Application of co-electroosmotic capillary electrophoresis for the determination of inorganic anions and carboxylic acids in soil and plant extract with direct UV detection

Summary Indirect UV detection in capillary zone electrophoresis (CZE) is frequently used for the determination of inorganic anions and carboxylic acids. However, there are few reports on direct UV detection of these solutes in real samples. This paper describes the use of direct UV detection of inorganic anions and organic acids in environmental samples using co-electroosmotic capillary zone electrophoresis (co-CZE) at 185 nm. The best separation and detection of the solutes was achieved using a fused silica capillary with an electrolyte containing 25 mM phosphate, 0.5 mM tetradecyltrimethylammonium bromide (TTAB) and 15% acetonitrile (v/v) at pH 6.0. Four common inorganic anions (Cl⁻, NO₂ ⁻, NO₃ ⁻ and SO₄ ²⁻) and 11 organic acids (oxalic, formic, fumaric, tartaric, malonic, malic, citric, succinic, maleic, acetic, and lactic acid), were determined simultaneously in 15 min. Linear calibration plots for the test solutes were obtained in the range 0.02–0.5 mM with detection limits ranging from 1–9 μM depending on the analyte. The proposed method was successfully used to determine inorganic anions and carboxylic acids in soil and plant tissue extracts with direct injection of the sample.     

On-line preconcentration of niobium(V) and tantalum(V) as 4-(2-pyridylazo) resorcinol-citrate ternary complexes in geological samples by ion interaction high-performance liquid chromatography

4-(2-Pyridylazo) resorcinol (PAR) and citrate were used as pre-column complexing agents for the determination of Nb(V) and Ta(V) as ternary complexes in geological samples. Aliquots of 2 ml of the standard and sample solutions containing the Nb(V) and Ta(V) complexes were loaded onto a concentrator column (C18, 0.4 cm x 4.6 mm) with a carrier mobile phase comprising 20% (v/v) methanol and containing 5 mM acetic acid, 5 mM citric acid and 10 mM tetrabutylammonium bromide (TBABr), pH 6.5 at 2 ml/min for 2 min, with the effluent being directed to waste. An automatic switching valve was then switched to flush both complexes from the concentrator column onto a C18 analytical column using a mobile phase comprising 32% (v/v) methanol and containing 5 mM acetic acid, 5 mM citric acid and 3 mM TBABr, pH 6.5 for 2.5 min. The switching valve was then switched back to the original position, and cleaned with methanol for 7 min to eliminate unwanted species still adsorbed to the concentrator column. This procedure prevented later eluting compounds from reaching the analytical column, which reduced the overall run time. The detection limits of Nb(V) and Ta(V) (determined at a signal-to-noise ratio of 3, detection wavelength of 540 nm and a 2-ml sample volume) were 0.012 and 0.039 ppb for Nb(V) and Ta(V), respectively. Recoveries of Nb(V) and Ta(V) were 99.4 and 96.2%, respectively. The HPLC results obtained from the reference granite and basalt samples agreed well with inductively coupled plasma MS and certified values, but the HPLC method yielded slightly low values of the Nb/Ta ratio.     

Development of a capillary electrophoresis method for the determination of soybean proteins in soybean-rice gluten-free dietary products

CE has been applied for the first time to the simultaneous separation of soybean and rice proteins. Treated and untreated capillaries with different effective lengths as well as separation media at different pHs were tested. For that purpose, samples and standard solutions were prepared in 25:75 ACN-water media containing 0.3% v/v acetic acid. The use of an untreated capillary of 50 cm effective length together with an 80 mM borate buffer (pH 8.5) modified with 20% v/v ACN and UV detection at 254 nm were the conditions working the best. These conditions enabled the determination of soybean proteins in gluten-free dietary commercial products elaborated with soybean protein and/or soybean flour and rice flour using the standard additions calibration method. The method was linear up to 26 mg/mL of soybean proteins, the precision (expressed as RSD) was always better than 6%, and recoveries obtained for soybean proteins when spiking commercial products were very close to 100%.     

Determination of textile dyes by means of non-aqueous capillary electrophoresis with electrochemical detection

Eight textile dye compounds including five cationic dyes, namely, basic blue 41, basic blue 9, basic green 4, basic violet 16 and basic violet 3, and three anionic dyes, acid green 25, acid red 1 and acid blue 324, were separated and detected by non-aqueous capillary electrophoresis (NACE) with electrochemical detection. Simultaneous separations of acid and basic dyes were performed using an acetonitrile-based buffer. Particular attention was paid to the determination of basic textile dyes. The optimized electrophoresis buffer for the separation of basic dyes was a solvent mixture of acetonitrile/methanol (75:25, v/v) containing 1 M acetic acid and 10 mM sodium acetate. The limits of detection for the basic dyes were in the range of 0.1–0.7 μg mL⁻¹. An appropriate solid-phase extraction procedure was developed for the pre-treatment of aqueous samples with different matrices. This analytical approach was successfully applied to various water samples including river and lake water which were spiked with textile dyes.     

Electromembrane extraction of heavy metal cations followed by capillary electrophoresis with capacitively coupled contactless conductivity detection

Electromembrane extraction (EME) was used as an off-line sample pre-treatment method for the determination of heavy metal cations in aqueous samples using CE with capacitively coupled contactless conductivity detection (CE-C(4) D). A short segment of porous polypropylene hollow fibre was penetrated with 1-octanol and 0.5%?v/v bis(2-ethylhexyl)phosphonic acid and constituted a low cost, single use, disposable supported liquid membrane, which selectively transported and pre-concentrated heavy metal cations into the fibre lumen filled with 100?mM acetic acid acceptor solution. Donor solutions were standard solutions and real samples dissolved in deionized water at neutral pH. At optimized EME conditions (penetration time, 5?s; applied voltage, 75?V; and stirring rate, 750?rpm), 15-42% recoveries of heavy metal cations were achieved for a 5?min extraction time. Repeatability of the EME pre-treatment was examined for six independent EME runs and ranged from 6.6 to 11.1%. Limits of detection for the EME-CE-C(4) D method ranged from 25 to 200?nM, resulting into one to two orders of magnitude improvement compared with CE-C(4) D without sample treatment. The developed EME sample pre-treatment procedure was applied to the analysis of heavy metal cations in tap water and powdered milk samples. Zinc in the real samples was identified and quantified in a background electrolyte solution consisting of 20?mM L-histidine and 30?mM acetic acid at pH 4.95 in about 3?min.     

维生素C抗氧化-同位素内标稀释-超高效液相色谱-串联质谱同时测定新生期大鼠海马体神经递质的含量

脑区神经递质的测定对于研究神经系统的作用机制具有重要意义。该研究建立了同位素内标稀释-超高效液相色谱-串联质谱同时测定双酚A暴露后大鼠海马体中5种神经递质含量的方法，包括谷氨酸、γ-氨基丁酸、乙酰胆碱、多巴胺以及5-羟色胺。选用2%（体积分数）乙酸水-甲醇（9：1，v/v）溶液配制标准样品及溶解样品，以Ultimate AQ-C₁₈（150 mm×4.6 mm，3 μ m）为分离柱，0.1%（体积分数）甲酸水和甲醇为流动相，在28℃柱温下，梯度洗脱，所有化合物在3 min内出峰。添加维生素C能显著减少多巴胺和5-羟色胺在前处理以及样品储存过程中的氧化，极大地提高神经递质的稳定性，从而准确定量。该方法标准曲线线性关系良好，相关系数R²>0.998，检出限和定量限低，日内和日间精密度为0.39%~13.6%，加标回收率为92.9%~119%，实际进样残留低，已被成功地应用于新生期双酚A暴露后大鼠海马体神经递质含量的测定。     

Simple and sensitive HPLC method for determination of amrubicin and amrubicinol in human plasma: application to a clinical pharmacokinetic study

A simple and sensitive high‐performance liquid chromatographic (HPLC) method was developed for determination of amrubicin and its metabolite amrubicinol in human plasma. After protein precipitation with methanol without evaporation procedure, large volume samples were injected and separated by two monolithic columns with a guard column. The mobile phase consisted of tetrahydrofuran–dioxane–water (containing 2.3 mM acetic acid and 4 mM sodium 1‐octanesulfonate; 2:6:15, v/v/v). Wavelengths of fluorescence detection were set at 480 nm for excitation and 550 nm for detection. Under these conditions, linearity was confirmed in the 2.5–5000 ng/mL concentration range of both compounds. The intra‐ and inter‐day precision and intra‐ and inter‐day accuracy for both compounds were less than 10%. The method was successfully applied to a clinical pharmacokinetic study of amrubicin and amrubicinol in cancer patients. Copyright © 2009 John Wiley & Sons, Ltd.     

Determination of water-soluble UV-filters in sunscreen sprays by liquid chromatography

Liquid chromatography was used for the determination of the three most used water-soluble UV filters, benzophenone-4 (BZ4), terephthalylidene dicamphor sulfonic acid (TDS), and phenylbenzimidazole sulphonic acid (PBS), in aqueous sunscreen sprays. A C18 stationary phase and an isocratic mobile phase of EtOH-20 mM sodium acetate buffer of pH 4.6 (30:70, v/v) were used at a flow-rate of 0.5 ml min(-1). Mobile phase was also used as solvent for samples and standards. UV detection was at 313 nm. The analytical run took 5.5 min. The limits of detection were 0.5, 0.9 and 2 microg ml(-1) for BZ4, TDS and PBS, respectively. The proposed method does not involve highly toxicsolvents.     

Capillary electrophoresis analysis of fosfomycin in biological fluids for clinical pharmacokinetic studies

A feasible capillary zone electrophoresis (CZE) method with indirect UV and contactless conductivity detection was developed for the determination of fosfomycin, an antibiotic, in human plasma and microdialysis samples. Samples were collected from test persons during a clinical trial. The background electrolytes used consisted of 25 mM benzoic acid and 0.5 mM hexadecyltrimethylammonium bromide, adjusted with tris(hydroxymethyl)aminomethane solution to pH 6.95 for plasma, and to pH 8.05 for microdialysis samples. CZE separations of the anionic analyte were carried out with reversed electroosmotic flow directed towards the anode. The limit of detection was between 0.6 and 2 microg/mL, depending on the matrix and the detection method. No sample preparation was needed for microdialysis samples; for plasma samples, proteins were precipitated with methanol (1+2, v+v), and the supernatant was analyzed. The yield determined with spiked samples was about 100%, the reproducibility of the entire method, expressed by the RSD% of three independent determinations of fosfomycin in triplicate after spiking Ringer's solutions and plasma samples, respectively, was better than 8%. The method is thus well-suited for clinical studies for the determination of the antibiotic in biological fluids.     

Simultaneous HPLC-UV determination of ketamine, xylazine, and midazolam in canine plasma

An isocratic reversed-phase high-performance liquid chromatography method with UV detection is developed and validated for the simultaneous determination of ketamine, xylazine, and midazolam in canine plasma. Analytes are extracted from alkalinized samples into diethyl ether-methylene chloride (7:3, v:v) using single-step liquid-liquid extraction. Chromatographic separation is performed on a C(18) column using a mobile phase containing an acetonitrile-methanol-10 mM sodium heptanesulfonate buffer adjusted to pH 3, with glacial acetic acid (44:10:46, v:v) at a detection wavelength of 210 nm, with a total runtime of 10 min. The calibration is linear over the range of 78.125-5000 ng/mL for ketamine and 15.625-1000 ng/mL for xylazine and midazolam. The limits of detection are 17.8, 10.3, and 15.1 ng/mL for ketamine, xylazine, and midazolam, respectively. The extraction recoveries are 76.1% for ketamine, 91.0% for midazolam, and 78.2% for xylazine. The method is successfully used for clinical and pharmacokinetic studies of the three-drug fixed dose combination formulations.     

Determination of acidic herbicides in surface water by solid-phase extraction followed by capillary zone electrophoresis

A rapid solid-phase extraction-capillary zone electrophoresis (CZE) method for determining 2,4-dichlorophenoxyacetic acid, 4-(2,4-dichlorophenoxy) butyric acid, and 2,4,5-trichlorophenoxyacetic acid in real water samples is described. Factors affecting the recoveries and detection of the targets are investigated. With samples being acidified to pH 2 and salted by sodium sulfate to 2% (w/w), an average recovery of greater than 85% is obtained using ethyl acetate as the eluent on an octadecylsilane-bonded silica cartridge. A running buffer of 5 mM sodium tetraborate in a water-acetonitrile mixture (70:30, v/v) adjusted to pH 9 is employed in the CZE analysis, and the targets can be analyzed within 7 min with good reproducibility and acceptable sensitivity. The method is suitable for detecting herbicide residues of sub-parts-per-billion levels in surface water. A local pond water is analyzed, and the concentrations of 2,4-dichlorophenoxyacetic acid and 4-(2,4-dichlorophenoxy) butyric acid are detected to be 0.27 +/- 0.03 ppb and 0.61 +/- 0.08 ppb, respectively.     

Photochemical-fluorimetric method for the determination of total chlorophenoxyacid herbicides

A room temperature photochemically-induced fluorescence (RTPF) method is proposed for the quantitative analysis of seven widely-used chlorophenoxyacid herbicides. The influence of organic solvent, pH (in aqueous solutions), methanol percentage, and UV irradiation time on the excitation and emission wavelengths and fluorescence intensity was investigated. It was found that the largest fluorescence signals were obtained in a mixture of methanol and pH 5 buffer (50/50, v/v), while organic solvents and water produced generally lower signals. The tri- and bichlorinated phenoxyacid herbicides were photolysed significantly more slowly than the monochlorinated derivatives, and the derivatives of 2-propionic acid were photodegraded more quickly than the corresponding derivatives of acetic and butyric acid. Selected UV irradiation times were found to be 15 min for all herbicides under study. Linear calibration graphs were established over about one to two orders of magnitude in the interval 0.1-10 mug ml(-1). The RTPF limits of detection were between 36 ng ml(-1) and 179 ng ml(-1), according to the compound. Analytical application of RTPF to river water samples containing chlorophenoxyacid herbicides is discussed.     

Determination of valproic acid (2-propylpentanoic acid) in human plasma by capillary electrophoresis with indirect UV detection

A rapid capillary zone electrophoresis method with indirect UV detection was developed and validated for the determination of valproic acid (VPA) in human plasma. The analyses were carried out under optimized conditions, using a buffer system composed of 15 mM benzoate and 0.5 mM cetyltrimethylammonium bromide at pH 6.0, and 25% v/v methanol; 2-hydroxybutyric acid was selected as the internal standard (IS). The capillary electrophoresis (CE) separation was carried out at a negative potential of 30 kV and the indirect UV detection was operated at 210 +/- 20 nm for all assays. The influence of buffer pH, ionic strength, concentration of electroosmotic flow (EOF) modifier and organic modifier on indirect signal response and migration behavior of the organic acid was investigated. Isolation of VPA from plasma was accomplished by a carefully implemented procedure using methanol as the precipitant agent. Using a high ratio of methanol to plasma for deproteinization (4:1), good absolute recovery of the analyte and satisfactory selectivity was obtained. The calibration line for VPA was linear over the 1-100 microg/mL concentration range. Sensitivity was high; in fact, the limit of detection (LOD) of VPA was 150 ng/mL and 450 ng/mL the limit of quantitation (LOQ). The results obtained analyzing real plasma samples from schizophrenic patients under polytherapy with VPA as well as antipsychotic drugs were satisfactory in terms of precision, accuracy and sensitivity.     

Validated and optimized high-performance liquid chromatographic determination of tizoxanide, the main active metabolite of nitazoxanide in human urine, plasma and breast milk

A high-performance liquid chromatographic method was optimized and validated for the determination of desacetyl nitazoxanide (tizoxanide), the main active metabolite of nitazoxanide in human plasma, urine and breast milk. The proposed method used a CN column with mobile phase consisting of acetonitrile-12mM ammonium acetate-diethylamine in the ratio of 30:70:0.1 (v/v/v) and buffered at pH 4.0 with acetic acid, with a flow rate of 1.5 mL/min. Quantitation was achieved with UV detection at 260 nm using nifuroxazide as internal standard. A simplified direct injection of urine samples without extraction in addition to the urinary excretion pattern were calculated using the proposed method. Also, the effectiveness of protein precipitation and a clean-up procedure were investigated for biological plasma and human breast milk samples. The validation study of the proposed method was successfully carried out in an assay range between 0.2 and 20 μg/mL.     

Simultaneous determination of ropivacaine and antipyrine by high performance liquid chromatography and its application to the in vitro transplacental study

A simple and reliable RP-HPLC method has been developed for the simultaneous determination of ropivacaine and antipyrine in perfusate samples. Samples were analyzed on an ODS column with UV detection at 210 nm after liquid-liquid extraction. The mobile phase consisted of potassium dihydrogenphosphate (25 mM, adjusted to pH 5.0 with phosphoric acid)-acetonitrile (79:21, v/v). The method has been validated to be precise, accurate and linear. It has been applied to the investigation of placental transfer of ropivacaine via a dually perfused cotyledon model of human placenta in vitro.     

Analytical potential of 6-oxy-(N-succinimidyl acetate)-9-(2'-methoxycarbonyl) fluorescein for the determination of amino compounds by capillary electrophoresis with laser-induced fluorescence detection

The analytical potential of a fluorescein analogue, 6-oxy-(N-succinimidyl acetate)-9-(2'-methoxycarbonyl) fluorescein (SAMF), for the first time synthesized in our laboratory, as a labeling reagent for the labeling and determination of amino compounds by capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection was investigated. Biogenic monoamines and amino acids were chosen as model analytes to evaluate the analytical possibilities of this approach. The derivatization conditions and separation parameters for the biogenic amines were optimized in detail. The derivatization was performed at 30 degrees C for 6 min in boric acid buffer (pH 8.0). The derivatives were baseline-separated in 15 min with 25 mM boric acid running buffer (pH 9.0), containing 24 mM SDS and 12.5% v/v acetonitrile. The concentration detection limit for biogenic amines reaches 8 x 10(-11) mol.L(-1) (signal-to-noise ratio = 3). The application of CE in the analysis of the SAMF-derivatized amino acids was also exploited. The optimal running buffer for amino acids suggested that weak acidic background electrolyte offered better separation than the basic one. The proposed method was applied to the determination of biogenic amines in three different beer samples with satisfying recoveries varying from 92.8% to 104.8%. Finally, comparison of several fluorescein-based probes for amino compounds was discussed. With good labeling reaction, excellent photostability, pH-independent fluorescence (pH 4-9), and the resultant widely suited running buffer pH, SAMF has a great prospect in the determination of amino compounds in CE.     

Analysis of formic and acetic acid in rain water by capillary electrophoresis

A simple technique is described for the routine capillary electrophoretic determination of formic and acetic acid in rain water. These acids were determined simultaneously in approximately 6 min using a carrier electrolyte containing lO mM phosphate and 0.5 mM myristyltrimethylammonium bromide (MTAB) as electroosmotic flow (EOF) modifier at pH 6.5 and direct UV detection at 185nm. The method is quantitative, with recoveries in the 99-101% range and linear up to 5mgL^-1. The precision is better than 2.1% and the procedure shows the appropriate sensitivity, with detection limits between 0.042 and 0.055mg L^-1. The proposed method was successfully employed for the determination of formic and acetic acid in 57 rain water samples, collected from October 2000 to February 2001 in four different sampling stations located in Galicia (NW Spain), by direct sample injection after filtration.     

Determination of caffeine and associated compounds in food, beverages, natural products, pharmaceuticals, and cosmetics by micellar electrokinetic capillary chromatography

A method is described for quantitating caffeine, theobromine, theophylline, paracetamol, propyphenazone, acetylsalicylic acid, salicylic acid, and codeine phosphate in corresponding real samples of food, beverages, natural products, pharmaceuticals, and cosmetic preparations by micellar electrokinetic capillary chromatography. The separation is carried out at 25 degrees C and 25 kV, using a 20mM phosphate buffer (pH 9.0), 80mM sodium dodecyl sulfate, and 7.5% (v/v) acetonitrile. UV detection is at 210 nm. The method is shown to be specific, accurate (recoveries over the range 98.9-101.2%), linear over the tested range (correlation coefficients >/= 0.9993), and precise (relative standard deviation below 2.1%). The method is applied for the quantitative analysis of these compounds in different foods, beverages, natural products, pharmaceuticals, and cosmetic products.     

Simultaneous Separation and Determination of Benzoic Acid Compounds in the Plant Medicine by High Performance Capillary Electrophoresis

A simple and inexpensive high performance capillary electrophoresis (HPCE) was applied to separate five benzoic acid compounds simultaneously. The investigation was carried out by micellar electrokinetic capillary chromatography (MECC). To avoid a time‐consuming and tedious procedure, orthogonal experimental design OA₉ (3⁴) for separation experiments was applied to find the optimal conditions in terms of the resolution and analytical time. The best conditions for separation were obtained using a 20 mM borax and 30 mM sodium dodecyl sulfate (SDS) buffer (pH 9.8) containing 2 mM β‐CD and 4% methanol (v/v). Online UV detection was performed at 250 nm. A voltage of 16 kV was applied and the temperature was controlled at 25 °C. Injection was performed for 5 s. The method was validated for the quantification of benzoic acid, salicylic acid and ortho‐aminobenzoic acid in Radix Isatidis, a traditional plant medicine with removal of endotoxin. The separation and determination were satisfactory and quick.     

Determination of egg white lysozyme by on-line coupled capillary isotachophoresis with capillary zone electrophoresis

An on-line coupled capillary isotachophoresis - capillary zone electrophoresis method for the determination of lysozyme in selected food products is described. The optimized electrolyte system consisted of 10 mM NH(4)OH + 20 mM acetic acid (leading electrolyte), 5 mM epsilon -aminocaproic acid +5 mM acetic acid (terminating electrolyte), and 20 mM epsilon -aminocaproic acid +5 mM acetic acid +0.1% m/v hydroxypropylmethylcellulose (background electrolyte). A clear separation of lysozyme from other components of acidic sample extract was achieved within 15 min. Method characteristics, i.e., linearity (0-50 micrograms/mL), accuracy (recovery 96+/-5%), intra-assay (3.8%), quantification limit (1 microgram/ml), and detection limit (0.25 microgram/mL) were determined. Low laboriousness, sufficient sensitivity and low running costs are important attributes of this method. The developed method is suitable for the quantification of the egg content in egg pasta.