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Capillary Electrophoresis (capillary + electrophoresis)
Kinds of Capillary Electrophoresis Terms modified by Capillary Electrophoresis Selected AbstractsCoupling Capillary Electrophoresis and Pulsed Electrochemical DetectionELECTROANALYSIS, Issue 13 2005Carlos Abstract Pulsed electrochemical detection (PED) is an excellent method for detection of analytes that normally foul electrodes. In PED, the detection electrode is first cleaned at a high positive potential, then reactivated at a negative potential dissolving the surface oxide, and finally used to oxidize the analyte at a moderate positive potential. Due to the advantages and versatility of PED, many different variations of the detection waveform can be found in literature. This review focuses on application of PED to CE and in particular, the most commonly used modes: pulsed amperometric detection (PAD) and integrated pulsed amperometric detection (iPAD). [source] Contactless Conductivity Detection in Capillary Electrophoresis: A ReviewELECTROANALYSIS, Issue 24 2004Pavel Kubá Abstract The popularity of contactless conductivity detection in capillary electrophoresis has been growing steadily over the last few years. Improvements have been made in the design of the detector in order to facilitate its handling, to allow easy incorporation into available instruments or to achieve higher sensitivity. The understanding of its fundamental working principles has been advanced and the detection approach has also been transferred to lab-on-chip devices. The range of applications has been extended greatly from the initial work on small inorganic ions to include organic species and biomolecules. Concurrent determination of cations and anions by dual injection from opposite ends has been demonstrated as well as sample introduction by using flow-injection systems for easy automation of the process. [source] Determination of Reserpine in Urine by Capillary Electrophoresis with Electrochemiluminescence DetectionELECTROANALYSIS, Issue 3 2004Weidong Cao Abstract A fast and sensitive approach to detect reserpine in urine using micellar electrokinetic capillary chromatography with electrochemiluminescence (ECL) of Ru(bpy)32+ detection is described. Using a 25,,m i.d. capillary as separation column, the ECL detector was coupled to the capillary in the absence of an electric field decoupler. Field-amplified injection was used to minimize the effect of ionic strength in the sample and to achieve high sensitivity. In this way, the sample was analyzed directly without any pretreatment. The method was validated for reserpine in the urine over the range of 1×10,6,1×10,4,mol/L with a correlation coefficient of 0.996. The RSD for reserpine at a level of 5,,mol/L was 4.3%. The LOD (S/N=3) was estimated to be 7.0×10,8,mol/L. The average recoveries for 10,,mol/L reserpine spiked in human urine were 94%. [source] Cover Picture: Electrophoresis 19'2009ELECTROPHORESIS, Issue 19 2009Article first published online: 2 OCT 200 Issue no. 19 is a special issue on "Electrochemistry in Microfluidics and Capillary Electrophoresis". It has 25 contributions providing an "actual overview on the different aspects of electrochemistry in general (not necessarily only electrochemical detection) involving principles, designs, and applications in microfluidics (not necessarily only in electrophoretic mode) and CE". [source] Cover Picture: Electrophoresis 3'09ELECTROPHORESIS, Issue 3 2009Article first published online: 11 FEB 200 This is a regular issue with an emphasis on "Fundamentals Methodologies and Instrumentation" assembling 11 articles in various research areas on fundamentals, methods development, instrumental design, detection and sensitivity enhancement approaches. The remaining articles are on proteins and proteomics analyses by various electrophoretic approaches. Selected topics of issue 3 are: Capillary Electrophoresis-based detection of Methicillin-resistant Staphylococcus aureus (MRSA CE-based detection of methicillin-resistant Staphylococcus aureus A portable capillary electropherograph equipped with a cross-sampler and a contactless-conductivity detector for the detection of the degradation products of chemical warfare agents in soil extracts Two-dimensional phosphate-affinity gel electrophoresis for the analysis of phosphoprotein isotypes [source] Cover Picture: Electrophoresis 8/2008ELECTROPHORESIS, Issue 8 2008Article first published online: 17 APR 200 Regular issues provide a wide range of research and review articles covering all aspects of electrophoresis. Here you will find cutting-edge articles on methods and theory, instrumentation, nucleic acids, CE and CEC, miniaturization and microfluidics, proteomics and two-dimensional electrophoresis. On April 2 Professor Hjertén celebrated his 80th birthday, and it is an honor to take this opportunity to congratulate him on this special occasion and at the same time on his fruitful work. Stellan Hjertén's distinguished personality in research and life makes this celebration very special. It is therefore appropriate to devote a separate laudation in ELECTROPHORESIS to his achievements through which he has attained renown within the separation science community: indeed, he is considered undoubtedly to be the "Father of Capillary Electrophoresis". Professor Hjertén's preliminary work with Arne Tiselius motivated him to commit his career to electrophoresis: the development of free zone electrophoresis certainly revolutionized separation science, and since the construction of the first "capillary electrophoresis" equipment, one of the most cited works in this field carries his name. His friends were very keen to contribute manuscripts to this Issue, covering almost all areas in which Professor Hjertén has worked in his distinguished career. [source] Comparison of Cyclodextrin-Dipeptide Inclusion Complexes in the Absence and Presence of Urea by Means of Capillary Electrophoresis, Nuclear Magnetic Resonance and Molecular ModelingEUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 18 2007Benjamin Waibel Abstract The use of capillary electrophoresis (CE) modified with cyclodextrin (CD) for the separation of stereoisomers of peptides is well established. To increase the solubility of ,-CD, urea is often added to the buffer which may influence the complexation of a CD with a guest molecule. The aim of the present study was to investigate the influence of urea on the complexation between dipeptides and ,-CD using Ala-Phe and Ala-Tyr as model compounds. For this purpose three different analytical methods were employed: capillary electrophoresis (CE), 1H-NMR spectroscopy and molecular dynamics simulations (MD). Electropherograms of the peptide enantiomers were different in the presence and absence of urea. For example, at pH,2.5 in the absence of urea the enantiomers of Ala-Tyr are not separated in contrast to the use of buffers containing urea. Applying "complexation-induced chemical shift (CICS)" in NMR spectroscopy and rotating frame Overhauser enhancement spectroscopy (ROESY) revealed differences in the complexation of the peptide enantiomers by ,-CD in the absence and presence of urea suggesting the stabilization of the complex through the phenolic hydroxyl group of tyrosine. MD simulations for different complexes were carried out with consideration of both water and urea molecules in solution. Simulations were performed for 1 ns. In conclusion, NMR spectroscopy and MD methods help to understand the structure of peptide-CD complexes and the separation and migration behavior in CE. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007) [source] Quantitative Determination of Surface Concentration of Human Apolipoprotein H with Capillary ElectrophoresisIUBMB LIFE, Issue 5 2000Shao-xiong Wang Abstract The phospholipid monolayer at an air/water interface is widely used to mimic the biological membrane. The dynamic process of the protein or peptide interacting with lipid molecules can be reflected in the change in surface pressure of the monolayer. But the conventional method used to measure the surface pressure change gives results that cannot easily be correlated with the contribution of a single protein molecule. Previously, measuring the surface concentration of the protein molecules at the air/water interface has required the protein to be labeled with radioactivity or fluorescence. Here, a new method using capillary electrophoresis is introduced to measure the surface concentration of the protein. The results show at least two advantages of the new method: The numerical results of protein concentration can be obtained in a more precise and rapid way; and there is no need to label the protein sample or to build a special monolayer setup. [source] Pesticide analysis by capillary electrophoresisJOURNAL OF SEPARATION SCIENCE, JSS, Issue 12 2004J. Hernández-Borges Abstract In this work, a critical and updated revision of the current situation of the analysis of pesticides by Capillary Electrophoresis (CE) is presented. The review has been written in two main sections. The first one presents a thorough revision of the various off-line and on-line sample preconcentration procedures that have been used in conjunction with CE to analyze these compounds. The second part reviews the various detection strategies (i. e., UV, LIF, MS, and electrochemical) and CE modes that have been applied to the analysis of pesticides. Future trends that can be expected from this hot research area are also discussed. [source] Differential Increase in Taurine Levels by Low-Dose Ethanol in the Dorsal and Ventral Striatum Revealed by Microdialysis With On-Line Capillary ElectrophoresisALCOHOLISM, Issue 7 2004A Smith Ethanol increases taurine efflux in the nucleus accumbens or ventral striatum (VS), a dopaminergic terminal region involved in positive reinforcement. However, this has been found only at ethanol doses above 1 g/kg intraperitoneally, which is higher than what most rats will self-administer. We used a sensitive on-line assay of microdialysate content to test whether lower doses of ethanol selectively increase taurine efflux in VS as opposed to other dopaminergic regions not involved in reinforcement (e.g., dorsal striatum; DS). Adult male rats with microdialysis probes in VS or DS were injected with ethanol (0, 0.5, 1, and 2 g/kg intraperitoneally), and the amino acid content of the dialysate was measured every 11 sec using capillary electrophoresis and laser-induced fluorescence detection. In VS, 0.5 g/kg ethanol significantly increased taurine levels by 20% for 10 min. A similar increase was seen after 1 g/kg ethanol, which lasted for about 20 min after injection. A two-phased taurine efflux was observed with the 2.0 g/kg dose, where taurine was increased by 2-fold after 5 min but it remained elevated by 30% for at least 60 min. In contrast, DS exhibited much smaller dose-related increases in taurine. Glycine, glutamate, serine, and ,-aminobutyric acid were not systematically affected by lower doses of ethanol; however, 2 g/kg slowly decreased these amino acids in both brain regions during the hour after injection. These data implicate a possible role of taurine in the mechanism of action of ethanol in the VS. The high sensitivity and time resolution afforded by capillary electrophoresis and laser-induced fluorescence detection will be useful for detecting subtle changes of neuronally active amino acids levels due to low doses of ethanol. [source] Analysis of Platinum Adducts with DNA Nucleotides and Nucleosides by Capillary Electrophoresis Coupled to ESI-MS: Indications of Guanosine 5,-Monophosphate O6,N7 ChelationCHEMBIOCHEM, Issue 11 2004Ulrich Warnke Dr. Abstract DNA is the ultimate target of platinum-based anticancer therapy. Since the N7 of guanine is known to be the major binding site of cisplatin and its analogues, adduct formation with model nucleotides, especially 2,-deoxyguanosine 5,-monophosphate (dGMP), has been studied in detail. During the last few years a coupled capillary eletrophoresis/electrospray-ionization mass spectrometry (CE/ESI-MS) method has been advantageously used in order to separate and identify platinum adducts with nucleotides in submillimolar concentrations in aqueous solutions. Beside the bisadduct, [Pt(NH3)2(dNMP)2]2,(NMP=2,-deoxynucleoside 5,-monophosphate), and the well-known monochloro and monohydroxo adducts, [Pt(NH3)2Cl(dNMP)],and [Pt(NH3)2(dNMP)OH],, respectively, a third kind of monoadduct species with a composition of [Pt(NH3)2(dNMP)],can be separated by CE and detected through the m/z values measured with ESI-MS. Different experimental setups indicate the existence of an O6,N7 chelate, whereas the formation of N7,,PO4macrochelates or dinuclear species is unlikely. Additionally, offline MS experiments with 2,-deoxyguanosine (dG) and stabilization of the controversially discussed O6,N7 chelate by oxidation with hydrogen peroxide support the assumption of the existence of O6,N7 chelation. [source] Chiral Discrimination of Aromatic Amino Acids by Capillary Electrophoresis in (+)- and (-)-(18-Crown-6)-2,3,11,12-tetracarboxylic Acid Selector Modes.CHEMINFORM, Issue 50 2003Wonjae Lee Abstract For Abstract see ChemInform Abstract in Full Text. [source] Protein Fractionation of Cowpea (Vigna unguiculata (L.) Walp) Leaf, Flower and Seed by Capillary Electrophoresis and Its Potential for Variety IdentificationCHINESE JOURNAL OF CHEMISTRY, Issue 4 2010Sirithon Siriamornpun Abstract The proteins of different faction of cowpea [Vigna unguiculata (L.) Walp] were fractionated by capillary electrophoresis (CE). The extracting solvent system was one of the most critical factors in the optimization exercise. To improve reproducibility, seed samples needed to be defatted with chloroform/methanol (V:V=2:1) as preferred prior to protein extraction. Proteins were extracted from seeds, leaves and flowers with 50% aqueous 1-propanol and separated on a 50 (m×20 cm fused silica capillary column using a UV detector at 200 nm. Separation was conducted at constant voltage (10 kV, 40°C), using iminodiacetic acid (pH 2.5) containing 0.05% hydroxypropylmethylcellulose (HPMC) and 20% acetonitile. The results showed that proteins extracted from all fraction of cowpea were successfully separated by CE in less than 20 min. Seed extracts provided the greatest number of eluted protein peaks, followed by flower and leaf, respectively. The seed-protein extracts provided unique CE patterns for different varieties; hence the seed was the tissue chosen as being most suitable for variety identification. As a result, an optimized procedure was developed to provide rapid identification of cowpea varieties, based on capillary electrophoregram patterns. [source] Determination of Bioactive Components in Polygonum perfoliatum L. by Capillary Electrophoresis with Electrochemical DetectionCHINESE JOURNAL OF CHEMISTRY, Issue 4 2009Shuping JIN Abstract The major bioactive components in a Chinese herb named Polygonum perfoliatum L. including ferulic acid, vanillic acid, quercetin, caffeic acid and protocatechuic acid were simultaneously determined by capillary electrophoresis with electrochemical detection (CE-ED) in this paper. 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 five analytes could be separated within 17 min at a separation voltage of 18 kV in 10 mmol· L,1 phosphate buffer (pH 9.2). A 300 µm diameter carbon disk electrode generated good responses at +0.95 V (vs. SCE) to the five analytes. The response was linear over three orders of magnitude with detection limits (S/N=3) ranging from 7.1×10,8 to 9.3×10,8 g·mL,1 for the analytes. This proposed method could be successfully applied to the analysis of the real samples with relatively simple extraction procedures and satisfactory results. [source] Analysis of Trace Ingredients in Green Tea by Capillary Electrophoresis with Amperometric DetectionCHINESE JOURNAL OF CHEMISTRY, Issue 3 2008Ping LI Abstract In this paper, four trace ingredients (rutin, gallic acid, quercetin, chlorogenic acid) in green tea were simultaneously determined by capillary electrophoresis coupled with amperometric detection (CE-AD). Effects of several important factors such as the pH and concentration of running buffer, separation voltage, injection time and detection potential were investigated to acquire the optimum conditions. Under the optimum conditions, the analytes could be separated within 20 min at a separation voltage of 18 kV in a 60 mmol/L borate buffer (pH 8.7). A 300 µm diameter carbon disk electrode generated good responses at 950 mV (vs. SCE) for all analytes. The relationship between the peak currents and concentrations of the analytes was linear over about three orders of magnitude with detection limits (S/N=3) ranging from 1.0×10,7 to 1.0×10,4 g·mL,1 for all the analytes. This proposed method demonstrated long-term stability and reproducibility with relative standard deviations less than 3% for both migration time and peak current (n=7), which could be successfully used for the determination of the analytes in green tea with satisfactory assay results. [source] Fast Determination of Clenbuterol and Salbutamol in Feed and Meat Products Based on Miniaturized Capillary Electrophoresis with Amperometric DetectionCHINESE JOURNAL OF CHEMISTRY, Issue 12 2007Qing-Cui CHU Abstract The fast separation capability of a novel miniaturized capillary electrophoresis with an amperometric detection (,CE-AD) system was demonstrated by determining clenbuterol and salbutamol in real samples. The effects of several factors such as the acidity and concentration of the running buffer, the separation voltage, the applied potential and the injection time on CE-AD were examined and optimized. Under the optimum conditions, the two , -agonists could be baseline separated within 60 s at a separation voltage of 2 kV in a 90 mmol/L H3BO3 -Na2B4O7 running buffer (pH 7.4), which was not interfered by ascorbic acid and uric acid. Highly linear response was obtained for above compounds over three orders of magnitude with detection limits ranging from 1.20×10,7 to 6.50×10,8 mol/L (S/N=3). This method was successfully used in the analysis of feed and meat products with relatively simple extraction procedures. [source] Capillary Electrophoresis for the Simultaneous Determination of Metals by Using Ethylenediamine Tetraacetic Acid as Complexing Agent and Vancomycin as Complex SelectorCHINESE JOURNAL OF CHEMISTRY, Issue 12 2006Jirasak Threeprom Abstract A new separation system of capillary electrophoresis for the simultaneous determination of metals by using ethylenediamine tetraacetic acid (EDTA) as complexing agent and employing vancomycin as complex selector was described. The Z-shape cell capillary electrophoresis was used to enhance the sensitivity for the determination of the complexes of Cu(II), Ni(II), Co(II) and Fe(III) with EDTA. The partial filling method (co-current mode) was used in order to increase the selectivity of the electrophoretic method, meanwhile vancomycin was not present at the detector path during the detection of metal-EDTA complexes. The vancomycin concentration, phosphate concentration and pH of the buffer strongly influenced mobility, resolution and selectivity of the studied analytes. Under the optimal condition, the relative standard deviations (n=5) of the migration time and the peak area were less than 3.14% and 7.35%, respectively. Application of the Z-shape cell capillary electrophoresis method with UV detection and vancomycin loading led to the reliable determination of these metal ions in tap water and the recoveries were97%,101%. The detection limits based on a signal to noise ratio of 3:1 were found in the range of 2,10 µg·L,1. [source] Capillary electrophoresis for the analysis of contaminants in emerging food safety issues and food traceabilityELECTROPHORESIS, Issue 13 2010Belinda Vallejo-Cordoba Abstract This review presents an overview of the applicability of CE in the analysis of chemical and biological contaminants involved in emerging food safety issues. Additionally, CE-based genetic analyzers' usefulness as a unique tool in food traceability verification systems was presented. First, analytical approaches for the determination of melamine and specific food allergens in different foods were discussed. Second, natural toxin analysis by CE was updated from the last review reported in 2008. Finally, the analysis of prion proteins associated with the "mad cow" crises and the application of CE-based genetic analyzers for meat traceability were summarized. [source] Cover Picture: Electrophoresis 2'2010ELECTROPHORESIS, Issue 2 2010Article first published online: 18 JAN 2010 Issue no. 2 has 19 contributions including one Fast Track contribution on "Thermophoresis of ssDNA". The remaining 18 articles cover a wide variety of investigation involving nucleic acids, cells, viruses, interactive CE, detection methodologies in CE, enantioseparations, and studies on proteins and proteomics. Further selected articles include: Capillary electrophoresis of bone marrow stromal cells with uptake of heparin-functionalized poly(lactide-co-glycolide) nanoparticles during differentiation towards neurons. ((10.1002/elps.200900336)) Ghrelin-liposome interactions. Characterization of liposomal formulations of an acylated 28-amino acid peptide using capillary electrophoresis. ((10.1002/elps.200900394)) [source] Capillary electrophoresis of liposomes functionalized for protein bindingELECTROPHORESIS, Issue 20 2006Gerhard Bilek Abstract CE enabled assessing the attachment of hexa-histidine-tagged proteins to functionalized phospholipid liposomes. The liposomes were made of 1-palmitoyl-2-oleoyl- sn -glycero-3-phosphocholine, phosphatidyl-ethanolamine, cholesterol and distearoyl-glycero-3-phosphoethanolamine- N -methoxy(polyethylene glycol) in a molar ratio of 29:26:40:5. The unilamellar vesicles, which had an average diameter of 170,nm, were labelled by inclusion of FITC-dextran for fluorescence detection. CE was carried out in poly(vinyl alcohol) (PVA)-coated capillaries at 25°C with a BGE consisting of Tris-HCl (50,mM, pH,8.0). For conjugation of the liposomes with the proteins (soluble synthetic receptor fragments with molecular mass of 60 and 70,kDa, respectively), Ni2+ was implanted into the vesicle surface by an anchor lipid containing a nitrilotriacetate acid (NTA) group as complexation agent for the metal ions. The difference in surface charge enabled the separation of the different species of interest by CE: plain vesicles, vesicles functionalised with Ni-NTA, vesicle,protein complexes and the species formed upon removal of the Ni-ions by complexation with EDTA. Loss of the Ni-ions resulted in the release of the proteins and the reappearance of the plain Ni-free NTA-liposome species in the electropherograms. [source] Capillary electrophoresis of affinity complexes between subviral 80S particles of human rhinovirus and monoclonal antibody 2G2ELECTROPHORESIS, Issue 13 2006Leopold Kremser Dr. Abstract Human rhinoviruses (HRVs), the main etiologic agents of the common cold, transform into subviral B- or 80S particles (they sediment at 80S upon sucrose density gradient centrifugation) during infection and, in,vitro, upon exposure to a temperature between 50 and 56°C. With respect to the native virion they lack the genomic RNA and the viral capsid protein VP4. 80S particles are unstable and easily disintegrate into their components, VP1, VP2, and VP3 in buffers containing SDS. However, this detergent was found to be a necessary constituent of the BGE for the analysis of these viruses and their complexes with receptors and antibodies by CE. We here demonstrate that dodecylpoly(ethyleneglycol ether) (D-PEG) a nonionic detergent, is suitable for analysis of subviral particles as it preserves their integrity, in contrast to SDS. Electrophoresis of the 80S particles in borate buffer (pH,8.3, 100,mM) containing 10,mM D-PEG resulted in a well-defined electrophoretic peak. The identity of the peak was confirmed, among other means, by complexation with mAb,2G2, which recognizes a structural epitope exclusively present on subviral particles but not on native virus. Upon incubation of the 80S particles with mAb,2G2 the peak disappeared, but a new peak, attributed to the antibody complex emerged. The separation system allowed following the time course of the transformation of intact HRV serotype,2 into 80S particles upon incubation at temperatures between 40 and 65°C. We also demonstrate that subviral particles derived from HRV2 labeled with the fluorescence dyes FITC or Cy3.5 were stable in the separation system containing D-PEG. Dye-modified particles were still recognized by mAb,2G2, suggesting that the exposed lysines that are derivatized by the reagent do not form part of the epitope of the antibody. [source] Capillary electrophoresis using copolymers of different composition as physical coatings: A comparative studyELECTROPHORESIS, Issue 5-6 2006Guillaume L. Erny Abstract In this work, a comparative study on the use of different polymers as physically adsorbed coatings for CE is presented. It is demonstrated that the use of ad hoc synthesized polymers as coatings allows tailoring the EOF in CE increasing the flexibility of this analytical technique. Namely, different polymers were synthesized at our laboratory using different percentages of ethylpyrrolidine methacrylate (EpyM) and N,N -dimethylacrylamide (DMA). Thus, by modifying the percentage of EpyM and DMA monomers it is possible to manipulate the positive charge of the copolymer, varying the global electrical charge on the capillary wall and with that the EOF. These coated capillaries are obtained by simply flushing a given EpyM,DMA aqueous solution into bare silica capillaries. It is shown that by using these coated capillaries at adequate pHs, faster or more resolved CE separations can be achieved depending on the requirements of each analysis. Moreover, it is demonstrated that these coated capillaries reduce the electrostatic adsorption of basic proteins onto the capillary wall. Furthermore, EpyM,DMA coatings allow the reproducible chiral separation of enantiomers through the partial filling technique (PFT). The EpyM,DMA coated capillaries are demonstrated to provide reproducible EOF values independently of the pH and polymer composition with%RSD values lower than 2% for the same day. It is also demonstrated that the coating procedure is reproducible between capillaries. The compatibility of this coating protocol with CE in microchips is discussed. [source] Capillary electrophoresis versus differential scanning calorimetry for the analysis of free enzyme versus enzyme-ligand complexes: In the search of the ligand-free status of cholinesterasesELECTROPHORESIS, Issue 2 2006Daniel Rochu Dr. Abstract Cholinesterases (ChEs) are highly efficient biocatalysts whose active site is buried in a deep, narrow gorge. The talent of CE to discover inhibitors in the gorge of highly purified preparations has fairly altered the meaning of a ChE ligand-free status. To attempt at a description of this one, we investigated the stability of Bungarus fasciatus acetylcholinesterase (AChE), alone or complexed with different inhibitors. Determination of midtransition temperature for thermal denaturation, using differential scanning calorimetry (DSC) and CE, provided conflicting results. Discrepancies strongly question the reality of a ligand-free AChE state. DSC allowed estimation of the stability of AChE-ligands complexes, and to rank the stabilizing effect of different inhibitors. CE acted as a detector of hidden ligands, provided that they were charged, reversibly bound, and thus dissociable upon action of electric fields. Then, CE allowed quantification of the stability of ligand-free AChE. CE and DSC providing each fractional and nonredundant information, cautious attention must be paid for actual estimation of the conformational stability of ChEs. Because inhibitors used in purification of ChEs by affinity chromatography are charged, CE remains a leading method to estimate enzyme stability and detect the presence of bound hidden ligands. [source] Capillary electrophoresis of polycationic poly(amidoamine) dendrimersELECTROPHORESIS, Issue 15 2005Xiangyang Shi Abstract Generation,2 to generation,5 poly(amidoamine) (PAMAM) dendrimers having different terminal functionalities were analyzed by capillary electrophoresis (CE). Polyacrylamide gel electrophoresis was also used to assess the composition of the individual generations for comparison with the CE results. Separation of PAMAMs can be accomplished by either using uncoated silica or silanized silica capillaries, although reproducibility is poor using the uncoated silica capillary. To improve run-to-run reproducibility, silanized capillary was used and various internal standards were also tested. Relative and normalized migration times of primary amine terminated PAMAM dendrimers were then determined using 2,3-diaminopyridine (2,3-DAP) as an internal standard. Using silanized capillaries and internal standards, the relative and normalized migration times are fully reproducible and comparable between runs. Apparent dimensionless electrophoretic mobilities were determined and the results were compared to theoretical calculations. It is concluded that for PAMAMs a complex separation mechanism has to be considered in CE, where the movement of the ions is due to the electric field, but the separation is rather the consequence of the adsorption/desorption equilibria on the capillary wall ("electrokinetic capillary chromatography"). The described method may be used for quality control and may serve as an effective technique to analyze polycationic PAMAM dendrimers and their derivatives with different surface modifications. [source] Capillary electrophoresis of amphipathic ,-helical peptide diastereomersELECTROPHORESIS, Issue 1 2004Traian V. Popa Abstract We have made a rigorous assessment of the ability of capillary electrophoresis to resolve peptide diastereomers through its application to the separation of a series of synthetic 18-residue, amphipathic ,-helical monomeric peptide analogues, where a single site in the centre of the hydrophobic face of the ,-helix is substituted by 19 L - or D -amino acids. Such L - and D -peptide pairs have the same mass-to-charge ratio, amino acid sequence and intrinsic hydrophobicity, varying only in the stereochemistry of one residue. CE approaches assessed in their ability to separate diastereomeric peptide pairs included capillary zone electrophoresis (uncoated capillary), micellar electrokinetic chromatography (uncoated capillary in the presence of 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate, CHAPS), open-tubular capillary electrochromatography (C8 -coated capillary in the presence of 25% 2,2,2-trifluoroethanol (TFE) or 25% ethanol). Overall, the OT-CEC methods were the most effective at separating the most peptide pairs, particularly for those containing hydrophilic side chains. However, the MEKC approach proved most effective for separation of peptide pairs containing hydrophobic or aromatic side chains. [source] Capillary zone electrophoresis with a dynamic double coating for analysis of carbohydrate-deficient transferrin in human serum: Impact of resolution between disialo- and trisialotransferrin on reference limitsELECTROPHORESIS, Issue 24 2003Christian Lanz Abstract Capillary electrophoresis with a dynamic double coating formed by charged polymeric reagents represents a very effective tool for the separation of iron-saturated transferrin (Tf) isoforms and thus the determination of carbohydrate-deficient transferrin (CDT) in human serum. The resolution between di- and trisialo-Tf is dependent on the applied voltage and capillary temperature. With a 50 ,m inside diameter (ID) capillary of about 60 cm total length mounted into the P/ACE MDQ, 28 kV and 40°C, the resolution of the two Tf isoforms is shown to be between 1.0 and 1.4, whereas with reduced voltage and/or temperature, increased resolution at the expense of elongated run times is observed. Best data with complete resolution (Rs , 1.4) are obtained at 20 kV and 30°C. For the determination of CDT in serum, incomplete separation of di- and trisialo-Tf is demonstrated to have an impact on the reference limits. Analysis of the sera of 54 healthy individuals with no or moderate alcohol consumption and using valley-to-valley peak integration, the upper (lower) reference limits for CDT in relation to total Tf at the two power levels are 1.33 (0.52) and 1.57 (0.81)%, respectively, representing intervals that are significantly different (P < 0.001). Furthermore, the reference intervals are shown to be strongly dependent on the peak integration approach used. Valley-to-valley peak integration should only be employed for conditions with complete resolution between disialo- and trisialo-Tf. [source] Capillary electrophoresis as a probe of enantiospecific interactions between photoactive transition metal complexes and DNAELECTROPHORESIS, Issue 15 2003James P. Schaeper Abstract Recently, we have demonstrated the capacity to separate chiral transition metal (TM) complexes of the type [M(diimine)3]n+ using CE buffers containing chiral tartrate salts. In separate work, several chromium(III)- tris -diimine complexes in particular have been shown to bind enantioselectively with calf-thymus (CT) DNA, and a qualitative assessment of the relative strength and enantiospecificity of this interaction is of significant interest in the characterization of these complexes as potential DNA photocleavage agents. Here, we describe two convenient approaches to investigate such binding behavior using chiral CE. For complexes with lower DNA affinities exhibiting primarily surface binding, DNA itself is used as the chiral resolving agent in the electrophoretic buffer. In this approach, resolution of the TM complexes into their , and , isomers is achieved with the isomer eluting later exhibiting superior binding affinity toward DNA. For more strongly bound TM complexes containing ligands known to intercalate with DNA, the [Cr(diimine)3]3+ complexes are preincubated with oligonucleotide and subsequently enantiomerically resolved in a dibenzoyl- L -tartrate buffer system that facilitates analysis of the unbound TM species only. Differences in isomer binding affinity are distinguished by the relative peak areas of the ,- and ,-isomers, and relative binding strengths of different complexes can be inferred from comparison of the total amount of unbound complex at equivalent DNA/TM ratios. [source] Analysis of mitochondria by capillary electrophoresis: cardiolipin levels decrease in response to carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazoneEUROPEAN JOURNAL OF LIPID SCIENCE AND TECHNOLOGY, Issue 9 2010Wenfeng Zhao Abstract Cardiolipin is an important phospholipid present in the inner membrane of mitochondria. It plays a critical role in adenosine triphosphate (ATP) synthesis mediated by oxidative phosphorylation. Exposure of HepG2 cells to carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone (FCCP) caused the inhibition of ATP synthesis and the depolarization of mitochondria. Capillary electrophoresis with laser-induced fluorescence (CE-LIF) analysis of fluorescent mitochondrion-selective probe 10-N-nonyl acridine orange (NAO) labeled mitochondria was employed to in situ estimate the cardiolipin levels under FCCP-induced de-energization of mitochondria. NAO, stoichiometriclly bound to cardiolipin at a 1:1 or 2:1 molar ratio (NAO/cardiolipin), emitted green and red fluorescence, respectively. Green fluorescence was chosen for cardiolipin content analysis because it was more intense than red fluorescence. A significant decrease in the cardiolipin content, up to 11% of the control, was evident when the ATP content and mitochondrial membrane potential (MMP) correspondingly decreased. These related findings suggested that CE-LIF may provide a sensitive strategy to determine cardiolipin content in response to exposure to chemical uncouplers. This reinforces the hypothesis that alterations in ATP synthesis and MMP have a close association with cardiolipin content, which correlated tightly with mitochondrial membrane assembly and activity. [source] Kinetics of aspartic acid isomerization and enantiomerization in model aspartyl tripeptides under forced conditionsJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 10 2010Uwe Conrad Abstract The aim of the present study was the determination of the isomerization and enantiomerization of aspartic acid (Asp) in tripeptides. Capillary electrophoresis (CE) assays were developed and validated allowing the simultaneous determination of the diastereomeric ,- D/L -Asp and ,- D/L -Asp peptides. Rapid isomerization and enantiomerization were noted for peptides with the Phe-Asp-GlyOH sequence at pH 10 and 80°C while Gly-Asp-PheOH proved to be more stable due to the steric influence of the phenyl side chain. A kinetic model assuming a central role of the succinimide intermediate was used to fit the concentration versus time data. In incubations of L -Phe-,- L -Asp-GlyOH the ratio of ,-Asp/,-Asp peptides was about 1:4 in agreement with literature data. With regard to L -Asp and D -Asp peptides an ,-Asp/,-Asp ratio of about 1:3 and 1:5, respectively, was observed. The stereochemistry of Phe at the X,,,1 position affected the ratio of L -Asp/D -Asp implying an effect of the stereochemistry of neighboring amino acids on Asp enantiomerization. Modeling only overall Asp enantiomerization rate constants in accordance to literature data were observed for Asp peptides. In case of the asparagine (Asn) peptide the data could only be fitted to the models considering a direct conversion of L -Asn to a D -configured succinimide via an alternative pathway. © 2010 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 99:4162,4173, 2010 [source] Hydrolysis of aliphatic naphthalene diimides: effect of charge placement in the side chainsJOURNAL OF PHYSICAL ORGANIC CHEMISTRY, Issue 9 2008Michelle B. Kim Abstract Water-soluble naphthalene diimides (NDIs) have found uses in a wide variety of applications including as electron acceptors in electron transfer reactions and as molecules that undergo spontaneous organization in aqueous solution. Many studies have looked at their interaction with nucleic acids including work with DNA duplexes, triplexes, quadruplexes, hairpins, and DNA,RNA heteroduplexes. In many of these interactions the NDIs serve as threading intercalators. Herein we describe the reversible hydroxide-catalyzed hydrolysis of NDIs bearing aliphatic side chains, with ring opening first to the monoimide and then to the diamide. Examples with N -methylpyrrolidinium groups placed two (1) and three (5) atoms from the central core were studied. The Ka values for the first and second hydrolyses for 1 were 2.5,±,0.2,×,105 and 2.0,±,0.1,×,102,M,1, respectively; for 5 they were 1.4,±,0.1,×,105 and 44,±,2,M,1, respectively. NDI 1 hydrolyzed 6.8 times faster than 5. The rates for the first and second ring opening of 1 in 100,mM hydroxide measured by stopped-flow were 17.0,±,0.2 and 0.53,±,0.01,s,1, respectively. Capillary electrophoresis in borate buffer showed separation of the diimide and monoimide with the former eluting first. Nuclear magnetic resonance (NMR) showed both the syn and anti isomers of the diamide species. Overall, the rate of hydrolysis of the NDI is increased when the cationic charge is moved closer to the NDI core. Copyright © 2008 John Wiley & Sons, Ltd. [source] | ||||||||||||||||