Determination of Active Ingredients of Origanum Vulgare L. and Its Medicinal Preparations by Capillary Electrophoresis with Electrochemical Detection

Abstract

A method based on capillary electrophoresis with electrochemical detection (CE‐ED) has been developed for the first time for the separation and determination of isovanillic acid, vanillic acid, quercetin, rosmarinic acid, caffeic acid, and protocatechuic acid in Origanum vulgare L. and its medicinal preparations. The effects of working electrode potential, pH level, concentration of running buffer, separation voltage, and injection time on CE‐ED were investigated. Under the optimum conditions, the analytes could be separated in a 50 mmol L^?1 borate buffer (pH 8.7) within 21 min. A 300‐µm diameter carbon disk electrode has a good response at +0.95 V (vs. SCE) for all analytes. The response was linear over three orders of magnitude with detection limits (S/N=3) ranging from 4×10^?8 g mL^?1 to 2×10^?7 g mL^?1 for the analytes. The method has been successfully applied to the analysis of real sample, with satisfactory results.     

毛细管电泳-电化学检测法测定中药杠版归中的活性成分

采用毛细管电泳-电化学检测法（CE-ED）同时分离测定了中药杠板归中的阿魏酸、香草酸、槲皮素、咖啡酸、原儿茶酸等主要生物活性成分的含量。考察了工作电极电位、运行缓冲液的pH值和浓度、分离电压和进样时间等实验参数对实验结果的影响。在优化的实验条件下,以直径300 μm的碳圆盘电极为工作电极,检测电位为＋0.95V(vs. SCE),在10mmol/L磷酸盐（pH 9.2）的运行缓冲溶液中,五个分析物能够在17min内实现很好的基线分离,被测物浓度与峰电流在3个数量级呈良好的线性,检测限（S/N＝3）范围从7.1×10-8 到 9.3×10-8g mL-1。该方法已应用于实际样品的分析,样品处理简单,获得了令人满意的结果。     

Determination of Active Ingredients of Hawthorn by Capillary Electrophoresis with Electrochemical Detection

A method based on capillary electrophoresis with electrochemical detection has been developed for the separa-tion and determination of epicatechin,kaempferol,chlorogenic acid,4-hydroxybenzoic acid,quercetin and proto-catechuic acid in hawthorn for the first time.The effects of working electrode potential,pH and concentration ofrunning buffer,separation voltage and injection time on CE-ED were investigated.Under the optimum conditions,the analytes could be separated in a 60 mmol·L~(-1) borate buffer(pH 8.7)within 21 min.A 300 μm diameter carbondisk electrode has a good response at 0.95 V(vs.SCE)for all analytes.The response was linear over three ordersof magnitude with detection limits(S/N=3)ranging from 3×10~(-8) to 2×10~(-7) g·mL~(-1) for the analytes.The methodhas been successfully applied to the analysis of real sample,with satisfactory results.     

毛细管区带电泳法分析绿茶中的痕量成份

采用毛细管电泳/安培检测法(CE/AD)同时分离测定了绿茶中的芦丁、没食子酸、槲皮素、绿原酸等生物活性成分的含量, 考察了运行缓冲液酸度、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。在最优化条件下, 以300 μm碳圆盘电极为检测电极, 检测电位为+ 950 mV (vs. SCE) , 60 mmol/L硼酸盐运行缓冲液(pH 8.7)中, 上述各组分在20 min内可实现基线分离。各组分浓度与峰电流在3个数量级范围内呈良好线性, 检出限(S/N=3)在1.0×10^-7到1.0×10^-4g^.mL^-1范围,四种标样7次平行进样的相对标准偏差(RSD)小于3.0 %。该方法已成功地应用于绿茶中生物活性成分的测定, 结果令人满意。     

Study on the Effect of Moderate Exercise on Lactic Acid Content in Breast Milk by Indirect CE with Amperometric Detection

An indirect high-performance capillary electrophoresis with amperometric detection (CE-AD) method has been developed for determination of lactic acid (LA) in body fluids of lactating postpartum women. Several important factors such as the running buffer additive and concentration, the working electrode potential, the pH value and concentration of the running buffer, the separation voltage and injection time were investigated. Under the optimum conditions, LA could be well separated with co-existing interferences including uric acid (UA) in real samples in a 90-cm-length capillary at separation voltage of 12 kV in 4.0 × 10^?6 g mL^?1 3,4-dihydroxybenzylamine (DHBA)/40 mmol L^?1 H₃BO₃?CNa₂B₄O₇ buffer (pH 7.8). The linearity between peak current and concentration of LA was over three orders of magnitude with detection limit of 5.00 × 10^?7 g mL^?1 (S/N = 3). This proposed method has been successfully used to study the effects of moderate exercise on LA content in breast milk and urine samples of lactating postpartum women, and assay results showed that LA content in breast milk can return to normal level through 60 min rest without decreasing acceptance by breast-feeding infants, although the LA level did increase by 4?C6 times in both breast milk and urine samples at 10 min after moderate exercise.     

Simultaneous Determination of Phenolic Additives in Cosmetics by Micellar Electrokinetic Capillary Chromatography with Electrochemical Detection

A micellar electrokinetic capillary chromatography method with electrochemical detection (MECC‐ED) has been developed for the simultaneous determination of eight phenolic additives, including propyl gallate (PG), tert‐butylhydroquinone (TBHQ), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), methylparaben (MP), ethylparaben (EP), propylparaben (PP) and butylparaben (BP) in cosmetic products. Method development involved optimization of the working electrode, the pH value of running buffer, the concentration of sodium dodecyl sulfate (SDS), the separation voltage, and the sample injection time. Under the optimum conditions, all analytes can be well separated within 26 min at the separation voltage of 18 kV in a 9 mmol·L^?1 sodium dodecyl sulfate (SDS) ?60 mmol·L^?1 borate running buffer (pH 8.0). A 300 μm diameter carbon disk electrode generated good response at +0.90 V (vs. SCE) for all analytes. Linearity of the present method was over three orders of magnitude of analyte concentration with detection limits (S/N=3) ranging from 1.1×10^?7 to 1.2×10^?6 g·mL^?1 for all analytes. This proposed method has been successfully applied to the simultaneous determination of the above additives in commercial cosmetics, and the assay results were satisfactory.     

Capillary Electrophoresis of the Active Ingredients of Dioscorea bulbifera L. and its Medicinal Preparations

A method based on capillary electrophoresis with electrochemical detection has been developed for the separation and determination of epicatechin, isovanillic acid, vanillic acid and myricetin in Dioscorea bulbifera L. and its medicinal preparations. The effects of working electrode potential, pH and concentration of running buffer, separation voltage and injection time were investigated. Under optimum conditions, the analytes could be separated in a 40 mmol L^?1 borate buffer (pH 8.7) within 15 min. A 300 μm diameter carbon disk electrode has a good response at + 0.95 V (vs. SCE) for all analytes. The response was linear over three orders of magnitude with detection limits (S/N = 3) ranging from 3.0 × 10^?8 g mL^?1 to 1.0 × 10^?7 g mL^?1. The method has been successfully applied to the analysis of real samples.     

CE-ED Separation and Determination of Seasonal Content Variations of Some Active Ingredients in Toona sinensis (A. Juss) Roem Leaves

A method based on capillary zone electrophoresis with electrochemical detection (CE-ED) has been employed for the separation and determination of epicatechin, scopoletin, quercetin and gallic acid in Toona sinensis (A. Juss) Roem leaves. Seasonal variations of these ingredients in T. sinensis (A. Juss) Roem leaves collected in April, June and August, were discussed. Several important factors including running buffer acidity, separation voltage, working electrode potential and etc., were evaluated to acquire optimum analysis conditions. Under the selected optimum conditions, these analytes could be well separated. Good linear relationship was established between the peak current and the concentration of each analyte over 2–3 orders of magnitude with detection limits (S/N = 3) ranging from 4.00 × 10^?8 to 1.73 × 10^?7g mL^?1. The proposed method has been successfully applied for the evaluation of the seasonal content variation of these four active ingredients in T. sinensis (A. Juss) Roem leaves with satisfactory results.     

Analysis of Atorvastatin and Related Substances by MEKC

A new micellar electrokinetic capillary chromatographic (MEKC) method has been developed for simultaneous quantitation of atorvastatin (AT) and its related substances. The separation was carried out in an extended light path capillary at applied voltage of 30 kV using a background electrolyte consisting of 10 mM sodium tetraborate buffer pH 9.5, 50 mM sodium dodecyl sulphate and 20% (v/v) methanol. The addition of methanol to the running buffer resulted in a very effective choice to achieve resolution between the peaks of charged substances adjacent to AT as well as the peaks of neutral drug-related substances. Linear calibration curves were established over the concentration range 100–1,200 μg mL^?1 for AT and 1.0–12.5 μg mL^?1 for related substances. The proposed MEKC procedure has been validated with respect to selectivity, precision, linearity, limits of detection, and quantitation, accuracy and robustness. The method has been successfully applied to the determination of AT and purity evaluation of bulk drug and formulated products.     

Determination of Four Phenolic Compounds in Scirpus yagara Ohwi by CE with Amperometric Detection

A method based on capillary electrophoresis with amperometric detection has been developed for the determination of trans-resveratrol, scirpusin A, scirpusin B, and p-hydroxycinnamic acid in the rhizomes of Scirpus yagara Ohwi. The effects of the acidity and the concentration of the running buffer, separation voltage, injection time, and detection potential were investigated to acquire the optimum conditions. The detection electrode was a 300 μm diameter carbon disc electrode at a detection potential of +0.90 V. The four analytes could be well separated within 12 min in a 40 cm length fused silica capillary at a separation voltage of 12 kV in a 50 mM borate buffer (pH 9.2). The relation between peak current and analyte concentration was linear over about three orders of magnitude with detection limits (S/N = 3) ranging from 32.2 to 63.4 μg L⁻¹ for the four analytes.

     

Determination of Bioactive Flavonoids in Rhododendron Dauricum L. by Capillary Electrophoresis with Electrochemical Detection

A method based on capillary electrophoresis with electrochemical detection (CE-ED) has been developed for the determination of bioactive flavonoids in the traditional Chinese herbal medicine, Rhododendron dauricum L. The effects of working electrode potential, pH and concentration of running buffer, separation voltage and injection time on CE-ED were investigated. Under the optimum conditions, the analytes could be separated in a 70 mmol L^?1 borate buffer (pH=9.2) within 20 min. A 300 μm diameter carbon disk electrode has a good response at + 0.90 V (vs. SCE) for all analytes. The response was linear over three orders of magnitude with detection limits (S/N = 3) ranging from 2 × 10^?8 g mL^?1 to 2 × 10^?7 g m^?1 for the analytes. The method has been successfully applied for the analysis of real sample, with satisfactory results.     

CE Determination of Creatinine and Uric Acid in Saliva and Urine During Exercise

A simple and reliable method based on capillary electrophoresis with electrochemical detection (CE–ED) was applied to study the effect of aerobic exercises on creatinine and uric acid concertration in saliva and urine. The pH value, the running buffer concentration, the SDS concentration, separation voltage, injection time and the potential applied to the working electrode were investigated to find the optimum conditions. The detection limits (S/N = 3) for creatinine and uric acid were 3.6 μmol L^?1 and 0.86 μmol L^?1, respectively. This method was successfully used in the rapid analysis of creatinine and uric acid in saliva samples. After aerobic exercises, creatinine concentration decreased, and uric acid concentration increased in saliva. In urine, the concentrations of creatinine and uric acid both increased after exercise.     

Optimization,Validation and Application of a Capillary Zone Electrophoresis Method for the Assay of Sparfloxacin in Pharmaceutical Formulation

Abstract

A capillary zone electrophoresis (CZE) method has been developed for the determination of the antibiotic sparfloxacin in tablets. The CZE separation was performed using 75 µm×35 cm fused-silica capillary under the following conditions: 25°C; applied voltage, 12 kV; 25 mM H₃PO₄-NaOH running buffer (pH 8.5). The detection wavelength was 254 nm. Flumequine was used as internal standard (IS). The method was suitably validated with respect to linearity, limit of detection and quantification, accuracy, precision, specificity, and robustness. The calibration was linear from 10 to 60 µg mL^?1 and the limit of detection and quantification were 5.38 and 9.46 µg mL^?1, respectively. Recoveries ranging from 95.68%–102.4% were obtained in the determination of sparfloxacin that were spiked to placebos. Excipients in the commercial tablets and degraded products from different stress conditions did not interfere in the assay. The method was successfully applied to the determination of sparfloxacin in pharmaceutical tablets.     

Adsorptive BSA Coating Method for CE to Separate Basic Proteins

A novel and simple coating method was developed by coating bovine serum albumin (BSA) onto the inner surface of a fused-silica capillary, to avoid the adsorption of analytes during CE. The advantage presented here was that the coating process is more simple, fast, stable, and reproducible. The coated capillary avoided the adsorption of analytes onto the inner surface of a fused-silica capillary and might be a promising candidate for separation of complex biological samples with further development. Meanwhile, the efficiencies of the coated capillary were evaluated by EOF, chromatographic peak shape, and theoretical plate number (N m^?1) of RNase A. The optimal coating conditions were obtained from the results. The pH value of coating buffer PB was 4.2, the standing time was 12 h at 4 °C, and the coating concentration of BSA was 1.5 mg mL^?1. The stability of the coating on the inner wall of the capillary and the reproducibility of the coated capillaries were good. The theoretical plate number values of RNase A were over 1.3 × 10⁵ (N m^?1) in the coated capillary. After successive electrophoresis for 48 h using the coated capillary, the RSD values of EOF and the theoretical plate number were 4.14 % and 9.14 %, respectively. In addition, the RSD values of EOF and the theoretical plate number (N m^?1) in the coated capillaries were 13.19 % and 8.96 %, respectively. Finally, the coated capillary was successfully applied to separate the mixture of four basic proteins (RNase A, lysozyme, trypsin and myoglobin).     

Determination of Sulfonamides in Pharmaceuticals and Rabbit Plasma by Microchip Electrophoresis with LED-IF Detection

In this paper, we describe a compact and low-cost light-emitting diode-induced fluorescence (LED-IF) detection coupled to microchip electrophoresis for the determination of sulfonamides in pharmaceutical formulations and rabbit plasma. Three fluorescein isothiocyanate-labeled sulfonamides in rabbit plasma were separated in the running buffer of 40 mM phosphate buffer (pH 7.0) at the separation voltage of 2.0 kV, and detected by LED-IF detector in which the high-power blue LED was driven at the constant current of 150 mA and the emitted fluorescence over 510 nm was collected by a planar photodiode. The linear concentration ranged from 2.0 to 125.0 μg mL^?1, both for sulfadiazine and sulfamethazine with the correlation coefficients (r ²) of 0.995 and 0.997, respectively, and from 2.0 to 100.0 μg mL^?1 with the correlation coefficients (r ²) of 0.997 for sulfaguanidine. The limits of detection for the three sulfonamides were 0.36–0.50 μg mL^?1 (S/N = 3). Intra-day and inter-day precision of migration time and peak area for the determination of sulfonamides were <4.5 %. This method has been successfully applied to the analysis of sulfonamides in pharmaceuticals, and could be used to study the pharmacokinetics of sulfonamides in rabbit.     

Simultaneous Analysis of Sugars and Polyphenols in Tobacco by CZE with Amperometric Detection Based on a Cu2O/MWCNT-COOH Composite Electrode

A composite electrode was fabricated from Cu₂O powder, carboxyl-functionalized multi-wall carbon nanotubes (MWCNT-COOH), and paraffin oil in the proportions 51:17:32 (w/w). This composite electrode was used for amperometric detection (CZE–AD) in simultaneous capillary zone electrophoretic analysis of chlorogenic acid, rutin, sucrose, glucose, mannose, and fructose in tobacco samples. Under the optimum conditions, the six analytes could be separated in 100 mmol L^?1 NaOH buffer within 30 min. Good linearity was achieved in the range 1 × 10^?7–1 × 10^?4 mol L^?1 for the two polyphenols and 5 × 10^?6–1 × 10^?3 mol L^?1 for the four sugars. The detection limits (S/N = 3) for the polyphenols and sugars were as low as 10^?8 mol L^?1 and 10^?6 mol L^?1, respectively.     

Ultrasonic-Assisted Drop-to-Drop Solvent Microextraction in a Capillary Tube for Analyzing Trace Benzene, Toluene, Xylene in One Drop of a Water Sample

A novel analytical technique termed ultrasonic-assisted drop-to-drop solvent microextraction (USA-DDSME) in a capillary tube was developed to determine trace benzene, toluene, xylene in one drop of a water sample, which was combined with gas chromatography–flame ionization detection (GC–FID). The advantages of this method are rapidity, convenience, ease of operation, simplicity of the device, and extremely little solvent and sample consumption. Extraction conditions including the type of extraction solvent, the volume of extraction solvent, the volume of sample, extraction time and effect of salt concentration were optimized. The best optimum parameters for extraction were achieved with 3 μL of extraction solvent. Chloroform was divided into four equal divisions in 20 μL water sample (without salt addition) in a capillary tube and ultrasonicated for 10 min, centrifugated at 2,500 rpm for 5 min to let the extraction solvent settle at the bottom of the capillary tube, then 1 μL of the separated extraction solvent was injected into the GC–FID for analysis. Linearity of the method was determined by analyzing spiked water samples over a concentration range of 0.1–50 μg mL^?1. Correspondingly, the LOD values were 0.01 μg mL^?1. All calibration curves were found to have good linearity with correlation coefficients (r ²) > 0.995. The precision (RSD) of the system, measured by six repeated determinations of the analytes at 1 μg mL^?1 were in the range of 1.6–3.5%.     

Nonextractive Trace Level Simultaneous Determination of Mercury(II) and Zinc(II) in Environmental Samples with 2‐(5‐Bromo‐2‐pyridylazo)‐5‐diethylaminophenol and Cetylpyridinium Chloride

Abstract

A simple, rapid, selective, and sensitive method for the derivative spectrophotometric determination of Hg(II) and its simultaneous determination in the presence of Zn(II) using 2‐(5‐bromo‐2‐pyridylazo)‐5‐diethylaminophenol in the presence of cetylpyridinium chloride, a cationic surfactant, has been developed. The molar absorption coefficient and analytical sensitivity of the 1∶1 Hg(II) complex at 558 nm (λ_max) are 5.78×10⁴ L mol^?1 cm^?1 and 0.67 ng mL^?1, respectively. The detection limit of Hg(II) is 1.40×10^?2 ng mL^?1, and Beer's law is valid in the concentration range 0.05–2.40 µg mL^?1. Overlapping spectral profiles of Hg(II) and Zn(II) complexes in zero‐order mode interfere in their simultaneous determination. However, 0.10–2.00 µg mL^?1 of Hg(II) and 0.065–0.650 µg mL^?1 of Zn(II), when present together, can be simultaneously determined at zero cross point of the derivative spectrum, without any prior separation. The relative standard deviation for six replicate measurements of solutions containing 0.134 µg mL^?1 of Hg(II) and 0.620 µg mL^?1 of Zn(II) is 1.72 and 1.47%, respectively. The proposed method has successfully been evaluated for trace level simultaneous determination of Hg(II) and Zn(II) in environmental samples.     

Simultaneous Determination of Theobromine,Paraxanthine, Theophylline,and Caffeine in Urine by Reversed-Phase High-Performance Liquid Chromatography with Diode Array UV Detection

A reversed-phase high performance liquid chromatographic method was improved for the simultaneous determination of theobromine, paraxanthine, theophylline, and caffeine in urine. The method includes a liquid-liquid extraction at alkaline pH with ethylacetate. The 7-(2,3-dihidroxypropyl) theophylline was used as an internal standard (ISTD). The separation was achieved on a C₁₈ column using 14:86 methanol:buffer (25 mM KH₂PO₄ adjusted to pH 4 with ortho-phosphoric acid) solution as mobile phase under isocratic conditions at a flow rate 1 mL min^?1. An ultraviolet absorption at 274 nm was monitored. In these conditions, the LOD was 0.03 μg mL^?1 for theobromine, 0.02 μg mL^?1 for paraxanthine, 0.04 μg mL^?1 for theophylline, and 0.08 μg mL^?1 for caffeine. The method has been applied to urine samples.     

On-Line Two-Dimension Liquid Chromatography for the Analysis of Ingredients in the Medicinal Preparation of Coptis Chinensis Franch

An on-line two-dimension microflow liquid chromatography was developed for better separation and analysis of the highly complex ingredients of medicinal preparation of traditional Chinese medicine Coptis Chinensis Franch. A two-valve switching system was utilized for two-dimension chromatography with strong cation exchange and reverse-phase capillary columns separation. The components were separated well by this system and yielded over 420 peaks. Under the optimal condition, 4 compounds were detected quantitatively. A good linear relationship was obtained from 0.2 µg mL^?1 to 24 µg mL^?1with detection limits (S/N = 3) ranging from 0.05 µg mL^?1 to 0.2 µg mL^?1for the compounds. We demonstrated that the method can be successfully applied to the analysis of a natural complex sample, with satisfactory results.