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Keqi Tang - One of the best experts on this subject based on the ideXlab platform.
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on a separation voltage polarity switching transient Capillary Isotachophoresis method for higher sample loading capacity and better separation performance
Analyst, 2021Co-Authors: Ji Tang, Junliang Zhang, Chenlu Wang, Tao Yin, Keqi TangAbstract:Limited sample loading capacity is one of the major reasons that prevents the utility of Capillary electrophoresis (CE) as a routine separation method as compared to liquid chromatography (LC). In our previous study, separation voltage polarity switching transient Capillary Isotachophoresis (PS-tCITP) was proposed. Both sample loading capacity and separation resolution could be improved using a single PS-tCITP instead of routine transient Capillary Isotachophoresis (tCITP). In this study, a detailed investigation on the optimization strategy of the PS-tCITP method was performed systematically. A possible mechanism of sample preconcentration in multiple PS-tCITP was first proposed to better understand the multiple PS-tCITP process. Several optimization experiments were then performed, including single PS-tCITP, paused PS-tCITP and multiple PS-tCITP, sequentially using a mixture of five peptides. By selecting an optimum polarity switching time, sample loading capacity of 100% Capillary volume could be achieved in a single PS-tCITP. Introducing an additional pause between each polarity switching in a single PS-tCITP further improved the separation resolution. Experimental results showed a baseline separation of five selected peptide standards at 100% sample loading volume using a 100 min pause in a single PS-tCITP. To further improve separation efficiency while still maintaining 100% sample loading volume, a multiple PS-tCITP technique was developed through this study. Compared to the separation performance of the optimal single PS-tCITP at 100% sample loading volume with a 10 min pause, the separation window was improved by 54% and the peak capacity was improved by 48% in the optimal four PS-tCITP with the same sample loading volume and pause.
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a separation voltage polarity switching method for higher sample loading capacity and better separation resolution in transient Capillary Isotachophoresis separation
Analyst, 2019Co-Authors: Roza Wojcik, Tujin Shi, Keqi TangAbstract:A separation voltage polarity switching transient Capillary Isotachophoresis (PS-tCITP) was developed to overcome a major sample loading volume limitation in transient Capillary Isotachophoresis (tCITP). The fundamental idea of PS-tCITP is to let sample ions move back and forth in a separation Capillary during their initial Isotachophoresis focusing stage by switching the polarity of the separation voltage, in order to both increase the sample loading volume and improve the separation efficiency as compared to the conventional tCITP method. The experimental evaluation of the novel PS-tCITP method by using two peptide standards at 2 μM concentration showed that the maximum sample loading volume could be increased from 45% of the total separation Capillary volume in tCITP to 70% in PS-tCITP, which resulted in a more than 1.5 fold increase in the peptide peak intensity at a given length/volume of the separation Capillary. Due to the consecutive focusing of sample volume from each polarity switching of the separation voltage, the separation time window at a given sample loading volume was also increased significantly in PS-tCITP as compared to tCITP. Experiment comparison between tCITP and PS-tCITP at 45% sample loading volume using the same setup showed that the migration time difference between the two peptide peaks increased from 0.3 min in tCITP to 0.363 min in PS-tCITP with similar peak widths and heights, resulting in roughly a 21% improvement in separation resolution. The performance advantages of PS-tCITP separation over tCITP separation were further verified by using a mixture of six peptide standards.
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Effective coupling of CE with nanoESI MS via a true sheathless metal-coated emitter interface for robust and high sensitivity sample quantification (ASMS 2016)
2017Co-Authors: Pnnl Omics, Keqi Tang, Xuejiang Guo, Thomas L. Fillmore, Yuqian GaoAbstract:Capillary electrophoresis (CE) coupled with mass spectrometry (MS) is a promising alternative to conventional liquid chromatography (LC) MS in chemical and biological sample analysis due to its high resolving power and fast separation speed. Reproducibility and ruggedness problems, suffered to a certain degree by almost all the CE-MS interfaces, limit its broad applications. We present the development of a new sheathless CE-MS interface aiming at overcoming these problems and pushing CE-MS suitable to routine sample analysis with high sensitivity. A systematic evaluation of the new interface was performed using a hybrid Capillary Isotachophoresis (CITP) and Capillary zone electrophoresis (CZE) separation coupled with electrospray ionization (ESI) MS for its achievable sensitivity and reproducibility in sample quantification.
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Capillary electrophoresis nanoelectrospray ionization selected reaction monitoring mass spectrometry via a true sheathless metal coated emitter interface for robust and high sensitivity sample quantification
Analytical Chemistry, 2016Co-Authors: Thomas L. Fillmore, Keqi TangAbstract:A new sheathless transient Capillary Isotachophoresis (CITP)/Capillary zone electrophoresis (CZE)–MS interface, based on a commercially available Capillary with an integrated metal-coated ESI emitter, was developed in this study aiming at overcoming the reproducibility and ruggedness problems suffered to a certain degree by almost all the available CE–MS interfaces, and pushing the CE–MS technology suitable for routine sample analysis with high sensitivity. The new CITP/CZE–MS interface allows the electric contact between ESI voltage power supply and the CE separation liquid by using a conductive liquid that comes in contact with the metal-coated surface of the ESI emitter, making it a true sheathless CE–MS interface. Stable electrospray was established by avoiding the formation of gas bubbles from electrochemical reaction inside the CE Capillary. Crucial operating parameters, such as sample loading volume, flow rate, and separation voltage, were systematically evaluated for their effects on both CITP/CZE...
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Capillary Isotachophoresis nanoelectrospray ionization selected reaction monitoring ms via a novel sheathless interface for high sensitivity sample quantification
Analytical Chemistry, 2013Co-Authors: Chenchen Wang, Richard D. Smith, Cheng S. Lee, Keqi TangAbstract:A novel sheathless Capillary Isotachophoresis (CITP/CZE)–mass spectrometry (MS) interface featuring a large inner diameter (i.d.) separation Capillary, and a detachable small i.d. porous electrospray ionization (ESI) emitter was developed in this study to simultaneously achieve large sample loading capacity and stable nanoESI operation. Crucial operating parameters, including sample loading volume, flow rate, and separation window, were systematically investigated to attain optimum CITP/CZE separation efficiency and MS detection sensitivity. The performance of CITP/CZE–nanoESI-MS using the new sheathless interface was evaluated for its achievable low limit of quantification (LOQ) by analyzing targeted peptides, leu-enkephalin and angiotensin II, spiked in a BSA tryptic digest matrix at different concentrations. A linear dynamic range spanning 4.5 orders of magnitude and a 10 pM LOQ with measurement reproducibility of the CV < 22% were obtained experimentally for both targeted peptides, representing a 5-fo...
Petr Boček - One of the best experts on this subject based on the ideXlab platform.
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analytical Capillary Isotachophoresis after 50 years of development recent progress 2014 2016
Electrophoresis, 2017Co-Authors: Zdena Mala, Petr Gebauer, Petr BočekAbstract:This review brings a survey of papers on analytical ITP published since 2014 until the first quarter of 2016. The 50th anniversary of ITP as a modern analytical method offers the opportunity to present a brief view on its beginnings and to discuss the present state of the art from the viewpoint of the history of its development. Reviewed papers from the field of theory and principles confirm the continuing importance of computer simulations in the discovery of new and unexpected phenomena. The strongly developing field of instrumentation and techniques shows novel channel methodologies including use of porous media and new on-chip assays, where ITP is often included in a preseparative or even preparative function. A number of new analytical applications are reported, with ITP appearing almost exclusively in combination with other principles and methods.
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recent progress in Capillary Isotachophoresis
Electrophoresis, 2000Co-Authors: Petr Gebauer, Petr BočekAbstract:This article is a continuation of previous reviews and summarizes the progress of analytical Capillary Isotachophoresis since 2000. Papers reviewed include methodological and instrumental aspects as well as analytical applications. Included are also papers using Isotachophoresis and/or isotachophoretic principles as part of multidimensional separation schemes.
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analysis of orotic acid in human urine by on line combination of Capillary Isotachophoresis and zone electrophoresis
Journal of Chromatography A, 1999Co-Authors: Andrea Prochazkova, Ludmila Křivánková, Petr BočekAbstract:Abstract The techniques of the on-line combination of Capillary Isotachophoresis with zone electrophoresis in two coupled capillaries (ITP–CZE) and a single Capillary zone electrophoresis (CZE) were used for the sensitive determination of orotic acid (OA) in human urine. The simple CZE system was successfully applied for fast and reliable analyses of urine of healthy adult volunteers (the detection limit 1.7·10−6 M OA, the total time of analysis 6 min). However, this method failed in analyses of OA in urine of ill children due to more complex matrix of the samples. Here, the ITP preconcentration and preseparation step coupled on-line with CZE proved to serve well with an electrolyte system developed and optimized for this purpose. The maximum selectivity and resolution of OA from other sample constituents in ITP–CZE was achieved by use of an electrolyte system of very low pH 2.15 both for ITP and CZE stage. The sensitivity of detection and simplicity of OA identification were enhanced by use of an external UV scanning detector. High sensitivity of ITP–CZE combination (limit of detection 3·10−7 M OA), low sample consumption (1 μl), good reproducibility of migration times (inter-day RSD
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Synergism of Capillary Isotachophoresis and Capillary zone electrophoresis.
Journal of chromatography. B Biomedical sciences and applications, 1997Co-Authors: Ludmila Křivánková, Petr BočekAbstract:The combination of Capillary Isotachophoresis and Capillary zone electrophoresis may enhance greatly the performance of analytical Capillary electrophoresis with respect to both separation power and the concentration sensitivity. The concentrating effects and the separation power of Isotachophoresis allow the analysis of diluted samples and the elimination of interferences due to bulk components. The separation process of zone electrophoresis enables one to resolve the stack of trace analytes and detect the resulting individual zones with high sensitivity. The transition of Isotachophoresis into zone electrophoresis plays the key role in the overall performance of this hyphenated technique. This article describes the dynamics of the conversion of Isotachophoresis into zone electrophoretic mode and shows that the key role is played by the segments of the leading and terminating zones from the isotachophoretic stage. The magnitude of these segments directly effects the detection time as well as the separation width of the peaks of analytes. It is shown that these effects are also important in the analyses by Capillary zone electrophoresis where Isotachophoresis is induced by the sample itself. Finally, the paper presents a list of recommended, user-friendly, electrolyte systems which enable one to simply predict the performance of the combination Isotachophoresis-zone electrophoresis.
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options in electrolyte systems for on line combined Capillary Isotachophoresis and Capillary zone electrophoresis
Journal of Chromatography A, 1993Co-Authors: Ludmila Křivánková, Petr Gebauer, Wolfgang Thormann, Richard A Mosher, Petr BočekAbstract:Abstract A theoretical description of the electrolyte systems that can be used in the on-line combination of Isotachophoresis and zone electrophoresis is given. A classification of these systems is presented, based on the type of electrolyte used for the zone electrophoretic separation step. It is shown that transient sample stacking effects always persist from the isotachophoretic step to the beginning of the zone electrophoretic step and that they may negatively influence the zone electrophoretic separation and detection of the sample components. A mathematical description of these effects is given that allows the calculation of their magnitude and consequently the selection of operating conditions such that the stacking is decreased to an acceptable extent. In order to verify the reliability of the theoretical model, a modified PC simulation pack was prepared and used for investigating the behaviour of some model systems.
Hanno Stutz - One of the best experts on this subject based on the ideXlab platform.
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monitoring of deamidation and lanthionine formation in recombinant mugwort allergen by Capillary zone electrophoresis cze uv and transient Capillary Isotachophoresis cze electrospray ionization tof ms
Analytical Chemistry, 2018Co-Authors: Lorenz Stock, Sabrina Wildner, Christof Regl, Gabriele Gadermaier, Christian G. Huber, Hanno StutzAbstract:The response to thermal stress is an important parameter relevant for characterizing the biological activity and long-term stability of recombinant proteins, which may show irreversible, pH dependent structural changes under these conditions. We selected the recombinant pollen allergen of mugwort (Artemisia vulgaris) rArt v 3.0201 as a relevant model to study structural changes due to thermal and pH stress by means of Capillary zone electrophoresis (CZE)-UV and Capillary zone electrophoresis (CZE)-electrospray ionization (ESI)-TOF-MS. Therefore, this recombinant protein was exposed to 95 °C under acidic (pH 3.4) and slightly alkaline (pH 7.3) conditions for up to 120 min. CZE-UV data showed a continuous degradation of the allergen accompanied by the gradual formation of several reaction products. Characterization of novel allergen variants occurring at longer migration times was done via CZE-ESI-TOF-MS using in-Capillary transient Capillary Isotachophoresis (tCITP) preconcentration to facilitate the ident...
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Monitoring of Deamidation and Lanthionine Formation in Recombinant Mugwort Allergen by Capillary Zone Electrophoresis (CZE)-UV and Transient Capillary Isotachophoresis-CZE-Electrospray Ionization-TOF-MS
2018Co-Authors: Lorenz G. Stock, Sabrina Wildner, Christof Regl, Gabriele Gadermaier, Christian G. Huber, Hanno StutzAbstract:The response to thermal stress is an important parameter relevant for characterizing the biological activity and long-term stability of recombinant proteins, which may show irreversible, pH dependent structural changes under these conditions. We selected the recombinant pollen allergen of mugwort (Artemisia vulgaris) rArt v 3.0201 as a relevant model to study structural changes due to thermal and pH stress by means of Capillary zone electrophoresis (CZE)-UV and Capillary zone electrophoresis (CZE)-electrospray ionization (ESI)-TOF-MS. Therefore, this recombinant protein was exposed to 95 °C under acidic (pH 3.4) and slightly alkaline (pH 7.3) conditions for up to 120 min. CZE-UV data showed a continuous degradation of the allergen accompanied by the gradual formation of several reaction products. Characterization of novel allergen variants occurring at longer migration times was done via CZE-ESI-TOF-MS using in-Capillary transient Capillary Isotachophoresis (tCITP) preconcentration to facilitate the identification of minor variants. MS data revealed various modifications of rArt v 3.0201 in response to heating. Variants with deamidations and sulfur-related modifications including both yield and loss of sulfur were identified at increased migration times. Desulfurization produced allergen variants with up to four lanthionines that replaced initial disulfide bonds. In addition, mass spectra revealed shifts in the charge state distribution which indicate concomitant conformational alterations. Moreover, several low-abundant oxidized variants were identified. With extended thermal stress, the portfolio of variants increased and progressively shifted toward rArt v 3.0201 with high lanthionine content. The kinetics of conversion and the complexity of variant composition were pH dependent and increased under alkaline conditions
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Capillary zone electrophoresis of metal binding proteins in formic acid with uv and mass spectrometric detection using cationic transient Capillary Isotachophoresis for preconcentration
Electrophoresis, 2004Co-Authors: Hanno Stutz, Guy Bordin, Adela Rosa RodriguezAbstract:A Capillary zone electrophoresis (CZE) method with preceding cationic transient Capillary Isotachophoresis (tCITP-CZE) was developed for uncoated fused-silica capillaries to analyze metal-binding proteins (MBPs) of clinical relevance. UV detection was followed by mass spectrometry (MS). Optimization was done with model proteins of properties similar to relevant human MBPs. Using 1.0 mol.L - 1 formic acid (pH 1.78) as electrolyte resulted in up to 165000 plates m - 1 in CZE and 230000 plates m - 1 in combination with tCITP and analysis time was less than 5 min in uncoupled mode. Cationic tCITP with 125 mmol.L - 1 ammonium formate, buffered to pH 4.00, as leading electrolyte improved sample loadability considerably in comparison with sample stacking without impairing resolution. Following systematic optimization of the electrospray ionization process (ESI) the coupled system ((tCITP)-CZE-UV-ESI-MS) was tested with protein model mixtures and human MBPs. Repeatability of migration times was < 0.64% in pure CZE mode and in tCITP-CZE mode and < 0.83% in CZE-ESI-MS coupled mode. Mass accuracy was < 0.015%. Limits of detection were found to be in the range 50-160 fmol.
František Kvasnička - One of the best experts on this subject based on the ideXlab platform.
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Determination of glycyrrhizin in liqueurs by on-line coupled Capillary Isotachophoresis with Capillary zone electrophoresis
Journal of chromatography. A, 2007Co-Authors: František Kvasnička, Michal Voldřich, Josef VyhnálekAbstract:Abstract An on-line coupled Capillary Isotachophoresis-Capillary zone electrophoresis method for the determination of glycyrrhizin in liqueurs is described. The optimised electrolyte system was 5 mM HCl + 11 mM ɛ-aminocaproic acid + 0.05% hydroxyethylcellulose + 30% methanol (leading electrolyte), 5 mM caproic acid + 30% methanol (terminating electrolyte) and 20 mM caproic acid + 10 mM histidine + 0.1% hydroxyethylcellulose + 30% methanol (background electrolyte). Method characteristics, i.e ., linearity (20–500 ng/ml), accuracy (recovery 99 ± 4%), intra-assay repeatability (2%), intermediate repeatability (3.8%) and detection limit (8 ng/ml) were determined. Speed of analysis, low laboriousness, high sensitivity and low-running cost are the typical attributes of the Capillary Isotachophoresis-Capillary zone electrophoresis method. Developed method was successfully applied to analysis of liqueurs with liquorice extract and some foods (sweets and food supplements) containing liquorice. Found levels of glycyrrhizin in liqueurs, sweets and food supplements varied between 1–16 mg/l, 850–1050 mg/kg and 1.6–1.8 g/kg, respectively.
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determination of egg white lysozyme by on line coupled Capillary Isotachophoresis with Capillary zone electrophoresis
Electrophoresis, 2003Co-Authors: František KvasničkaAbstract: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.
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determination of isocitric acid in citrus juice a comparison of hplc enzyme set and Capillary Isotachophoresis methods
Journal of Food Composition and Analysis, 2002Co-Authors: František Kvasnička, Michal Voldřich, Ivan VinšAbstract:Abstract Three different analytical procedures for the determination of dl -isocitric acid content in citrus fruit juices were compared. The dl -isocitric acid is an important marker in multicomponent procedures for the evaluation of authenticity and quality of fruit products. The compared procedures were the new proposed Capillary Isotachophoresis (CITP) procedure using coupled two capillaries, reverse-phase high-pressure liquid chromatography (HPLC) method and enzymic procedure (Boehringer Mannheim). The optimized CITP electrolyte system was: 6m m HCl+3.8m m BisTrisPropan+2m m CaCl 2 +0.05% hydroxypropylmethyl cellulose (leading electrolyte) and 5m m morpholinethanesulphonic acid+1sssssm m BisTrisPropan (terminating electrolyte). The best correlation was found between the CITP and enzymic procedure ( r =0.956, n =6), the other cross-correlation coefficients were 0.850 and 0.748 for HPLC–enzymic and HPLC–CITP, respectively.
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new configuration in Capillary Isotachophoresis Capillary zone electrophoresis coupling
Journal of Chromatography A, 2001Co-Authors: František Kvasnička, Michal Jaros, Bohuslav GasAbstract:The paper surveys possible configurations of a coupling Capillary column operating in various electromigration modes. Special attention is given to Capillary Isotachophoresis-Capillary zone electrophoresis (cITP-CZE) coupling and its description from the theoretical point of view. Computer simulations of separation are presented and compared with experiments. Further, we propose a new configuration of electrolyte systems in cITP-CZE coupling, which offers a possibility to perform complex analyses of micro- and macro-constituents in one run. The electrolyte system is verified by practical experiments for both anionic and cationic modes of analysis. The advantages and disadvantages of the new combination are discussed.
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determination of fumaric acid in apple juice by on line coupled Capillary Isotachophoresis Capillary zone electrophoresis with uv detection
Journal of Chromatography A, 2000Co-Authors: František Kvasnička, Michal VoldrichAbstract:Abstract An on-line coupled Capillary Isotachophoresis–Capillary zone electrophoresis (cITP–CZE) method for the determination of the fumaric acid content in apple juice is presented. A clear separation of fumaric acid in real samples is achieved within 20 min. The leading, terminating and background electrolyte of the employed system consist of 10 m M HCl+β-alanine+5 m M β-cyclodextrin+0.05% hydroxypropylmethylcelullose (HPMC), pH 3, 10 m M citric acid and 20 m M citric acid+β-alanine+5 m M β-cyclodextrin+0.1% HPMC, pH 3.3, respectively. The linearity, recovery, repeatability and detection limit of the developed method are 25–1000 ng/ml, 1.07%, 95.4±3.5 (± s )% and 10 ng/ml, respectively. Low laboriousness (no sample pretreatment), sufficient sensitivity and low running cost are the important attributes of the cITP–CZE method which was successfully applied to analyses of real samples of apple juices.
Ludmila Křivánková - One of the best experts on this subject based on the ideXlab platform.
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Determination of ethyl glucuronide in human serum by hyphenation of Capillary Isotachophoresis and zone electrophoresis.
Electrophoresis, 2008Co-Authors: Michaela Nováková, Ludmila KřivánkováAbstract:The determination of ethyl glucuronide (EtG), a marker of recent alcohol consumption, in human serum by hyphenation of Capillary ITP (CITP) and CZE is reported. For CITP step, 1 x 10(-2) M hydrochloric acid adjusted with epsilon-aminocaproic acid (EACA) to pH 4.4 was used as the leading electrolyte, and 1 x 10(-2) M nicotinic acid with EACA, pH 4.4, was used as the terminating electrolyte (TE). All electrolytes contained 0.2% hydroxypropylcellulose to suppress electroosmosis. In CITP, EtG was separated from fast serum macrocomponents chloride, phosphate, lactate, and acetate. Zones of microcomponents including EtG that migrated between acetate and nicotinate were forwarded to the second Capillary filled with a BGE identical with the TE. Conductivity detection was used in the CITP step. Sensitive detection in the CZE step was performed using indirect spectrophotometric detection at 254 nm. The assay is based on a 1:5 dilution of serum with deionized water and has a concentration LOD for EtG in diluted sample of 9.8 x 10(-9) M. The method was used for the determination of EtG in sera of volunteers consuming alcohol.
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determination of iodide in samples with complex matrices by hyphenation of Capillary Isotachophoresis and zone electrophoresis
Electrophoresis, 2007Co-Authors: Pavla Pantůckova, Marek Urbanek, Ludmila KřivánkováAbstract:A method has been developed for the determination of iodide in mineral water, seawater, cooking salt, serum, and urine based on hyphenation of Capillary ITP and zone electrophoresis. A commercially available instrumentation for Capillary ITP with column-switching system was used. ITP served for removal of chloride present in the analyzed samples in a ratio of 10(6)-10(7):1 to iodide, zone electrophoresis was used for evaluation. Isotachophoretic separation proceeded in a Capillary made of fluorinated ethylene-propylene copolymer of 0.8 mm id and 90 mm total length to the bifurcation point filled with a leading electrolyte (LE) composed of 8 mM HCl + 16 mM beta-alanine (beta-Ala) + 10% PVP + 2.86 mM N(2)H(4)x2HCl, pH 3.2; and a terminating electrolyte composed of 8 mM H(3)PO(4) + 16 mM beta-Ala + 10% PVP + 5 mM N(2)H(4), pH 3.85 for all the matrices except seawater. For ITP of seawater the LE consisted of 50 mM HCl + 100 mM beta-Ala + 10% PVP + 2.86 mM N(2)H(4)x2HCl, pH 3.52. Distance of conductivity detector from the injection point and bifurcation point was 52 and 38 mm, respectively. Zone electrophoresis was performed in a Capillary made of fused silica of 0.3 mm id and 160 mm total length filled with LE from isotachophoretic step. LODs reached for all matrices were 2-3x10(-8) M concentration (2.5-4 microg/L) enabled monitoring of iodide in all analyzed samples with RSD 0.4-9.3%. Estimated concentrations of iodide in individual matrices were 10(-6)-10(-8) M.
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analysis of orotic acid in human urine by on line combination of Capillary Isotachophoresis and zone electrophoresis
Journal of Chromatography A, 1999Co-Authors: Andrea Prochazkova, Ludmila Křivánková, Petr BočekAbstract:Abstract The techniques of the on-line combination of Capillary Isotachophoresis with zone electrophoresis in two coupled capillaries (ITP–CZE) and a single Capillary zone electrophoresis (CZE) were used for the sensitive determination of orotic acid (OA) in human urine. The simple CZE system was successfully applied for fast and reliable analyses of urine of healthy adult volunteers (the detection limit 1.7·10−6 M OA, the total time of analysis 6 min). However, this method failed in analyses of OA in urine of ill children due to more complex matrix of the samples. Here, the ITP preconcentration and preseparation step coupled on-line with CZE proved to serve well with an electrolyte system developed and optimized for this purpose. The maximum selectivity and resolution of OA from other sample constituents in ITP–CZE was achieved by use of an electrolyte system of very low pH 2.15 both for ITP and CZE stage. The sensitivity of detection and simplicity of OA identification were enhanced by use of an external UV scanning detector. High sensitivity of ITP–CZE combination (limit of detection 3·10−7 M OA), low sample consumption (1 μl), good reproducibility of migration times (inter-day RSD
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Synergism of Capillary Isotachophoresis and Capillary zone electrophoresis.
Journal of chromatography. B Biomedical sciences and applications, 1997Co-Authors: Ludmila Křivánková, Petr BočekAbstract:The combination of Capillary Isotachophoresis and Capillary zone electrophoresis may enhance greatly the performance of analytical Capillary electrophoresis with respect to both separation power and the concentration sensitivity. The concentrating effects and the separation power of Isotachophoresis allow the analysis of diluted samples and the elimination of interferences due to bulk components. The separation process of zone electrophoresis enables one to resolve the stack of trace analytes and detect the resulting individual zones with high sensitivity. The transition of Isotachophoresis into zone electrophoresis plays the key role in the overall performance of this hyphenated technique. This article describes the dynamics of the conversion of Isotachophoresis into zone electrophoretic mode and shows that the key role is played by the segments of the leading and terminating zones from the isotachophoretic stage. The magnitude of these segments directly effects the detection time as well as the separation width of the peaks of analytes. It is shown that these effects are also important in the analyses by Capillary zone electrophoresis where Isotachophoresis is induced by the sample itself. Finally, the paper presents a list of recommended, user-friendly, electrolyte systems which enable one to simply predict the performance of the combination Isotachophoresis-zone electrophoresis.
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options in electrolyte systems for on line combined Capillary Isotachophoresis and Capillary zone electrophoresis
Journal of Chromatography A, 1993Co-Authors: Ludmila Křivánková, Petr Gebauer, Wolfgang Thormann, Richard A Mosher, Petr BočekAbstract:Abstract A theoretical description of the electrolyte systems that can be used in the on-line combination of Isotachophoresis and zone electrophoresis is given. A classification of these systems is presented, based on the type of electrolyte used for the zone electrophoretic separation step. It is shown that transient sample stacking effects always persist from the isotachophoretic step to the beginning of the zone electrophoretic step and that they may negatively influence the zone electrophoretic separation and detection of the sample components. A mathematical description of these effects is given that allows the calculation of their magnitude and consequently the selection of operating conditions such that the stacking is decreased to an acceptable extent. In order to verify the reliability of the theoretical model, a modified PC simulation pack was prepared and used for investigating the behaviour of some model systems.
