Selectivity control in the non-aqueous capillary electrophoretic separation of amino acids

The separation of dansylated amino acids and underivatized amino acids in non-aqueous electrolytes was evaluated with direct and indirect UV detection. Different migration orders were achieved for dansylated amino acids in methanol compared to aqueous electrolyte systems. A reversed migration order was observed for some dansylated amino acids. Separation selectivity was different under acidic and basic conditions and was also a function of the solvation properties of the solvent. Underivatized amino acids were separated in basic and acidic electrolytes in methanol; different separation selectivities and, for some amino acids, a reversed migration order were also observed in these electrolyte systems. Analytical merits of the separation of both derivatized and underivatized amino acids were briefly evaluated; detection limits for dansylated amino acids were in the range of 2·10⁻⁷–4·10⁻⁷ mol/l and, for underivatized amino acids, were 2·10⁻⁶–4·10⁻⁵ mol/l.     

Direct sample injection for capillary electrophoretic determination of organic acids in cerebrospinal fluid

Organic acids in cerebrospinal fluid (CSF) are potential diagnostic markers for neurological diseases and metabolic disorders. A capillary electrophoretic (CE) method for the direct analysis, i.e., without any sample preparation, of six organic acids in CSF was developed. A capillary coating consisting of a triple layer of charged polymers (polybrene-dextran sulfate-polybrene) was used in combination with a negative separation voltage, providing fast and efficient analysis of acidic compounds. Separation conditions, such as background electrolyte (BGE) concentration and pH were optimized, and the influence of albumin and sodium chloride was systematically studied using a set of test compounds. With injection volumes of ca. 44 nL, plate numbers of up to ca. 150,000 were obtained with a BGE of 200 mM sodium phosphate (pH 6.0). It appeared that high sodium chloride concentrations in the sample hardly affected the peak width and shape of the organic acids, most probably due to transient isotachophoresis effects occurring in the sample zone. Adverse effects of CSF proteins, which frequently compromise the CE performance, could be effectively minimized by the triple layer coating in combination with rinses of 0.1 M hydrochloric acid. Overall, the developed CE system allowed direct injections of CSF samples, yielding good separation efficiencies and stable migration times (RSDs < 2%) for organic acids. Validation of the method with artificial and real CSF samples showed good linear responses (r > 0.99), and LODs for the organic acids were in the range of 2–8 μg/mL when applying UV detection. RSDs for migration times and peak areas were <2% and <7%, respectively. The applicability of the CE system is shown for the determination of organic acids in CSF samples.     

Low-molecular-mass anion screening for forensic environmental analyses by capillary zone electrophoresis with indirect UV detection

A method for low-molecular-mass anion screening is described using a buffer composed of 5-sulfosalicylate (SS) as a visualizing ion, hexadimethrine bromide as an electroosmotic flow modifier and Tris as a pH buffer component, at pH 8.6. All ions with effective mobility higher than 2610⁻⁹ m² s⁻¹ V⁻¹ can be separated within 7.5 min under −30 kV. By using the moderately mobile SS (5410⁻⁹ m² s⁻¹ V⁻¹), not only the sensitivity of the detection is improved due to its high UV absorptivity, but also a smaller overall overloading effect is achieved. Meanwhile, the resolution of the high mobility ions, which is normally critical, remains almost the same as compared to a chromate buffer. With an electrokinetic injection, the limit of detection (LOD) of the common ions is 2–13 nM and the detection range is linear up to 0.5–3 μM. With a hydrostatic injection the LOD is 0.15–1 μM and the detection range is linear up to 25–200 μM. The identification of ions is performed by comparing the mobility of the ions with that of standards, taking the apparent and effective mobility of HCO₃⁻, which is normally present in the sample solution, as a reference.     

Sample pretreatment for the capillary electrophoretic determination of organic acids in chromium(III) plating baths

This work deals with the development and optimization of the sample pretreatment and consequent electrophoretic analysis of two modern plating baths containing chromium(III) and either citric acid or oxalic acid. Some model mixtures containing known amounts of components of industrial baths have been prepared to simulate simplified bath matrices. Prior to analysis, a sample pretreatment consisting of the addition of some agents that could release acid from the stable chromium complex was tested. Determination of organic anions was accomplished by indirect UV detection. The best results were achieved by precipitation of chromium(III) hydroxide. The content of oxalate and citrate in real samples was calculated as 96.5% (SD 2.3%) and 97.3% (SD 0.8%), respectively, of the declared amount. Very good robustness of the method and satisfactory repeatability of migration time and peak area were obtained. This simple inexpensive method is suitable for routine determination of citric and oxalic acid in chromium(III)‐based plating baths.     

毛细管电泳法对铝胁迫下小麦根部有机酸的直接测定

在进行植物抗铝逆性研究中,从植物的培养液中发现从根部分泌出的有机酸与铝结合从而实现无毒化是其抗铝逆性机理的重要依据,但是其分析需繁杂的前处理过程。而本文用毛细管电泳法直接测定了铝胁迫下培育的小麦根中的铝和有机酸,确认了主要积累的是苹果酸和柠檬酸。发现随着提高培养液中的铝浓度,根部的铝量也相应增加。同时,虽然根中的柠檬酸量无明显变化,但苹果酸被诱导增加。在pH7．8的Na3PO4缓冲溶液中加入表面活性剂溴化十六烷基三甲铵作为电泳溶液体系,紫外检测波长214nm,可以有效地分离检测以阴离子形式存在的上述有机酸。探讨了植物抗铝胁迫研究中简便易行的植物根部有机酸的直接测定方法。     

Simultaneous determination of inorganic anions and organic acids in amine solutions for sour gas treatment by capillary electrophoresis with indirect UV detection

A new CE method has been developed for the simultaneous determination of selected inorganic anions (bromide, chloride, thiosulfate, nitrite, nitrate, sulfate, thiocyanate, fluoride and phosphate) and organic acids (oxalic, malonic, formic, tartric, acetic, glycolic, propionic, butyric and cyclohexanoic) in amine solutions from sour gas treatment units. An electrolyte composed of 10 mM trimellitic acid, 200 mM Tris (pH 9.0), 0.1% polyvinyl alcohol provides a satisfactory separation of all analytes of interest. The electroosmotic flow is reversed by using hexadimethrine bromide as a semi-permanent positively charged coating, making the electrolyte free of additive. Indirect UV detection at 240 nm is used because of the weak absorbing properties of most of analytes. The addition of 1% diethanolamine in standard mixtures permits to better preserve them, inhibiting potential degradation processes, especially for thiosulfate. The quantification is performed using internal standardization, by which molybdate is used as internal standard. Moreover, the use of relative migration times and the excellent repeatabilities obtained allow unambiguous identification of analytes in real samples by comparison with standard mixture. It has been shown that no significant matrix effect came from the presence of 30% diethanolamine in amine solution samples and the developed method was characterized in terms of calibration linearity and accuracy using recovery tests. In short, the developed method allows the simultaneous and rapid determination, in difficult matrices, of numerous inorganic anions and organic acids characterized by a large range of electrophoretic mobilities.     

Using capillary electrophoresis for the determination of organic acids in Port wine

The simultaneous separation and determination of organic acids in several samples of white and red Port wines was performed by capillary zone electrophoresis using indirect UV detection with 2,6-pyridinedicarboxylic acid as a background electrolyte buffer. Operational parameters like migration time, temperature, voltage and capillary length were optimized. Sixteen samples of red wine and four samples of white wine were used to analyze for tartaric, malic, lactic, succinic and acetic acids using glyoxylic acid as the internal standard. The method is rapid, sensitive and quantitative, and time-consuming sample preparation, such as solid-phase extraction or liquid-liquid extraction procedure, is not required.     

A facile and sensitive chemiluminescence detection of amino acids in biological samples after capillary electrophoretic separation

It was found that native amino acids enhanced the chemiluminescence (CL) reaction between luminol and BrO(-) in an alkaline aqueous solution. This has led to the development of a facile and highly sensitive CL detection scheme for the determination of amino acids in biological samples after capillary electrophoretic (CE) separation. The CE-CL conditions were optimized. An electrophoretic buffer of 2.5 x 10(-2) M sodium borate (pH 9.4) containing 1 x 10(-4) M luminol was used. The oxidizer solution of 8 x 10(-4) M NaBrO in 0.1 M sodium carbonate buffer solution (pH 12.5) was introduced post-column. Under the optimal conditions, the detection limits were 1.0 x 10(-7) M for glutamic acid (Glu) and 1.3 x 10(-7) M (S/N = 3) for aspartic acid (Asp). The relative standard deviations (RSDs) of peak area and migration time were in the ranges of 3.8-4.3% and 1.4-1.6%, respectively. The present method was applied to the determination of excitatory amino acids (i.e., Asp and Glu) in rat brain tissue and monkey plasma. The levels of these major excitatory amino acids in monkey plasma were quantified for the first time and found to be 1.17 +/- 0.17 x 10(-5) M (mean +/- SD, n = 6) for Glu and 1.64 +/- 0.19 x 10(-6) M for Asp, which were comparable with the levels in human plasma.     

Preconcentration and separation of haloacetic acids by ion chromatography

A comparative study was made of the chromatographic behaviour of five haloacetic acids (mono-, dibromoacetic and mono-, di-, trichloroacetic acids). The techniques investigated included reversed-phase ion interaction chromatography with UV detection, suppressed and non-suppressed anion-exchange chromatography. The systems are discussed studying the retention as a function of the mobile phase parameters and the stationary phases used (LiChrospher 100 RP-18, IonPac AS9, AS10 and AS11). A preconcentration step, performed on different substrates, namely LiChrolut-EN and activated vegetal carbon, has been optimized in order to reduce the method detection limits. Results obtained for drinking water samples are shown.     

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.     

Determination of organic acids in air by capillary electrophoresis and ion-exclusion chromatography

Summary A capillary electrophoretic (CE) method for the determination of organic acids in the low ppm range is described. The buffer consisted of 5 mM 2,6-pyridinedicarboxylic acid and 0.5 mM cetyltrimethylammonium bromide, pH 5.6. The former served as background electrolyte for indirect UV detection at 200 nm, whereas the latter was used to reverse electroosmotic flow. In <5 min 8 low molecular mass organic acids (oxalic, formic, malonic, glutaric, glycolic, acetic, lactic and propanoic) and two inorganic acids (hydrochloric and sulphuric) were separated. Detection limits for anions tested were 0.04 mg L⁻¹ (lactic acid) to 0.6 mg L⁻¹ (malonic acid). The method was applied to the determination of organic acids in air samples. The CE results when compared with ion-exclusion chromatography (IEC) agreed well. The use of electrokinetic injection in CE proved beneficial for preconcentration of organic acids in real samples. Using electrokinetic injection, preconcentration factors ranging from 14 (hydrochloric acid) to 3 (propanoic acid) were obtained. Presented at Balaton Symposium on High-Performance Separation Methods, Siófok, Hungary, September 1–3, 1999     

Capillary electrophoretic separation of dicarboxylic acids in atmospheric aerosol particles

2,3-Pyrazinedicarboxylic acid (PZDA), 2,6-pyridinedicarboxylic acid (PDA) and 2,3-pyridinedicarboxylic acid (QUIN) solutions were studied as background electrolytes (BGEs) in the capillary electrophoretic analysis of dicarboxylic acids in aerosol particles with indirect UV detection. The BGEs were selected on the basis of similarity in structure with the analytes so that mobilities would be compatible. Optimised pH values for PZDA, PDA and QUIN solutions were 10.6, 11.0 and 10.2, respectively. Myristyltrimethylammonium hydroxide and myristyltrimethylammonium bromide were added to reverse the electroosmotic flow in the solutions in the direction of anode to enable fast anion detection. Separation was obtained for nine dicarboxylic acids (C₂–C₁₀) differing in the number of CH₂ groups in their skeleton. The electrophoretic mobilities were determined to lie in the range 3.0×10⁻⁴–7.0×10⁻⁴ cm² V⁻¹ s⁻¹. The relative standard deviations (RSDs) for the absolute migration times of the analytes were mostly less than 0.5% (n=6) in PZDA solution. In PDA solution the within-day and day-to-day RSD values for migration were less than 1% and between 2 and 4%, respectively. Peak heights and areas mostly deviated between 1 and 15% in both PZDA and PDA solutions. Detection limits ranged between 1 and 5 mg/l. Methods were applied to the analysis of dicarboxylic acids isolated from aerosol particles.     

Determination of low-molecular-mass organic acids in any type of beer samples by coelectroosmotic capillary electrophoresis

A separation and determination of a mixture of 19 low-molecular-mass organic acids usually present in beer samples was developed using coelectroosmotic capillary zone electrophoresis. A polycation (hexadimetrine bromide, HDB) has been added to the electrolyte, which dynamically coats the inner surface of the capillary and causes a fast anodic electroosmotic flow. The main factors affecting reversal of the EOF such as type of modifier and concentration and influence of organic solvents were studied. Three types of modifiers, two alkylammonium salts (cethyltrimethylammonium bromide and tetradecyltrimethylammonium bromide) and a polycation (HDB) were investigated. The composition of the running buffer results on a 25% 2-propanol, 0.001% HDB and 50 mM sodium phosphate. The different instrumental parameters affecting the capillary electrophoretic separation were also optimized resulting on a -15 kV voltage with a hydrodynamic injection for 7 s with a UV detection at 210 nm. The applicability of the present method has been demonstrated for the determination of organic acids in different beer samples.     

Separation of phenolic acids by capillary electrophoresis with indirect contactless conductometric detection

A new method for the electrophoretic separation of nine phenolic acids (derivatives of benzoic and cinnamic acids) with contactless conductometric detection is presented. Based on theoretical calculations, in which the mobility of the electrolyte co- and counterions and mobility of analytes are taken into consideration, the electrolyte composition and detection mode was selected. This approach was found to be especially valuable for optimization of the electrolyte composition for the separation of analytes having medium mobility. Indirect conductometric detection mode was superior to the direct mode as predicted theoretically. The best performance was achieved with 150 mM 2-amino-2-methylpropanol electrolyte at pH 11.6. The separation was carried out in a counter-electroosmotic mode and completed in less than 6 min. The LODs achieved were about 2.3-3.3 microM and could be further improved to 0.12-0.17 microM by using a sample stacking procedure. The method compares well to the UV-Vis detection.     

Glass capillary columns for separation of free organic acids

Glass capillary columns have been prepared without acidic additive in the stationary phase, from which free organic acids elute as sharp and symmetrical peaks. The required surface in the borosilicate glass capillary was generated by a combination of leaching with aqueous HCl and deposition of colloidal silica particles; it can be coated with stationary phases have a broad range of polarity. Aqueous samples containing free organic acids can also be analyzed in such columns in an isothermal mode.     

Podand and macrocyclic amine receptors with urea functionalities for potentiometric detection of organic acids in HPLC

Ten synthetic receptors were tested in potentiometric coated wire electrodes. The electrodes were used as sensing devices in classical reversed phase HPLC determinations of dicarboxylic acids present in food materials. All receptors contained amine functionalities. Four of them were podand urea derivatives. The other six were macrocyclic polyamines with lipophilic groups. Three of them had urea functionalities in their side-chains. Glassy carbon (GC) substrate electrodes were coated with PVC-based “liquid membranes” containing plasticizers and receptors. All tested compounds strongly enhanced the detection limits for the dicarboxylic acids (low pg detection limits), in comparison to a non-specific electrode based on methyltridodecylammonium chloride (MTDDACl). Receptors of the podand urea type yielded high sensitivity, but the electrodes had a lower long-time stability (a few weeks) than the electrodes based on macrocyclic polyamines (at least 3 months).     

毛细管电泳-间接紫外检测法测定蜂蜜中的氨基酸

采用毛细管电泳-间接紫外检测法同时分离测定蜂蜜中的赖氨酸、色氨酸、谷氨酸等9种氨基酸。考察了磷酸浓度、进样方式和缓冲液pH对分离效率和重现性的影响。在分离电压为-15 kV、检测波长为220 nm条件下,以含有0.5 mmol/L十六烷基三甲基溴化铵、20 mmol/L烟酸、10%甲醇的10 mmol/L磷酸二氢钠缓冲溶液(pH 10.2)为运行缓冲液,9种组分在11 min内达到基线分离;检出限最低可达到0.3 mg/L;线性范围为1.0~1000 mg/L;日间及日内精密度为0.64%~5.83%。实际样品中除甲硫氨酸外的8种氨基酸的加标回收率为60.00%~118.37%。将该方法应用于不同蜜源植物和产地的蜂蜜样品的测定,在市售的5种蜂蜜中均检测到脯氨酸、丝氨酸和天冬氨酸,而只在荔枝蜜中检测到苏氨酸。该方法可以为蜂蜜的蜜源鉴别及质量评估提供借鉴方法。     

Determination of various inorganic anions and organic acids in the atmospheric particulate matter using capillary zone electrophoresis with indirect UV detection

Capillary zone electrophoresis with indirect UV detection was developed for the simultaneous analysis of inorganic anions and organic acids using a mixed solution of 2,6-pyridinedicarboxylic acid and cetyltrimethylammonium hydroxide as the background electrolyte (BGE). The parameters which influence the separation, such as indirect UV detection wavelength, BGE conditions, applied voltage and extraction conditions were investigated. Thirteen inorganic anions and organic acids were detected within 20 min. The calibration curves of each analyte were linear with correlation coefficients greater than 0.991. The relative standard deviations (n = 10) of the peak areas ranged from 0.6% to 3.3%. The detection limits for these species ranged from 0.4 to 1.4 mg/L at a signal-to-noise ratio of 3. The recovery rate of each analyte was more than 80% under optimised extraction conditions, except for nitrite. The proposed method was applied towards the analysis of inorganic anions and organic acids in the atmospheric particulate matter using an Andersen sampler. The particle size of the particulate matter was determined, but not the size of the anions.     

Potentiometric detection of organic acids in liquid chromatography using conducting oligomer electrodes

A conducting oligomer electrode was used for the potentiometric detection of organic acids in reversed phase liquid chromatography (LC). The conducting material consisted of a mixture of a phenylene vinylene trimer with a polycarbonate host polymer and iodine. A glassy carbon electrode was coated with this material by evaporation from a chloroform solution. A theoretical model was given to describe the observed potentiometric responses. The analysis conditions were optimized to obtain both efficient separations, and sensitive potentiometric responses. Detection limits in the nanogram level were attained when a 1 mM phosphoric acid solution was used as the eluent, which were comparable to the values obtained with low-wavelength UV detection. Calibration curves showed a logarithmic dependence on an injected amount for amounts higher than 5 nmol, and a linear dependence for injected amounts below this value. The response times of the electrode were smaller than 1 s at typical LC flow-rates. The reproducibility for consecutive injections was 5%.     

Development and optimization of organic acid analysis in tobacco with ion chromatography and suppressed conductivity detection

With the aid of a central composite face-centered design, an ion chromatographic method was developed and optimized for analyzing organic acids in tobacco. A Dionex-100 ion chromatograph with an ion suppressor and a conductivity detector, and a Bio-Rad Aminex HPX-87H column were employed. Only 13 analyses were required to optimize two factors: column temperature and eluent strength. Two sets of optimal conditions for separating nine acids were found: 1.8 mM HFBA eluent and 42 °C column temperature, and 0.8 mM HFBA eluent and 50 °C column temperature. The flow-rate was 0.6 ml min⁻¹ and the analysis time was 18 min or less. A sample preparation procedure included extraction of 2 g ground tobacco with 100 ml of 5 mM sulfuric acid solution for 3 h, filtration of the extract, and dilution of the filtrate 10-fold with deionized water.