Quantitative determination of glutathione in single human erythrocytes by capillary zone electrophoresis with electrochemical detection

Glutathione (GSH) in single human erythrocytes is determined by capillary zone electrophoresis with end-column amperometric detection at a gold/mercury amalgam microelectrode. A capillary of 10 microm inner diameter is suitable for determination of GSH in an individual erythrocyte with a good signal-to-noise ratio. The limit of detection is 1 x 10(-7) mol/L or 26 amol and the linear dynamic range is 2 x 10(-7) to 2 X 10(-5) mol/L for the capillary. In this method, the calibration line is obtained with a capillary adsorbed before a certain amount of hemoglobin can be used for the quantification of GSH in the external standardization. The whole cell injection and the lack of necessity of a derivatization reaction lead to more accurate and precise results, which are closer to the macroscopic values of glutathione in human red blood cell (i.e., hemolysate) than those determined by indirect laser-induced fluorescence detection.     

Determination of diclofenac sodium by capillary zone electrophoresis with electrochemical detection

Capillary zone electrophoresis was employed for the determination of diclofenac sodium using an end-column amperometric detection with a carbon fiber microelectrode, at a constant potential of 0.83 V vs. saturated calomel electrode. The optimum conditions of separation and detection are 4.90 x 10(-3) mol/l Na2HPO4-3.10 x 10(-3) mol/l NaH2PO4 (pH 7.0) for the buffer solution, 10 kV for the separation voltage, 5 kV and 10 s for the injection voltage and the injection time, respectively. The limit of detection is 2.5 x 10(-6) mol/l or 5.2 fmol (S/N=2). The relative standard deviation is 0.8% for the migration time and 4.7% for the electrophoretic peak current. The method was applied to the determination of diclofenac sodium in human urine.     

Analysis of amino acids by capillary zone electrophoresis with electrochemical detection

The precapillary derivatization of 20 amino acids with naphthalene-2,3-dicarboxaldehyde (NDA) and CN(-) was investigated. All these derivatized amino acids could be oxidized on the carbon fiber microdisk bundle electrode except proline. Capillary zone electrophoresis with electrochemical detection was employed for the analysis of 19 amino acids. The optimum conditions of separation and detection were borate, pH 9.48, for the electrolyte, 18 kV for the separation voltage and 1.15 V versus a saturated calomel electrode for the detection potential. Limits of detection of concentration or mass for individual amino acids were between 1.7 x 10(-7) and 1.8 x 10(-6) mol/L or 84 and 893 amol (according to the signal-to-noise ratio of 3) for the injection voltage of 6 kV and injection time of 10 s. The relative standard deviations were between 0.80 and 2.3% for the migration times and 1.4 and 6.4% for the electrophoretic peak currents. From a mixture of 19 amino acids, 10 amino acids (Arg, Lys, Orn, Try, Ser, Ala, Gly, Cys, Glu, Asp) could be well separated. The other 9 amino acids appeared on three electrophoretic peaks. From the samples, in which the nine amino acids do not exist simultaneously, some of them could also be detected. The method was applied to the determination of amino acids in beer by the standard addition method. The recovery for the amino acids in beer was 91-109%.     

Sub-femtomole determination of DABSYL-amino acids with capillary zone electrophoresis separation and laser-induced thermo-optical absorbance detection

Fifteen amino acids are determined as the DABSYL (4-4-dimethylaminoazobenzene-4'-sulfonyl chloride) derivative with capillary zone electrophoresis separation and low-power laser-induced thermo-optical detection. Separation efficiencies are on the order of 200 000 theoretical plates and the detection limit, 3 s, is 200 attomole of glycine injected onto the column. At the detection limit, 0.7 attomole of glycine is present within the 40 picoliter detection volume.     

A capillary electrophoresis method for the determination of soluble monosaccharides in Ginkgo biloba leaves

A method for the simultaneous determination of six monosaccharides by pre-column derivatization with 1-phenyl-3-methyl-5-pyrazolone and capillary electrophoresis was developed in this work. The derivatization (i.e., reaction temperature, capillary electrophoresis duration, and extraction number) and separation (i.e., pH and buffer concentration) conditions for capillary electrophoresis were optimized. Results showed that the limits of detection under optimal conditions were in the range of 0.036–0.35 mg/L with a mean correlation coefficient >0.99. The recoveries were in the range of 87.3–108.49%, and the relative standard deviations of intra- and inter-day variations were in the ranges of 2.2–3.8 and 3.2–5.0%, respectively. The method was successfully applied to the analysis of six free monosaccharides in three types of Ginkgo biloba leaves.     

Analysis of monosaccharides by capillary electrophoresis with electrochemiluminescence detection.

A capillary electrophoresis with tris(2,2'-bipyridyl)ruthenium(II)-based electrogenerated chemiluminescence detection method was investigated for the determination of monosaccharides using 2-diethylaminoethanethiol as a derivatizing reagent. Xylose, rhamnose, glucose and glucosamine were selected for the demonstration of the method. Under optimal conditions, the calibration curve of glucose was linear over the range of 1.0 x 10(-4) to 1.0 x 10(-7) mol/L (R = 0.999) and the detection limit was 6.0 x 10(-8) mol/L (S/N = 3). The method was successfully applied for the assay of glucose in angelica.     

毛细管区带电泳法同时分离测定香兰素和邻位香兰素异构体

建立了同时测定香兰素和其异构体邻位香兰素的毛细管区带电泳法(CZE)。考察了缓冲溶液的种类、浓度和pH以及分离电压等因素对分离结果的影响。在缓冲溶液为50 mmol/L硼砂-150 mmol/L磷酸氢二钠(pH 7.5)、分离电压15 kV的优化条件下,6 min内即可实现分离。香兰素和邻位香兰素在10～240 mg/L范围内线性关系良好,相关系数分别为0.9999和0.9997;方法的检出限均为1.0 mg/L (信噪比为3);样品的加标回收率为99.4%～101.2%,相对标准偏差为0.19%～0.73%。该方法操作简单、快速,已应用于实际样品的分析,并获得了令人满意的结果。     

The separation and determination of nitrophenol isomers by high-performance capillary zone electrophoresis

o-Nitrophenol, m-nitrophenol, and p-nitrophenol could well be separated by capillary zone electrophoresis (CZE) by only adjusting the run buffer with methanol. Efficiency up to 10⁵ theoretical plates per meter was achieved. The effects of several important factors were investigated to find optimum conditions. The linear range, regression equation, and the recovery were given. This method possessed the advantages of simplicity, rapidity, and good reproducibility; it can be developed for the separation of practical samples in environment analysis.     

Sensitive determination of rhodamine 110-labeled monosaccharides in glycoprotein by capillary electrophoresis with laser-induced fluorescence detection

Rhodamine 110 (Rho110) has been used in the highly sensitive analysis of monosaccharides, as it reacts with the reducing carbonyl group of the saccharides. The monosaccharide derivatives were investigated by capillary electrophoresis with laser-induced fluorescence detection. The derivatization was performed at 90 °C for 30 min for all monosaccharides. The derivatized monosaccharides were separated using 200 mM borate (pH 10.5) as running buffer within 20 min. The fluorescence intensities of Rho110-derivatives were significantly decreased by the presence of excess reducing agent, but were greatly increased by the addition of potassium hexacyanoferrate(III). The concentration and mass detection limits for monosaccharides were in the range of 1.4–2.8 nM and 36–70 amol, respectively. We have applied this derivatization method to the analysis of the composition of monosaccharides in glycoproteins (ribonuclease B, fetuin, and erythropoietin) following their subjection to strong acid hydrolysis. The results from these analyses were in good agreements with the reported values established previously.     

Study of uptake kinetics of vincristine for human erythrocytes by capillary zone electrophoresis with electrochemical detection

Capillary zone electrophoresis (CZE) was employed for the determination of vincristine using electrochemical detection with a carbon fiber microdisk bundle electrode at a constant potential of 1.0 V versus saturated calomel electrode (SCE). The optimum conditions of separation and detection are 1.7×10⁻² Na₂HPO₄− 3.2×10⁻³ mol/l NaH₂PO₄ (pH 7.5) for the buffer solution, 20 kV for the separation voltage. The limit of detection is 5.0×10⁻⁷ mol/l or 2.2 fmol (S/N=3) for the injection voltage of 5 kV and the injection time of 10 s. The recovery of the method is between 95 and 101% for the vincristine taken by human erythrocytes. The method was applied to investigate uptake and accumulation behavior of vincristine for human erythrocytes. The advantages of the method are the small sample volume of CZE and the high selectivity and sensitivity of electrochemical detection.     

Separation and determination of nitroaniline isomers by capillary zone electrophoresis with amperometric detection

In this paper, capillary zone electrophoresis with amperometric detection (CZE-AD) was firstly applied to the simultaneous separation and determination of nitroaniline positional isomers. The three analytes could be perfectly analyzed by using the buffer of extreme pH. The effects of several important factors were investigated to find optimum conditions. A carbon-disk electrode was used as working electrode. The optimal conditions were 40 mmol/L tartaric acid-sodium tartrate (pH 1.2) as running buffer, 17 kV as separation voltage and 1.10 V (versus saturated calomel reference electrode, SCE) as detection potential. Under the optimum conditions, o-, m- and p-nitroaniline were separated successfully and good linearity, reproducibility and recovery results were obtained. The detection limit for m-nitroaniline was as low as at 9.06 × 10⁻⁹ mol/L. This proposed method demonstrated long-term stability and reproducibility with relative standard deviations of less than 1.8% for migration time and 1.1% for peak areas. The utility of this method was demonstrated by monitoring dyestuff wastewater and the assay results were satisfactory.     

Direct electrochemical determination of pyruvate in human sweat by capillary zone electrophoresis.

Capillary zone electrophoresis was employed for the determination of pyruvate in human sweat using electrochemical detection with a carbon fiber microdisk bundle electrode at a constant potential of 1.60 V vs. saturated calomel electrode. The optimum separation conditions are 3.6 x 10(-3) mol/L Na2HPO4-1.4 x 10(-3) mol/L NaH2PO4 (pH 7.2) for the buffer solution, and 18 kV for the separation voltage. The limits of detection of pyruvate are 8.0 x 10(-6) mol/L or 24 fmol (S/N = 3) for the injection voltage of 6 kV and the injection time of 10 s. The relative standard deviation is 2.0% for the migration time and 5.7% for the electrophoretic peak current. The method was applied to determining pyruvate in human sweat.     

Assay of amino acids in individual human lymphocytes by capillary zone electrophoresis with electrochemical detection

Amino acids in individual human lymphocytes were determined by capillary zone electrophoresis with electrochemical detection (ED) at a carbon fiber bundle electrode after on-column derivatization with naphthalene-2,3-dicarboxaldehyde (NDA) and CN⁻. In order to inject cells easily, a cell injector was designed. In this method, a single human lymphocyte and then the lysing/derivatizing buffer were electrokinetically injected into the front end of the separation capillary as a chamber to lyse the lymphocyte and derivatize amino acids in the cell. Four amino acids (serine (Ser), alanine (Ala), taurine (Tau), and glycine (Gly)) in single human lymphocytes have been identified. Quantitation has been accomplished through the use of calibration curves.     

Measurement of ascorbic acid in single human neutrophils by capillary zone electrophoresis with electrochemical detection

A method has been developed for the determination of ascorbic acid (AA) in individual human neutrophils by capillary zone electrophoresis with electrochemical detection at a carbon fiber microdisk bundle electrode. The natively easily oxidized substances such as glutathione, dopa, dopamine, serotonin, epinephrine do not interfere with the determination of ascorbic acid. A procedure of treating capillaries, which can overcome the influence of the adsorption of the substances in cells on the inner surface wall of the capillary on the migration time and the number of theoretical plates of interests, has been described. The average amount of AA in an individual neutrophil is 0.557 fmol, which is consistent with the literature value.     

瑞香狼毒多糖中单糖组成的毛细管区带电泳分析

采用1-苯基-3-甲基-5-吡唑啉酮(PMP)和1-(2-萘基)-3-甲基-5-吡唑啉酮(NMP)为衍生试剂,以反相高效液相色谱(RP-HPLC)和毛细管区带电泳(CZE)法对9种单糖衍生物进行了分离研究,对比确定了NMP衍生化CZE法在植物多糖单糖组成分析中的优越性。以NMP为柱前标记试剂,建立了毛细管区带电泳定量测定9种单糖的新方法。方法具有较好的重复性(相对标准偏差RSD小于4.3%),单糖衍生物检测限为0.85～1.6μmol/L,回收率为96.4%～104%。已应用于瑞香狼毒多糖样品的单糖组成分析。     

毛细管区带电泳-间接紫外法快速测定食品中的甜蜜素

建立了毛细管区带电泳-间接紫外法快速测定食品中甜蜜素的新方法。液体样品用超纯水稀释后直接进样;固体样品经粉碎或剪碎后用超纯水超声提取后离心,上清液直接进样或用水稀释后进样。以未涂敷石英毛细管（80 cm×75 μm,有效长度：70 cm）为分离柱,以2 mmol/L苯甲酸钠+10 mmol/L碳酸钠+0.5 mmol/L十六烷基三甲基溴化铵为分离缓冲液;于200 nm波长处检测。检出限为8.9 mg/kg （S/N=3）,定量限为26.7 mg/kg （S/N=9）。低、中、高添加水平的加标回收率分别为93.4%、100.3%及101.9%,相应的RSD分别为6.7%、2.0%及2.2%（n=5）。日内及日间精密度分别为2.6%和4.5%。整个分析过程无需有机溶剂。在能力验证样品的分析结果与国家标准方法的结果相吻合的基础上,分析了7件食品样品,获满意结果。     

毛细管区带电泳法拆分手性药物环扁桃酯

近年来,随着不同种类的手性添加剂[1]在毛细管电泳(CZE)中的使用,毛细管电泳越来越显示出其强有力的手性拆分性能。具有特殊笼状结构并含有多个手性中心的环糊精及其衍生物是毛细电泳手性分离研究中最常采用的手性添加添[2-4]。本文合成了环糊精衍生物单3 O 苯基胺甲酰基 β CD[2]并以之作为手性选择剂分离了β CD及手性药物环扁桃酯。1　实验部分932 3 HVPS高压电源(山东省化工研究院),DD 2000型可调波长紫外检测器(中国科学院大连化学物理研究所),XWT型记录仪(上海大华仪表厂),pHS 25型酸度计(上海雷磁仪器厂),石英毛细管45cm…     

Separation and determination of homovanillic acid and vanillylmandelic acid by capillary electrophoresis with electrochemical detection

A method of separation and determination of homovanillic acid (HVA) and vanillylmandelic acid (VMA) was developed based on capillary zone electrophoresis/amperometric detection with high sensitivity, good resolution and selectivity. In order to achieve complete separation and good response, several factors including pH, buffer concentration, separation voltage, detection potential and the length of separation capillary, were studied in detail. The method has been used to determine both HVA and VMA in human urine. Uric acid (UA) in human urine did not interference with their determination. The limit of detection of the method was 1.3×10⁻⁶ mol/l (1.4 fmol) for HVA and 7.9×10⁻⁷ mol/l (0.87 fmol) for VMA at a signal-to-noise ratio of 3.     

双通道-双工作电极毛细管电泳电化学法快速检测大黄酸

报道了一种测定大黄酸的快速毛细管电泳电化学方法。采用简易制作的一种双通道-双工作电极电化学系统,可以实现电导法和安培法同时检测;优化选择了缓冲介质、工作电极、检测电位、毛细管长度和内径以及分离电压等实验参数,并对提高分析速度进行了初步研究和探讨。结果表明:大黄酸在100s内可以得到较好的分离测定,电导法和安培法的线性范围分别为6.83×10-4～1.07×10-5mol/L和3.41×10-4～2.67×10-6mol/L,最小检出浓度分别为5.28×10-6mol/L和3.16×10-7mol/L。采用设计的双通道 双工作电极检测装置,可以充分发挥电导法和安培法的优越性,对样品峰及样品的纯度进行确证;另外通过采用较短的毛细管与适当提高分离电压,可以提高分析速度。该法已用于中药大黄中大黄酸的测定。     

Separation and determination of flavonoids in Agrimonia pilosa Ledeb. by capillary electrophoresis with electrochemical detection

Five flavonoids (catechin, hyperoside, quercitrin, quercetin, and rutin) were separated and determined by capillary electrophoresis with electrochemical detection. Effects of several important factors, such as the pH and concentration of running buffer, separation voltage, injection time, and detection potential were investigated to determine the optimum conditions. The five flavonoids were baseline separated within 20 min in a 60 cm length capillary at a separation voltage of 19.5 kV with a running buffer consisting of 60 mmoL/L Na2B4O7 - 120 mmoL/L NaH2PO4 (pH = 8.8). The relationship between peak current and analyte concentration was linear over about two orders of magnitude with detection limits (S/N = 3) ranging from 0.02 to 0.05 microg/mL for all compounds. This method was successfully used to determine the above five flavonoids in Agrimonia pilosa Ledeb. with relatively simple extraction procedures, and the assay results were satisfactory.