Determination of hydroxyl radical by capillary electrophoresis and studies on hydroxyl radical scavenging activities of Chinese herbs

High-performance capillary electrophoresis (CE) with electrochemical detection (ED) was employed to determine hydroxyl radicals in the Fenton reaction. Hydroxyl radicals can react with salicylic acid to produce 2,3-dihydroxy benzoic acid and 2,5-dihydroxy benzoic acid, which can be analyzed by CE-ED. Based on this principle, hydroxyl radicals were determined indirectly. In a 20 mmol/L phosphate running buffer (pH 7.4), 2,3-dihydroxy benzoic acid and 2,5-dihydroxy benzoic acid would elute simultaneously from the capillary within 6 min. As the working electrode, a 300 m diameter carbon-disk electrode exhibits good responses at +0.60 V (vs. SCE) for the two analytes. Peak currents of the two analytes are additive. Excellent linearity was obtained in the concentration range from 1.0×10^-3 mol/L to 5.0×10^-6 mol/L for 2,3-dihydroxy benzoic acid. The detection limit (S/N=3) was 2.0×10^-6 mol/L. This method was successfully applied for studying hydroxyl radical scavenging activities of Chinese herbs. It is testified that Apocynum Venetum L., Jinkgo bibola L., Morus alba L. and Rhododendron dauricum L. have strong hydroxyl radical scavenging activities.     

Determination of ionization constants of saccharides by capillary zone electrophoresis with amperometric detection

Summary A method based on a linear model enabling the efficient determination of the ionization constants (K _a) of saccharides by capillary zone electrophoresis with amperometric detection has been demonstrated. TheK _a values obtained from the plots of the reciprocal effective mobility against the inverse concentration of sodium hydroxide were in agreement with literature values.     

毛细管电泳安培法测定脂可平胶囊中的姜黄素

采用毛细管电泳柱端安培检测对脂可平胶囊中的姜黄素进行测定。着重研究了缓冲溶液浓度和酸碱度、检测电位、进样时间和高压对分离测定的影响。以微Pt电极为工作电极,电极电位为 1.0 V,以V(甲醇)∶V(乙醇)∶V(水)=5∶2∶3为非水介质,磷酸二氢钾和硼砂(pH 9.5)为缓冲体系,并用二阶样条小波进行滤波处理,姜黄素在1.0~120 mg/L范围内,峰高与其质量浓度呈良好的线性关系,线性回归方程:Y=20.2 146ρ,检出限为0.02 mg/L。     

毛细管电泳安培法检测酚类化合物

使用自行设计组装的毛细管电泳柱端安培检测系统 ,对四个酚类化合物进行了分离检测。研究了工作电极、缓冲液及其 p H值、检测电压和分离电压对分离检测的影响。在优化条件下 ,4个酚在 5× 1 0 -6～ 5× 1 0 -4 mol/L范围内峰高与浓度成良好的线性关系 ,检测下限为 8.5× 1 0 -7mol/L     

Simultaneous determination of dihydroxybenzene and phenylenediamine positional isomers using capillary zone electrophoresis coupled with amperometric detection

In general capillary zone electrophoresis (CZE) separation models, o‐, m‐, and p‐phenylenediamine isomers can be separated in a weak acidic running buffer for their pK_a values being 4.52, 5.64, 6.04, respectively, while o‐, m‐, and p‐dihydroxybenzene isomers can be separated in a weak basic buffer for their pK_a values being 9.40, 9.40 and 10.04, respectively. So, it is hard to find a suitable running buffer at a fixed pH in normal CZE for simultaneous separation of these two groups of positional isomers. In this paper, a novel method based on alternately running two different pH buffers in CZE coupled with amperometric detection (CZE‐AD) was designed to simultaneously determine these two groups of positional isomers. It is found that when two different pH running buffers were employed alternately under appropriate order and time, the six analytes could be separated perfectly in less than 20 min and the detection limits were as low as 10^–7 mol/L. Furthermore, the effects of working electrode potential, pH and concentration of running buffer, separation voltage and injection time on CZE–AD were investigated. Experimental results demonstrated that the introduced method was practical to simultaneously determine two groups of positional isomers with different pK_a and had some advantages of high sensitivity, good repeatability and small sample requirement.     

Determination of enrofloxacin and its metabolite ciprofloxacin by high performance capillary electrophoresis with end-column amperometric detection

Enrofloxacin (ENR) and its metabolite ciprofloxacin (CIP) were determined by capillary zone electrophoresis (CZE) with end-column amperometric detection. The effect of several factors, such as pH and concentration of running buffer solution, separation voltage, injection time, and working potential, on CZE were investigated to establish the optimal conditions of separation and detection. Under a given set of conditions (pH 8.00 phosphate buffer solution (20 mmol/L); +0.95 V for the working potential; 18 kV for the separation voltage; sample injection at 18 kV for 10 s), the compounds investigated can be well separated and detected within 8 min. Excellent linearity was observed between peak currents and concentration of analytes in the range from 0.034 to 70.0 mg/kg for these two compounds. The detection limits (S/N= 3) for enrofloxacin and ciprofloxacin were 13.68 mg/kg and 14.35 mg/kg, respectively, which were about 7-fold lower than the maximum residue limits (MRLs) established by the European Union. A simple sample pretreatment method was developed and proved to be effective in obtaining good recoveries and short analysis time. The developed CE-AD method was simpler, faster, and less cost intensive than other reported methods, and allows the determination of ENR and its metabolite CIP in contaminated eel liver samples and other animal tissue samples at the required maximum residue limits.     

Simultaneous determination of phenylenediamine isomers and dihydroxybenzene isomers in hair dyes by capillary zone electrophoresis coupled with amperometric detection

The simultaneous determination of three isomers of phenylenediamines (o, m, and p-phenylenediamine) and two isomers of dihydroxybenzenes (catechol and resorcinol) in hair dyes was performed by capillary zone electrophoresis coupled with amperometric detection (CZE–AD). The effects of working electrode potential, pH and concentration of running buffer, separation voltage, and injection time on CZE–AD were investigated. Under the optimum conditions the five analytes could be perfectly separated in 0.30 mol L⁻¹ borate–0.40 mol L⁻¹ phosphate buffer (pH 5.8) within 15 min. A 300 μm diameter platinum electrode had good responses at +0.85 V (versus SCE) for the five analytes. Their linear ranges were from 1.0 × 10⁻⁶ to 1.0 × 10⁻⁴ mol L⁻¹ and the detection limits were as low as 10⁻⁷ mol L⁻¹ (S/N = 3). This working electrode was successfully used to analyze eight kinds of hair dye sample with recoveries in the range 91.0–108.0% and RSDs less than 5.0%. These results demonstrated that capillary zone electrophoresis coupled with electrochemical detection using a platinum working electrode as detector was convenient, highly sensitive, highly repeatable and could be used in the rapid determination of practical samples. Figure Electropherograms obtained from 10 mg mL⁻¹ hair dye sample solutions at a platinum working electrode under optimum CZE–AD conditions: (a) natural black (I), (b) golden: (1) p-phenylenediamine, (2) m-phenylenediamine, (3) o-phenylenediamine, (4) resorcinol, and (5) catechol     

Separation and determination of flavonoids in three traditional chinese medicines by capillary electrophoresis with amperometric detection

Flavonoids are important active ingredients in many traditional Chinese medicines. In this paper, capillary electrophoresis with amperometric detection was employed to separate and detect eight flavonoids, rutin, quercetrin, quercetin, kaempferol, kaempferide, catechin, apigenin, and luteolin, in a home‐made capillary electrophoresis device. Under the separation voltage of 2000 V, the eight flavonoids could be completely separated within 33 min in 18 mM borax running buffer at pH 10.2. Good linear relationships were obtained for all analytes and the detection limits for flavonoids ranged from 0.46 to 0.85 μM. Then, the method was applied to separate and determine the flavonoids in three traditional Chinese medicines, hippophae rhamnoides, hypericum perforatum, and cacumen platycladi. Finally, rutin, kaempferol, quercetin, and quercetrin were discovered in these medicines and the concentrations ranged from 0.28 to 9.94 mg/g. The recoveries of flavonoids ranged from 84.7 to 113%, which showed the high reliability of this method.     

Determination of four kinds of carbamate pesticides by capillary zone electrophoresis with amperometric detection at a polyamide-modified carbon paste electrode

In this paper, a polyamide-modified carbon paste electrode in capillary zone electrophoresis with amperometric detection (CZE-AD) was firstly applied to the determination of four carbamate pesticides: fenobucarb, isoprocarb, metolcarb and carbaryl. The four carbamates were hydrolyzed in alkalescent aqueous solutions, resulting in the formation of 2-sec-butylphenol, 2-isopropylphenol, m-cresol and α-naphthol, which could be determined by amperometry after capillary electrophoretic separation. Under the selected optimum conditions, the four analytes could be perfectly separated within 23 min. The linear ranges of 2-sec-butylphenol, 2-isopropylphenol and m-cresol were from 1.0 × 10⁻⁷ to 2.0 × 10⁻⁵ mol L⁻¹ and that of α-naphthol was from 2.0 × 10⁻⁷ to 2.0 × 10⁻⁵ mol L⁻¹ and their detection limits were 3.0 × 10⁻⁸, 3.0 × 10⁻⁸, 3.0 × 10⁻⁸ and 6.0 × 10⁻⁸ mol L⁻¹, respectively (S/N = 3). Fenobucarb, isoprocarb, metolcarb and carbaryl can be indirectly determined by this CZE-AD method with recovery of 105, 104, 110 and 98% and R.S.D. of 4, 3, 4 and 3%, respectively. Above results demonstrated that this method was of high sensitivity, good repeatability and could be used in the rapid determination of the pesticide residues.     

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.     

Determination of the compositions of polysaccharides from Chinese herbs by capillary zone electrophoresis with amperometric detection

In this paper, capillary zone electrophoresis with amperometric detection (CZE-AD) was applied to determine the compositions of hetero-polysaccharides from Chinese herbs, Angelica sinensis and flax by analyzing their hydrolyzed monosaccharides: fucose, galactose, glucose, arabinose, rhamnose and xylose. Under the selected optimum conditions, the six monosaccharides could be perfectly separated within 25 min and showed significant current responses at copper electrodes. The linear ranges of the six monosaccharides were all from 5.0 x 10(-6) to 2.0 x 10(-4) mol L(-1) and their detection limits were lower or near 1.0 x 10(-6) mol L(-1) (S/N = 3). Experiments showed that the Angelica sinensis polysaccharide was composed of fucose, galactose, glucose, arabinose, rhamnose and xylose (mole ratio 1.0:13.6:15.0:8.7:21.3:3.7), and the flax polysaccharide was composed of galactose, glucose and arabinose (mole ratio 1.0:4.98:1.1). The purity of these polysaccharides leached by the introduced leaching method was 98.3 and 97.6%, respectively. Analyzing polysaccharides by this method has some merits of speed, simple instrumentation and operation, high sensitivity and high reproducibility.     

Study on sensitivity development of 2,3-dihydroxybenzoic acid by capillary zone electrophoresis-amperometric detection with p-methyl benzoate as stacking agent

For its high reactivity and very short half-life, the hydroxyl radical (OH.) in vivo is very difficult to be detected. Usually, it is indirectly quantified by determining 2,3-dihydroxybenzoic acid (2,3-DHBA) and 2,5-dihydroxybenzoic acid (2,5-DHBA), which are the reaction products of salicylic acid (SA) and OH.. Because 2,5-DHBA could be directly formed by the P(450) enzyme, only 2,3-DHBA is regarded as the real biomarker of OH.in biological studies. But the very low concentration of OH* in human bodies makes its determination very difficult and complicated. In this paper, a simple online stacking capillary zone electrophoresis coupled with amperometric detection (CZE-AD) method was explored to improve the detection sensitivity of 2,3-DHBA to reach the requirements in biological analysis. A mixture solution of 12.5 mmol/L Na(2)B(4)O(7)-25 mmol/L NaH(2)PO(4) (pH 7.9) was used as the running buffer and p-methyl benzoate was selected as a suitable stacker. The effects of the concentration, pH value, and injection time of p-methyl benzoate on stacking efficiency were carefully studied. Under the optimum stacking CZE-AD conditions, the detection sensitivity of 2,3-DHBA was improved about 20-fold and its detection limit reached the 10(-9) mol/L level. The experimental results showed that this was a potential method to determine OH* in vivo.     

Simultaneous determination of flavonoids in chrysanthemum by capillary zone electrophoresis with running buffer modifiers

Despite the separation efficiency of capillary electrophoresis (CE) is much higher than other chromatographic methods, it is sometimes difficult to perfectly separate the complex ingredients in biological samples. One possible and simple way to develop the separation effect in CE is to add some modifiers in the running buffer. In this paper, the suitable running buffer modifiers were explored to simultaneously separate and detect six typical flavonoids (apigenin, luteolin, kaempferol, quercetin, (+)-catechin and (−)-epicatechin) which are the main active ingredients in chrysanthemum by capillary zone electrophoresis with amperometric detection (CZE-AD). It was found that when β-cyclodextrin (β-CD) and the mixture of methanol and ethanol were used as running buffer modifiers, a baseline separation of the six analytes could be accomplished in less than 20 min and the detection limits were as low as 10⁻⁷ or 10⁻⁸ g ml⁻¹. Other factors affecting the CZE separation, such as working potential, pH value and ionic strength of running buffer, separation voltage and sample injection time were extensively investigated. Under the optimum conditions, a successful practical application on the determination of chrysanthemum samples confirmed the validity and practicability of this method.     

毛细管电泳柱端安培检测装置的研制

研制了一种新型的毛细管电泳柱端型安培检测装置。以直径为6μm的碳纤维微电极为工作电极,在自组装的ACS-2000毛细管电泳仪上,考察了用不同内径毛细管分离时分离电压对背景噪声的影响。利用该装置同时测定了3种苯二酚的异构体。     

Fast determination of sugars in Coke and Diet Coke by miniaturized capillary electrophoresis with amperometric detection

The fast separation capability of a novel miniaturized capillary electrophoresis with amperometric detection (CE-AD) system was demonstrated by determining sugar contents in Coke and diet Coke with an estimated separation efficiency of 60,000 TP/m. Factors influencing the separation and detection processes were examined and optimized. The end-capillary 300 microm Cu wire amperometric detector offers favorable signal-to-noise characteristics at a relatively low potential (+0.50 V vs. Ag/AgCl) for detecting sugars. Three sugars (sucrose, glucose, and fructose) have been separated within 330 s in a 8.5 cm length capillary at a separation voltage of 1000 V using a 50 mM NaOH running buffer (pH 12.7). Highly linear response is obtained for the above compounds over the range of 5.0 to 2.0 x 10(2) microg/mL with low detection limit, down to 0.8 microg/mL for glucose (S/N = 3). The injection-to-injection repeatability for analytes in peak current (RSD < 3.6%) and for migration times (RSD < 1.4%) was excellent. The new miniaturized CE-AD system should find a wide range of analytical applications involving assays of carbohydrates as an alternative to conventional CE and micro-CE.     

Simultaneous determination of electroactive and non-electroactive food preservatives by novel capillary electrophoresis with amperometric detection

A novel capillary electrophoresis and amperometric detection method was achieved by adding an electroactive additive (3,4-dihydroxybenzylamine, 3,4-DHBA) to the running buffer, so that both electroactive and non-electroactive food preservatives were simultaneously determined. Under the selected optimum conditions, four electroactive preservatives (methylparaben, ethylparaben, propylparaben and butylparaben) and two non-electroactive preservatives (potassium sorbate and sodium lactate) were well separated and sensitively detected with detection limits (S/N = 3) ranging from 1.06 × 10⁻⁸ to 2.73 × 10⁻⁶ g mL⁻¹. This method has been successfully employed for the determination of both electroactive and non-electroactive preservatives in several food commodities.     

Study on Rhizoma Chuanxiong based on capillary electrophoresis with amperometric detection

<正>A high-performance capillary electrophoresis with amperometric detection(CE-AD) method has been developed for the analysis of seven bioactive ingredients,namely ferulic acid(FA),vanillin,vanillic acid,p-hydroxybenzoic acid,caffeic acid,gallic acid and protocatechuic acid,in Rhizoma Chuanxiong.The effects of several factors such as the acidity and concentration of running buffer,the separation voltage,the applied potential to working electrode and the injection time were investigated.Under the optimum conditions,the seven analytes could be well separated within 21 min at the separation voltage of 16 kV in a 60 mmol/L borax running buffer(pH 8.7).A 300μm diameter carbon disk electrode has a good response at potential of +950 mV(vs.SCE) for all analytes.Good linear relationship was established over three orders of magnitude with detection limits(S/N = 3) ranged from 3.3×10~(-7) to 6.7×10~(-9)g/mL.This proposed method has been successfully applied for the determination and comparison of different batches of Rhizoma Chuanxiong samples based on their characteristic electrochemical profiles.     

A new sequential injection analysis‐capillary electrophoresis system with amperometric detection

A novel and fully automated sequential injection analysis manifold coupled to a capillary electrophoresis apparatus with amperometric detection, is described. The sequential injection manifold was isolated from the high voltage by inserting an air plug into the circuit. Small buffer reservoirs were used to avoid the need to pump fresh buffer to the interface during the electrophoretic separation. No decoupling device was used to mitigate the interference from the high voltage electric field, instead the potential shift induced by the separation voltage, was accounted for. The new hydrodynamic injection method presented is based on the overpressure created in the circuit when a pinch valve is closed for a predetermined time. The injection method yields RSD values of peak height and area below 2.55 and 1.82%, respectively, at different durations of valve closure (n = 5). The capillary and working electrode alignment was achieved by adapting a commercial available capillary union. When the electrode was replaced, the alignment method proved to be very reliable, yielding RSD values of peak height and area lower than 2.64 and 2.08%, respectively (n = 8). Using this system with a gold microelectrode, dopamine, and epinephrine could be quantified within the concentration range of 1–500 μM and detected at a concentration of 0.3 μM. The methods here presented could be applied for the development of new capillary electrophoresis systems with amperometric detection and/or to the design of fully automated systems for online process monitoring purposes.     

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.     

Fabrication and performance of a three-dimensionally adjustable device for the amperometric detection of microchip capillary electrophoresis

A microchip CE-amperometric detection (AD) system has been fabricated by integrating a two-dimensionally adjustable CE microchip and an AD cell containing a one-dimensionally adjustable disk detection electrode in a Plexiglas holder. It facilitates the precise 3-D alignment between the channel outlet and the detection electrode without a complicated 3-D manipulator. The performance of this unique system was demonstrated by separating five aromatic amines (1,4-phenyldiamine, aniline, 2-methylaniline, 4-chloroaniline, and 1-naphthylamine) of environmental concern. Factors influencing their separation and detection processes were examined and optimized. The five analytes have been well separated within 140 s in a 74 cm long separation channel at a separation voltage of +2500 V using a 10 mM phosphate buffer (pH 3.5). Highly linear response is obtained for the five analytes over the range 20-200 microM with the detection limits ranging from 0.46 to 1.44 microM, respectively. The present system demonstrated long-term stability and reproducibility with RSDs of less than 5% for the peak current (n = 9). The new approach for the microchannel-electrode alignment should find a wide range of applications in CE, flowing injection analysis, and other microfluidic analysis systems.