Microchip capillary electrophoresis/electrochemical detection of hydrazine compounds at a cobalt phthalocyanine modified electrochemical detector

This article reports on the use of cobalt(II) phthalocyanine (CoPc)-modified carbon paste amperometric detector for monitoring hydrazine compounds following their microchip separation. The marked catalytic electrochemical properties of CoPc-modified electrode display enhanced sensitivity compared with unmodified carbon pastes at a relatively low detection potential (+0.5 V versus Ag/AgCl). Factors influencing the on-chip separation and detection processes have been optimized. Three hydrazines (hydrazine, 1,1 dimethylhydrazine, and phenylhydrazine) have been separated within 130 s at a separation voltage of 1 kV using a 10 mM phosphate run buffer (pH 6.5). The detection limits obtained from using the CoPc-modified carbon paste electrodes for hydrazine and phenylhydrazine are 0.5 and 0.7 μM, respectively, with linearity over the 20–200 μM range examined. Such miniaturization and speed advantages of microchip CE are coupled to the highly sensitivity and convenient preparation of CoPc-modified carbon paste electrode. The resulting microsystem should be attractive for field monitoring of toxic hydrazine compounds in environmental applications.     

Microchip capillary electrophoresis with a boron-doped diamond electrochemical detector for analysis of aromatic amines

The attractive features of a boron-doped diamond (BDD) thin-film detector for microchip capillary electrophoretic (CE) separations of dye-related amino-substituted aromatic compounds are described. The diamond electrode was employed in the end-column amperometric detection of 4-aminophenol (4-AP), 1,2-phenylenediamine (1,2-PDA), 2-aminonaphthalene (2-AN), 2-chloroaniline (2-CA), and o-aminobenzoic acid (o-ABA), and its attractive behavior was compared to commonly used screen-printed carbon and glassy-carbon electrodes. These conventional electrode materials exhibit a significant degree of passivation and low sensitivity to the above-mentioned environmental pollutants. The diamond-based electrochemical detection system displayed a favorable analytical performance, including lower noise levels, higher peak resolution with enhanced sensitivity, and improved resistance against electrode passivation. Factors influencing the on-chip analysis were assessed and optimized. The diamond detector displayed detection limits of 2.0 and 1.3 microM for 4-AP and 2-AN, respectively, and a wide linear response for these compounds over the 2-50 microM range. The enhanced stability was demonstrated by relative standard deviation (RSD) values of 1.4% and 4.7% for 100 microM 1,2-PDA and 200 microM 2-CA, respectively, for repetitive detections (n = 7). Besides, the simultaneously observed current decrease was 2.4 and 9.1% for 1,2-PDA and 2-CA, respectively (compared to 21.8 and 41.0% at the screen-printed carbon electrode and 28.3 and 34.1% at the glassy carbon electrode, respectively). The favorable properties of the diamond electrode indicate great promise for environmental applications in CE and other microchip devices.     

Microchip capillary electrophoresis with electrochemical detector for fast measurements of aromatic amino acids

A method based on microchip capillary electrophoresis with amperometric detection was developed for the rapid separation and direct detection of oxidizable aromatic amino acids (without prior derivatization). The working electrode was a thick-film carbon strip electrode positioned opposite the outlet of the separation channel. Factors influencing the separation and detection processes were examined and optimized. The five aromatic amino acids, tyrosine, 5-hydroxytryptophan, tryptophan, p-aminobenzoic acid, and m-aminobenzoic acid, can be well separated within 5 min using a separation voltage of 2000 V and a 25 mM phosphate buffer (pH 7.0) run buffer containing 50 mM sodium dodecylsulfate. Most favorable amperometric detection was obtained at +0.95 V. Linear calibration plots are observed for micromolar concentrations of the oxidizable amino acids. The new protocol offers good stability and for reproducibility, with relative S.D. of less than 5% for both migration times and peak currents (n=8). It should be useful for the analysis of aromatic amino acids, as desired for life sciences.     

Microchip capillary electrophoresis with amperometric detection for several carbohydrates

Microchip capillary electrophoresis (μCE) with amperometric detection at Cu electrode benefited fast separation and direct detection of carbohydrates. The working electrode of 50-μm Cu wire attached nearly against the channel outlet—4 μm, where it benefited collecting detection current and suppressing overwhelming noise. The use of alkaline medium was essential to separating and detecting carbohydrates, which dissociated into the sensitive alcolate anions. The 10-cm serpentine chip, though lengthening the migration time, it provided better efficiency. Sucrose, cellobiose, glucose, and fructose migrated from the outlet in 400 s +2000 V. The linear calibration plots ranging from 10 to 1000 μM with regression coefficients better than 0.996 were obtained. The injection-to-injection reproducibility of 1.24% (n=7) for glucose in peak current and 0.6% for migration times were excellent. The detection limit was low, down to 2.3 μM for glucose (S/N=3) or 27.6 attomole in mass detection.     

Tear analysis of ascorbic acid,uric acid and malondialdehyde with capillary electrophoresis

Tears have a significant role in antioxidant defense in ocular tissues and since their collection is quick and noninvasive, their analysis would facilitate monitoring of pathophysiological changes. However, their low volume and low content of antioxidants makes analysis difficult; methods of high sensitivity are needed. In this paper, we present a method for tear analysis of two antioxidant molecules (ascorbic and uric acid) and of a lipid peroxidation indicator (malondialdehyde) with capillary electrophoresis. Tears were collected with Schirmer strips, extracted with a low‐pH phosphate buffer, centrifuged through membrane filters and an antioxidant was added. They were stable at ?70°C for 15 days. After pilot experiments, optimum electrophoretic separation was achieved in a 25 mm borate buffer, pH 10.0, containing 100 mm sodium dodecyl sulfate at 25°C and 20 kV. The developed method has good repeatability (<5% RSD), precision (<15% relative error values) and high sensitivity (LLOQ values of 20, 2.3 and 2.5 μM for ascorbate, urate and malondialdehyde, respectively). It was applied to the analysis of tears from healthy individuals and the antioxidant levels are in agreement with those obtained with other techniques. This method might serve as a tool to clarify the role of endogenous antioxidants in the pathophysiology of ocular diseases. Copyright © 2009 John Wiley & Sons, Ltd.     

Microchip capillary electrophoresis with a cellulose-DNA-modified screen-printed electrode for the analysis of neurotransmitters

A microfluidic chip based on capillary electrophoresis coupled with a cellulose-single-stranded DNA (cellulose-ssDNA) modified electrode was used for the simultaneous analysis of dopamine (DA), norepinephrine (NE), 3,4-dihydroxy-L-phenylalanine (L-DOPA), 3,4-dihydroxyphenylacetic acid (DOPAC), and ascorbic acid (AA). The modification of the electrode improved the electrophoretic analysis performance by lowering the detection potential and enhancing the signal-to-noise characteristic without surface poisoning of the electrode. The sensitivity of the modified electrode was about 12 times higher than those of the bare ones. The test compounds were separated using a 62 mm long separation channel at the separation field strength of +200 V/cm within 220 s in a 10 mM phosphate buffer (pH 7.4). The most favorable potential for the amperometric detection was 0.7 V (vs. Ag/AgCl). A reproducible response (relative standard deviation of 1.3, 1.3, 2.1, 3.1, 3.4% for DA, NE, L-DOPA, DOPAC, and AA, respectively, for n = 9) for repetitive sample injections reflected the negligible electrode fouling at the cellulose-ssDNA modified electrode. Square-wave voltammetric analyses reflected the sensitivities of the modified electrode for DA, NE, L-DOPA, DOPAC, and AA which were 1.78, 0.82, 0.69, 2.45, and 1.23 nC/microM with detection limits of 0.032, 0.93, 1.13, 0.31, and 0.62 microM, respectively. The applicability of this microsystem to real sample analysis was demonstrated.     

Microchip electrophoresis with chemiluminescence detection for assaying ascorbic acid and amino acids in single cells

A method based on microchip electrophoresis (MCE) with chemiluminescence (CL) detection was developed for the determination of ascorbic acid (AA) and amino acids including tryptophan (Trp), glycine (Gly) and alanine (Ala) present in single cells. Cell injection, loading, lysing, electrophoretic separation and CL detection were integrated onto a simple cross microfluidic chip. A single cell was loaded in the cross intersection by electrophoretic means through applying a set of potentials at the reservoirs. The docked cell was lysed rapidly under a direct electric field. The intracellular contents were MCE separated within 130 s. CL detection was based on the enhancing effects of AA and amino acids on the CL reaction of luminol with K₃[Fe(CN)₆]. Rat hepatocytes were prepared and analyzed as the test cellular model. The average intracellular contents of AA, Trp, Gly and Ala in single rat hepatocytes were found to be 38.3, 5.15, 3.78 and 3.84 fmol (n = 12), respectively.     

毛细管电泳免疫分析法研究癌胚抗原与抗体相互作用

采用毛细管电泳免疫分析法研究癌胚抗原和抗体相互作用.探讨了缓冲体系、癌胚抗原和抗体的配比、进样时间,进样电压等因素对分离检测的影响.结果表明分离电压为14 kV,进样时间为10 s, 在pH值为5.92的Tris-乙酸缓冲体系（TAE）中, 癌胚抗原及其复合物得到较满意的分离.     

Microchip micellar electrokinetic chromatography coupled with electrochemical detection for analysis of synthetic oestrogen mimicking compounds

A miniaturised analytical system for separating and detecting a range of steroidal oestrogens, based on the coupling of a micromachined capillary electrophoresis chip with glassy-carbon electrode amperometric detector, is described. Factors influencing the on-chip separation utilising the technique micellar electrokinetic chromatography (MEKC) and detection processes are optimised. The addition of modifiers such as organic solvents and surfactants improve separation and resolution of these hydrophobic compounds. Using a borate running buffer (5 mM, pH 11) with 20% methanol and SDS (20 mM) and a separation voltage of 2000 V, baseline resolution is observed for 16-keto-17β-oestradiol, oestriol, 11β-hydroxyoestradiol, oestrone, and β-oestradiol in 420 s with limits of detection 16–84 μM. The implications for on-site environmental analysis are discussed.     

Simultaneous electrochemical and electrochemiluminescence detection for microchip and conventional capillary electrophoresis

A simultaneous electrochemical (EC) and electrochemiluminescence (ECL) detection scheme was introduced to both microchip and conventional capillary electrophoresis (CE). In this dual detection scheme, tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy)3(2+)) was used as an ECL reagent as well as a catalyst (in the formation of Ru(bpy)3(3+)) for the EC detection. In the Ru(bpy)3(2+)-ECL process, Ru(bpy)3(3+) was generated and then reacted with analytes resulting in an ECL emission and a great current enhancement in EC detection due to the catalysis of Ru(bpy)3(3+). The current response and ECL signals were monitored simultaneously. In the experiments, dopamine and three kinds of pharmaceuticals, anisodamine, ofloxacin, and lidocaine, were selected to validate this dual detection strategy. Typically, for the EC detection of dopamine with the presence of Ru(bpy)3(2+), a approximately 5 times higher signal-to-noise ratio (S/N) can be achieved than that without Ru(bpy)3(2+), during the simultaneous EC and ECL detection of a mixture of dopamine and lidocaine using CE separation. The results indicated that this dual EC and ECL detection strategy could provide a simple and convenient detection method for analysis of more kinds of analytes in CE separation than the single EC or ECL detection alone, and more information of analytes could be achieved in analytical applications simultaneously.     

Simplified current decoupler for microchip capillary electrophoresis with electrochemical and pulsed amperometric detection

There is a need to develop broadly applicable, highly sensitive detection methods for microchip CE that do not require analyte derivatization. LIF is highly sensitive but typically requires analyte derivatization. Electrochemistry provides an alternative method for direct analyte detection; however, in its most common form, direct current (DC) amperometry, it is limited to a small number of easily oxidizable or reducible analytes. Pulsed amperometric detection (PAD) is an alternative waveform that can increase the number of electrochemically detectable analytes. Increasing sensitivity for electrochemical detection (EC) and PAD requires the isolation of detection current (nA) from the separation current (muA) in a process generally referred to as current decoupling. Here, we present the development of a simple integrated decoupler to improve sensitivity and its coupling with PAD. A Pd microwire is used as the cathode for decoupling and a second Au or Pt wire is used as the working electrode for either EC or PAD. The electrode system is easy to make, requiring no clean-room facilities or specialized metallization systems. Sensitive detection of a wide range of analytes is shown to be possible using this system. Using this system we were able to achieve detection limits as low as 5 nM for dopamine, 74 nM for glutathione, and 100 nM for glucose.     

Assay for the simultaneous determination of guanidinoacetic acid, creatinine and creatine in plasma and urine by capillary electrophoresis UV-detection

Guanidinoacetic acid (GAA) measurement has recently become of great interest for the diagnosis of creatine (Cn) metabolism disorders, and research calls for rapid and inexpensive methods for its detection in plasma and urine in order to assess a large number of patients. We propose a new assay for the measurement of GAA by a simple CZE UV-detection without previous sample derivatization. Plasma samples were filtered by Microcon-10 microconcentrators and directly injected into the capillary, while for urine specimens a simple water dilution before injection was needed. A baseline separation was obtained in less than 8 min using a 60.2 cm x 75 microm uncoated silica capillary, 75 mmol/L Tris-phosphate buffer pH 2.25 at 15 degrees C. The performance of the developed method was assessed by measuring plasma creatinine and Cn in 32 normal subjects and comparing the data obtained by the new method with those found with the previous CE assay. Our new method seems to be an inexpensive, fast and specific tool to assess a large number of patients both in clinical and in research laboratories.     

Chemiluminescence detection coupled to capillary electrophoresis

In recent years, chemiluminescence (CL)-based detection coupled to capillary electrophoresis (CE) as separation technique has attracted much interest due to new advances in home-made configurations, sample-treatment techniques for application to real matrixes, development of a commercial instrument and use of miniaturization techniques to obtain micro total analysis systems incorporating CE separation and CL detection in microchips. We present some developments, key strategies and selected analytical applications of CE-CL since the year 2000 in diverse fields (e.g., clinical and pharmaceutical, environmental or food analysis).     

Simultaneous determination of aminothiols, ascorbic acid and uric acid in biological samples by capillary electrophoresis with electrochemical detection

A method based on capillary electrophoresis with electrochemical detection has been employed for the separation and determination of homocysteine, cysteine, reduced glutathione, ascorbic acid and uric acid. Effects of several important factors such as the acidity and concentration of the running buffer, separation voltage, injection time and detection potential were investigated to acquire the optimum conditions. The detection electrode was a 500 microm diameter platinum disk electrode at a working potential of +1.05 V (vs saturated calomel electrode). The five analytes were well separated within 10 min in a 50 cm long fused silica capillary at a separation voltage of 18 kV in a 100 mm phosphate buffer (pH 7.8). The relation between peak current and analyte concentration was linear over about 3 orders of magnitude with the detection limits (S/N = 3) ranging from 0.83 to 2.58 microm. The proposed method was successfully applied to determine cysteine, reduced glutathione, ascorbic acid and uric acid in human whole blood and rat brain tissues with satisfactory assay results and should find a wide range of bioanalytical applications.     

高效毛细管区带电泳法快速测定尿液中的肌酐

建立了一种快速测定尿中肌酐浓度的高效毛细管电泳方法。利用非涂层石英毛细管(64.5 cm×50μm i.d),以pH 2.5,0.1 mmol/L H3PO4作为电泳缓冲液,检测波长191 nm,用0.05 Pa压力进样4 s,在电压16 kV快速分离尿液中的肌酐,采用外标法定量。肌酐的迁移时间约为5.5 min,肌酐浓度在34.5~8840μmol/L范围内呈良好的线性(r2=0.999)。平均日内精密度为2.5%,日间精密度为3.0%。回收率94.1%~99.0%。与全自动生化分析仪碱性苦味酸速率法相比有良好的相关性(r=0.990,n=56)。高效毛细管电泳法测定尿肌酐可应用于临床样品的检测。     

Determination of barbituric acid and 2-thiobarbituric acid with end-column electrochemical detection by capillary electrophoresis

Capillary electrophoresis (CE) with end-column electrochemical detection (EC) of barbituric acid (BA) and 2-thiobarbituric acid (TA) has been described. Under optimum condition, BA and TA were separated satisfactorily, and a response of high sensitivity and stability was obtained at a detection potential of 1.25 V versus Ag/AgCl. Optimized end-column detection provides detection limit as low as 0.5 and 0.1 μM for BA and TA, respectively. The calibration graph was linear over three orders of magnitude. The relative standard deviations (n=10) of peak currents and migration times obtained for both BA and TA were 3.4, 3.7, and 1.7, 1.2%, respectively. The proposed method has been applied to analyze water sample with satisfactory results.     

An electrochemical detector cell for open tubular liquid chromatography and capillary electrophoresis

Summary An electrochemical detector cell has been developed for micro-flow separation systems (OTLC, CE). The cell contains two electrodes, a disk-shaped working electrode made from a carbon fiber bundle, and a tubular Ag/AgCl quasi-reference electrode. The effective cell volume and the coulometric yield have been determined, for different electrode diameters and at different flow rates, in an OTLC system. An effective cell volume of less than 1 nl was observed. The applicability of the cell was demonstrated with the detection of OPA-derivatized amino acids. For use in CE, the cell was equipped with an additional compartment, housing a semi-permeable joint for the decoupling of the high electric field used for the electrophoretic separation. Results are shown on the determination of catecholamines by CE with electrochemical detection. Detection limits with both OTLC and CE were well below 1 fmole.     

Cationic amylopectin derivatives as additives for analysis of proteins in capillary electrophoresis

Positively charged amylopectin, which is a major constituent of cationic starch, was used to modify the inner surface of fused-silica capillaries by addition to the running solution, which was subsequently employed in CE. Capillaries filled with cationic amylopectin derivatives were shown to generate a stable reversed EOF in the investigated range of pH 4-8. Among the additives studied, quaternary ammonium amylopectin derivatives with high amino and low hydroxypropyl groups showed fast electroosmotic mobility and very effectively suppressed the adsorption of proteins. The run-to-run and batch-to-batch repeatability of the procedures were satisfactory with RSDs of 0.5% and 2.4%, respectively. A basic protein, alpha-chymotrypsinogen, migrated within 6 min and the theoretical plate number of it reached 560 000 plates/m.     

Characterization of sepia ink in ancient graphic documents by capillary electrophoresis

A simple and rapid capillary electrophoresis with diode array detection method was developed for sepia ink identification in ancient graphic documents.Separation was performed in a fused-silica capillary (64.5 cm length, 50 μm i.d.). The running buffer was 20 mM sodium tetraborate solution, pH 9.2. The applied potential was 25 kV, temperature 25 °C and detection was at 220 nm. An appropriate extraction procedure was applied for the take and treatment of sample from the reference substances and ancient graphic documents. This method was successfully applied to the collection of drawings and maps from the Royal Chancellery Archives of Granada (Spain).     

毛细管电泳在完整哺乳动物细胞分析中的应用

作为人体形态结构、生理功能和生长发育的基本单位,细胞的结构功能及行为分析具有重要的研究价值。毛细管电泳作为日渐成熟的微量分析技术在细胞分析及应用方面已取得显著进展。本文综述了毛细管电泳在完整哺乳动物细胞分析中的应用,包括群体细胞分析和完整单细胞分析。所述内容涉及血红细胞、公猪精子、人宫颈癌细胞、人神经母细胞瘤细胞、人结直肠腺癌细胞、人慢性髓系白血病细胞以及鼠小脑颗粒细胞。总结了完整哺乳动物细胞分析的毛细管电泳方法和条件,归纳分析了完整细胞分析中存在的细胞破碎、聚集、沉降、吸附及电泳异质性等关键问题及解决方法。对毛细管电泳在细胞分析方面未来的应用方向进行了展望。综述文献49篇。