients. We report here a novel method for the determination of aminothiols in body fluids by capillary electrophoresis (CE). Determination is fast and sensitive enough for diagnostic purposes.     

毛细管电泳安培检测技术进展

对毛细管电泳离柱和柱端安培检测方式、不同形式电极在安培检测中的应用、安培检测在芯片毛细管电泳中的应用、安培检测池等内容进行了总结和讨论 ,并预测了安培检测技术未来发展方向     

Determination of myo-inositol phosphates in food samples by flow injection-capillary zone electrophoresis

A capillary electrophoresis (CE) method with indirect photometric detection was developed to identify and quantify myo-inositol phosphates in food samples. A flow-injection (FI) system including a micro-column containing anionic exchange resin was used for the solid-phase extraction of the myo-inositol phosphates with a view to their preconcentration. The FI system was automatically coupled to CE equipment via a mechanical interface. The overall analysis time was shortened by incorporating an FI system for myo-inositol hexakisphosphate monitoring. The limit of detection for myo-inositol phosphates as determined by FI-CE ranged from 11 to 26 micromol/L and the coefficient of variation from 3.9 to 5.0%. On the other hand, the limit of detection and coefficient of variation for myo-inositol hexakisphosphate as monitored by the FI system were 75 micromol/L and 2.9%, respectively. The proposed method was successfully applied to a variety of food samples with recoveries ranging from 96.0 to 107.7% and the precision from 3.9 to 7.9%. Based on the results, the content of myo-inositol hexakisphosphate in nuts was two or three times higher than that in legumes.     

Determination of catechin isomers in human plasma subsequent to green tea ingestion using chiral capillary electrophoresis with a high-sensitivity cell

A simple and reliable analytical electrophoretic method using chiral capillary electrophoresis (CCE) with a high-sensitivity cell of special design has been established for simultaneous determination of (+)-catechin (C) and (-)-epicatechin (EC) in aqueous and human plasma media. The application of a capillary with high-sensitivity cell has led to an improvement of 10-fold and 5-fold time-corrected peak area over a standard cell and a capillary with bubble cell, respectively. Analysis has involved the electrophoretic separation of C and EC in less than 4.0 min at 210 nm. The running buffer consist of 50.0 mmol L(-1) borate buffer with 1.0 mmol L(-1) beta-cyclodextrin at pH 8.5. CCE system has been proved for its intended use by applying procedure starting from calibration of CE instrument into validation of all experimental parameters. The resolution between catechin isomers under optimal conditions has been found to be more than 3.0. The detection limits of C and EC have been calculated to be 3.2 and 1.0 ng mL(-1), respectively. Good linearity has been obtained with correlation coefficient (r(2)) ranging between 0.995 and 0.996 at 99% confidence level (CL). Application of the proposed method to human plasma after ingestion of green tea has successfully been achieved and has statistically been proved. The unchanged amounts of C and EC in plasma were about 17.4 and 1.8% of the administered dose after 2 h of starting tea ingestion. The detection limits of C and EC in human plasma at 210 nm were 4.1 and 1.5 ng mL(-1), respectively.     

New technique for capillary electrophoresis directly coupled with end-column electrochemiluminescence detection

A new end-column electrochemiluminescence (ECL) detection technique coupling to capillary electrophoresis (CE) is characterized. A 300 microm diameter Pt working electrode was used to directly couple with a 75 microm inner diameter separation capillary without an electric field decoupler. The hydrodynamic cyclic voltammogram (CV) of Ru(bpy) 3 2+ showed that electrophoretic current did not affect the ECL reaction. The presence of high-voltage (HV) field only resulted in the shift of the ECL detection potential. The distance of capillary to electrode was an important parameter for optimizing detection performance as it determined the characteristics of mass transport toward the electrode and the actual concentration of Ru(bpy) 3 2+ in the detection region. The optimum distance of capillary to electrode was decided by the inner diameter of the capillary, too. For a 75 microm capillary, the working electrode should be placed away from the capillary outlet at a distance within the range of 220-260 microm. The effects of pH value of ECL solution and molecular structure of analytes on peak height and theoretical plate numbers were discussed. Using the 75 microm capillary, under the optimum conditions, the method provided a linear range for tripropylamine (TPA) between 1 x 10(-10) and 1 x 10(-5) mol/L with correlation coefficient of 0.998. The detection limit (signal-to-noise ratio S/N = 3) was 5.0 x 10(-11) mol/L. The relative standard deviation in peak height for eight consecutive injections was 5.6%. By this new technique lidocaine spiked in a urine sample was determined. The method exhibited the linear range for lidocaine from 5.0 x 10(-8) to 1.0 x 10(-5) mol/L with correlation efficient of 0.998. The limit of detection (S/N = 3) was 2.0 x 10(-8) mol/L.     

Analysis of flavonoids in <Emphasis Type="Italic">Ginkgo biloba</Emphasis> L. and its phytopharmaceuticals by capillary electrophoresis with electrochemical detection

A high-performance capillary electrophoretic (CE) method with electrochemical detection (ED) has been developed for determination of the pharmacologically active flavonoids in Ginkgo biloba L. and phytopharmaceuticals containing its extract. Epicatechin, catechin, rutin, apigenin, luteolin, and quercetin are important flavonoids in this plant. Operated in a wall-jet configuration, a 300 micro m diameter carbon-disk electrode was used as working electrode with good response to the six analytes at +1000 mV (relative to the SCE). Under the optimum conditions, the analytes were separated within 22 min in a borax buffer (pH 9.0). Excellent linearity was obtained over two orders of magnitude and detection limits (S/N=3) ranged from 1.4 x 10(-7) to 5.0 x 10(-7) g mL(-1) for all six analytes. The method was successfully used for assay of Ginkgo biloba L. and its phytopharmaceuticals after a relatively simple extraction procedure; the results obtained were satisfactory.     

Capillary electrophoresis/electrochemical detection system with on-line deoxygenation

A capillary electrophoresis (CE)/electrochemical detection system with on-line deoxygenation was developed, consisting of a deoxygenation injector, a deoxygenation protector, and an electrochemical detection cell. When the system was utilized for 60 min, the steady-state current of oxygen detected could be dropped to 3% of the original value for the gold/mercury amalgam electrode and to 8% of the original value for the gold electrode, and the limit of detection could be decreased two orders of magnitude for the reducible analytes such as TI+ (from 3.1 x 10-5 mol/L to 8.0 x 10-7 mol/L) and metronidazole (from 3.8 x 10-5 mol/L to 4.0 x 10-7 mol/L).     

Analysis of serotonin in brain microdialysates using capillary electrophoresis and native laser-induced fluorescence detection

Serotonin or 5-hydroxytryptamine (5-HT) is a major neurotransmitter in the central nervous system. In this work, a method for analyzing 5-HT in brain microdialysis samples using a commercially available capillary electrophoresis (CE) system has been developed. A pH-mediated in-capillary preconcentration of samples was performed, and after separation by capillary zone electrophoresis, native fluorescence of 5-HT was detected by a 266 nm solid-state laser. The separation conditions for the analysis of 5-HT in standard solutions and microdialysates have been optimized, and this method has been validated on both pharmacological and analytical bases. Separation of 5-HT was performed using a 80 mmol/L citrate buffer, pH 2.5, containing 20 mmol/L hydroxypropyl-beta-cyclodextrin (HP-beta-CD) and +30 kV voltage. The detection limit was 2.5 x 10(-10) mol/L. This method allows the in vivo brain monitoring of 5-HT using a simple, accurate CE measurement in underivatized microdialysis samples.