Home About us Contact
|
Home About us Contact | ||
|
|
||
Derivatization
Kinds of Derivatization Terms modified by Derivatization Selected AbstractsDetermination of amino acids in rat vitreous perfusates by capillary electrophoresisELECTROPHORESIS, Issue 17 2004Kongthong Thongkhao-On Abstract In vivo determinations of amino acids are important for improving our understanding of physiological states of biological tissue function and dysfunction. However, the chemically complex matrix of different biological fluids complicates the assay of this important class of molecules. We introduce a method for characterizing the amino acid composition of submicroliter volumes of vitreous humor perfusates. Low-flow push-pull perfusion sampling is compatible with collecting small volume samples in a complicated matrix that are potentially difficult to separate. An efficient, sensitive, and rapid analysis of amino acids from in vivo perfusates of the vitreous is presented with 3-(4-carboxybenzoyl)-2-quinoline-carboxaldehyde (CBQCA) derivatitation and capillary electrophoresis (CE) separation with laser-induced fluorescence detection (LIF). Derivatization with CBQCA for up to 2 h provided high sensitivity and low detection limits at the nM level. Seventeen amino acids including D -serine (D -Ser) and D -aspartate (D -Asp) were resolved in less than 10 min. Importantly, D -Ser is separated from its enantiomeric pair. Characterization of vitreal amino acids with this assay technique will be useful for understanding ocular diseases and physiological mechanisms in vision. [source] Derivatization of inorganic ions in capillary electrophoresisELECTROPHORESIS, Issue 12-13 2003Audrius PadarauskasArticle first published online: 8 JUL 200 Abstract This review gives a short overview of the main approaches to the derivatization of inorganic ions in capillary electrophoresis (CE) with emphasis on the most recent works. Various derivatization procedures and detection methods are discussed. A brief account of their advantages and limitations is given. More specific areas such as microchip CE, simultaneous separation of anions and cations, and speciation analysis are also briefly discussed. [source] Systematic Studies on Photoluminescence of Oligo(arylene-ethynylene)s: Tunability of Excited States and Derivatization as Luminescent Labeling Probes for ProteinsEUROPEAN JOURNAL OF ORGANIC CHEMISTRY, Issue 14 2006Yong-Gang Zhi Abstract Functionalized oligo(phenylene-ethynylene)s (OPEs) with different conjugation lengths, p -X(C6H4C,C)nSiMe3 (n = 1,4; X = NH2, NMe2, H) were synthesized by Sonogashira coupling of (phenylene-ethynylene)s and 1-iodo-4-(trimethylsilylethynyl)benzene, followed by desilylation of the p -substituted (trimethylsilylethynyl)benzenes with potassium hydroxide. The photoluminescent properties for the OPE series with different chain lengths and their solvatochromic responses were examined. The absorption maxima were red-shifted with increasing numbers of ,(C6H4C,C), units (n), and a linear plot of the absorption energy maxima vs. 1/n was obtained for each series. The emission spectra in dichloromethane showed a broad and structureless band, the energies of which (in wavenumbers) also fit linearly with 1/n. Both the absorption and emission wavelength maxima of the NH2 - and NMe2 -substituted OPEs exhibited significant solvent dependence, whereas the parent OPEs (X = H) showed only minor shifts of the ,max values in different solvents. Substituent effects upon the photoluminescent characteristics of the OPEs and the tunability of the excited states were examined with the p -X(C6H4C,C)nSiMe3 (n = 2, 3; X = NH2, NMe2, H, SMe, OMe, OH, and F) series. The H- and F-substituted counterparts exhibited high-energy vibronically structured emissions attributed to the 3(,,*) excited states of the (arylene-ethynylene) backbone. For compounds bearing NH2 and NMe2 groups, a broad red-shifted emission with a remarkable Stokes shift from the respective absorption maximum was observed, which can be assigned to an n , ,* transition. The n , ,* assignment was supported by MO calculations on the model compounds p -X(C6H4C,C)2SiH3 (X = NH2, H). Functionalization of the oligo(arylene-ethynylene)s with the N -hydroxysuccinimidyl (NHS) moiety enabled covalent attachment of the fluorophore to HSA protein molecules. A series of fluorescent labels, namely p -X(C6H4C,C)nC6H4NHS, (n = 1, X = NH2, NMe2, SMe, OMe, OH, F; n = 2, X = NH2, NMe2) and p -Me2NC6H4C,C(C4H2S)C,CC6H4NHS were synthesized, and their conjugates with HSA (human serum albumin) were characterized by MALDI-TOF mass spectrometry, UV/Vis absorption spectroscopy, and gel electrophoresis. (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006) [source] Clenbuterol in the horse: urinary concentrations determined by ELISA and GC/MS after clinical dosesJOURNAL OF VETERINARY PHARMACOLOGY & THERAPEUTICS, Issue 1 2001J. D. Harkins Clenbuterol is a ,2 agonist/antagonist bronchodilator marketed as Ventipulmin® and is the only member of this group of drugs approved by the US Food and Drug Administration (FDA) for use in horses. Clenbuterol is a class 3 drug in the Association of Racing Commissioners International (ARCI) classification system; therefore, its identification in postrace samples may lead to sanctions. Recently, the sensitivity of postrace testing for clenbuterol has been substantially increased. The objective of this study was to determine the ,detection times' for clenbuterol after administration of an oral clinical dose (0.8 g/kg, b.i.d.) of Ventipulmin syrup. Five horses received oral clenbuterol (0.8 g/kg, b.i.d.) for 10 days, and urine concentrations of clenbuterol were determined by an enhanced enzyme-linked immunoabsorbent assay (ELISA) test and gas chromatography/mass spectrometric (GC/MS) analysis by two different methods for 30 days after administration. Twenty-four hours after the last administration, urine concentrations of apparent clenbuterol, as measured by ELISA, averaged about 500 ng/mL, dropping to about 1 ng/mL by day 5 posttreatment. However, there was a later transient increase in the mean concentrations of apparent clenbuterol in urine, peaking at 7 ng/mL on day 10 postadministration. The urine samples were also analysed using mass spectral quantification of both the trimethylsilyl (TMS) and methane boronic acid (MBA) derivatives of clenbuterol. Analysis using the TMS method showed that, at 24 h after the last administration, the mean concentration of recovered clenbuterol was about 22 ng/mL. Thereafter, clenbuterol concentrations fell below the limit of detection of the TMS-method by day 5 after administration but became transiently detectable again at day 10, with a mean concentration of about 1 ng/mL. Derivatization with MBA offers significant advantages over TMS for the mass spectral detection of clenbuterol, primarily because MBA derivatization yields a high molecular weight base peak of 243 m/z, which is ideal for quantitative purposes. Therefore, mass spectral analyses of selected urine samples, including the transient peak on day 10, were repeated using MBA derivatization, and comparable results were obtained. The results show that clenbuterol was undetectable in horse urine by day 5 after administration. However, an unexpected secondary peak of clenbuterol was observed at day 10 after administration that averaged ,1 ng/mL. Because of this secondary peak, the detection time for clenbuterol (0.8 g/kg, b.i.d. × 10 days) is at least 11 days if the threshold for detection is set at 1 ng/mL. [source] Derivatization for liquid chromatography/electrospray mass spectrometry: synthesis of tris(trimethoxyphenyl)phosphonium compounds and their derivatives of amine and carboxylic acidsRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 5 2002William J Leavens A simple method for the derivatization of primary amines and carboxylic acids with tris(trimethoxyphenyl)phosphonium (TMPP) reagents to enhance their detection by electrospray mass spectrometry (ESI-MS) has been developed. The synthesis of novel TMPP reagents and their stable isotopically labelled analogues is described. Through the use of stable isotopically labelled TMPP ,tags', incorporation of a doublet (1:1, 1H/2H or 12C/13C) into the target molecule can be achieved, enabling the use of isotopic target analysis to detect compounds of unknown molecular weight but with a characteristic isotope pattern and accurate mass difference. Copyright © 2002 John Wiley & Sons, Ltd. [source] ChemInform Abstract: The Use of Formamidine Protection for the Derivatization of Aminobenzoic Acids.CHEMINFORM, Issue 15 2009Paul E. Zhichkin Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a "Full Text" option. The original article is trackable via the "References" option. [source] Novel One-Pot Synthesis of 5-Alkenyl-15-alkynylporphyrins and Their Derivatization to a Butadiyne-Linked Benzoporphyrin Dimer.CHEMINFORM, Issue 20 2006Hiroko Yamada Abstract ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 200 leading journals. To access a ChemInform Abstract, please click on HTML or PDF. [source] Reaction of Halothane with sec-Butyllithium in the Presence of Zinc Halides , One-Pot Preparation of Chlorodifluorovinylzinc Reagent and Its Derivatization to ,-Chloro-,,,-difluorostyrene.CHEMINFORM, Issue 48 2003Masakazu Nishihara Abstract For Abstract see ChemInform Abstract in Full Text. [source] New Polyalkynyl Dendrons and Dendrimers: "Click" Chemistry with Azidomethylferrocene and Specific Anion and Cation Electrochemical Sensing Properties of the 1,2,3-Triazole-Containing DendrimersCHEMISTRY - A EUROPEAN JOURNAL, Issue 12 2009Jérémy Camponovo Abstract Dendrimers for ion sensing: The synthesis and use of new tris-alkynyl dendrons are reported. So-called "click" reactions of the dendrimers described with azidomethylferrocene give 27-ferrocenyl, 81-ferrocenyl, and 243-ferrocenyl dendrimers. Electrochemical recognition of oxo-anions and Pd2+ cations has been compared using the three polyferrocenyl dendrimers. The synthesis and use of the new tris-alkynyl dendrons 2 to 5 are reported, including the Williamson reaction of 5 with 9-iodo (9), 27-iodo (11), and 81-iodo (12) dendritic cores to yield 27-alkynyl (13), 81-alkynyl (14), and 243-alkynyl (15) dendrimers. So-called "click" reactions of these three dendrimers with azidomethylferrocene (20) give 27-ferrocenyl (16), 81-ferrocenyl (17), and 243-ferrocenyl (18) dendrimers. Electrochemical recognition of oxo-anions (H2PO4, and ATP2,) and Pd2+ cation has been compared using the three polyferrocenyl dendrimers. Derivatization of Pt electrodes with the dendrimers for recognition becomes more facile with increasing size of the dendrimer. This first "click" dendrimer bearing 243-ferrocenyl groups is the best one in the series to obtain robust, recyclable modified Pt electrodes, whereas previous "click" ferrocenyl dendrimers have not been suitable for this purpose. Nous reportons ici la synthèse et l'utilisation de nouveaux dendrons tris-alcynes (composés 2 à 5). La réaction de Williamson entre 5 et les c,urs dendritiques polyiodés comportant 9, 27 ou 81 branches (composés 9, 11 et 12) conduit aux dendrimères poly-alcynes à 27, 81 et 243 branches respectivement (composés 13 à 15). La réaction "click" de ces dendrimères avec l'azidométhylferrocène (20) permet d'obtenir des dendrimères polyferrocéniques à 27, 81 et 243 branches (composés 16 à 18). La reconnaissance électrochimique d'oxo-anions (H2PO4,et ATP2,) et du cation Pd2+est comparée avec trois dendrimères polyferrocéniques, et l'obtention d'électrodes de Pt modifiées à l'aide de ces dendrimères pour cette reconnaissance est de plus en plus facile lorsque la taille du dendrimère augmente. Le premier dendrimère "click" comportant 243 ferrocènes est le meilleur de la série pour la modification d'électrodes de Pt. Ces électrodes sont robustes et recyclables avec ce dendrimère, alors que les dendrimères "click" précédemment publiés n'étaient pas utilisables pour cette fonction. [source] Selenium Derivatization of Nucleic Acids for X-Ray Crystal-Structure and Function StudiesCHEMISTRY & BIODIVERSITY, Issue 4 2010Jia Sheng Abstract It is estimated that over two thirds of all new crystal structures of proteins are determined via the protein selenium derivatization (selenomethionine (Se-Met) strategy). This selenium derivatization strategy via MAD (multi-wavelength anomalous dispersion) phasing has revolutionized protein X-ray crystallography. Through our pioneer research, similarly, Se has also been successfully incorporated into nucleic acids to facilitate the X-ray crystal-structure and function studies of nucleic acids. Currently, Se has been stably introduced into nucleic acids by replacing nucleotide O-atom at the positions 2,, 4,, 5,, and in nucleobases and non-bridging phosphates. The Se derivatization of nucleic acids can be achieved through solid-phase chemical synthesis and enzymatic methods, and the Se-derivatized nucleic acids (SeNA) can be easily purified by HPLC, FPLC, and gel electrophoresis to obtain high purity. It has also been demonstrated that the Se derivatization of nucleic acids facilitates the phase determination via MAD phasing without significant perturbation. A growing number of structures of DNAs, RNAs, and protein,nucleic acid complexes have been determined by the Se derivatization and MAD phasing. Furthermore, it was observed that the Se derivatization can facilitate crystallization, especially when it is introduced to the 2,-position. In addition, this novel derivatization strategy has many advantages over the conventional halogen derivatization, such as more choices of the modification sites via the atom-specific substitution of the nucleotide O-atom, better stability under X-ray radiation, and structure isomorphism. Therefore, our Se-derivatization strategy has great potentials to provide rational solutions for both phase determination and high-quality crystal growth in nucleic-acid crystallography. Moreover, the Se derivatization generates the nucleic acids with many new properties and creates a new paradigm of nucleic acids. This review summarizes the recent developments of the atomic site-specific Se derivatization of nucleic acids for structure determination and function study. Several applications of this Se-derivatization strategy in nucleic acid and protein research are also described in this review. [source] Quantitation of valproic acid in pharmaceutical preparations using dispersive liquid-liquid microextraction followed by gas chromatography-flame ionization detection without prior derivatizationDRUG TESTING AND ANALYSIS, Issue 7 2010Hamid Reza Sobhi Abstract Dispersive liquid-liquid microextraction (DLLME), coupled with gas chromatography-flame ionization detection (GC-FID), has been successfully used for the extraction and determination of valproic acid (VPA) in pharmaceutical preparations. In the developed method, an appropriate mixture of extracting and disperser solvents was rapidly injected into an aqueous sample. Having formed a cloudy solution, the mixture was centrifuged and then the extracting solvent was sedimented at the bottom of a conical test tube. The extract was then injected into a GC system directly, without any further pretreatment. Initially, microextraction efficiency factors were optimized and the optimum experimental conditions found were as follows: tetrachloroethylene (9.0 µL) as extracting solvent; acetone (1.0 mL) as disperser solvent; 5 mL acidic aqueous sample (pH 1) without salt addition. Under the selected conditions, the calibration curve showed linearity in the range of 0.1,5.0 mg/L with regression coefficient corresponding to 0.9998. The limit of detection was found to be 0.05 mg/L. Finally, the method was applied for the determination of VPA in two different pharmaceutical preparations. A reasonable intra-assay (3.9,10.8%, n = 3) and inter-assay (5.6,11.4%, n = 3) precision illustrated the good performance of the analytical procedure. The protocol proved to be rapid and cost-effective for screening purposes. Copyright © 2010 John Wiley & Sons, Ltd. [source] Preparation, Electrochemistry, and Electrocatalytic Activity of Lead Pentacyanonitrosylferrate Film Immobilized on Carbon Ceramic ElectrodeELECTROANALYSIS, Issue 21 2008H. Razmi Abstract Lead pentacyanonitrosylferrate (PbPCNF), a new Prussian blue analog, was immobilized on the surface of a carbon ceramic electrode (CCE) prepared by sol-gel method. The immobilization process consists of adding a certain amount of metallic lead to the electrode matrix before gelation, and chemical derivatization of Pb on the electrode surface to a PbPCNF solid film by immersing the electrode in a solution of sodium pentacyanonitrosylferrate (PCNF). The composition of the synthesized PbPCNF was characterized by FTIR, scanning electron microscopy (SEM), and energy-dispersive X-ray (EDX) techniques. The resulting modified electrode showed electroactivity at two redox centers. The electrochemical behavior of the PbPCNF modified carbon ceramic electrode (PbPCNF|CCE) was studied by cyclic voltammetry. Under optimized conditions the peak-to-peak separation is only 39,mV, indicative of a surface reaction. Ion effects of the supporting electrolyte suggest that cations have a considerable effect on the electrochemical behavior of the modified electrode. The transfer coefficient (,) and the charge transfer rate constant at the modifying film|electrode interface (ks) were calculated. The electrocatalytic activity of the modified electrode toward the electro-reduction of peroxodisulfate was studied in details. [source] Selective Analysis of Secondary Amines Using Liquid Chromatography with Electrochemical Detection (LC-EC)ELECTROANALYSIS, Issue 21 2006Celia Abstract In a mixture of primary and secondary aliphatic amines, the primary amines were derivatized (masked) with o -phthalaldehyde (OPA) followed by derivatization of the remaining secondary amines with ferrocenecarboxylic acid chloride (FAC). The "tagged" amines were analyzed by LC-EC (liquid chromatography with electrochemical detection) using in-series dual electrode detection. Chemically-reversible oxidation of the FAC tagged secondary amines and their subsequent complementary oxidation and reduction signals coupled with chemically-irreversible oxidation of OPA tagged primary amines provided the selectivity for quantitative secondary amine analysis. The procedure was also applied for the selective identification of fragment 4,11 (N -terminus-proline) of Substance P in the presence of other Substance P fragments with primary amino acids as their N -termini. [source] Voltammetric Determination of Free and Total Sulfur Dioxide in BeerELECTROANALYSIS, Issue 5-6 2003J. Almeida Abstract A voltammetric method for the determination of free and total sulfur dioxide in beer is described. First, volatile aldehydes (mainly acetaldehyde) are purged with nitrogen from a beer sample diluted in alkaline medium, collected in an appropriate electrolyte trapping solution and determined, after derivatization with hydrazine, by voltammetry using a hanging mercury drop electrode. Then, the remaining beer solution is strongly acidified and (total) sulfur dioxide is purged with nitrogen, collected in an appropriate electrolyte trapping solution and determined by voltammetry. The free sulfur dioxide concentration is calculated by difference between (total) sulfur dioxide and acetaldehyde concentrations. The proposed method has a relative standard deviation of about 2.1% and 4.4%, respectively for (total) sulfur dioxide and free sulfur dioxide concentrations normally found in beer, and results are in good agreement with those obtained by the p -rosaniline reference method. [source] Recent developments and applications of EMMA in enzymatic and derivatization reactionsELECTROPHORESIS, Issue 1 2010Jie Zhang Abstract This review covers the time period of 2007 until mid-2009 and describes new developments in the field of electrophoretically mediated microanalysis. The review is subdivided in two parts dealing with (i) enzymatic and (ii) derivatization or chemical reactions. A compilation of the relevant literature is given for each part. [source] Cover Picture: Electrophoresis 21'2009ELECTROPHORESIS, Issue 21 2009Article first published online: 27 OCT 200 Issue no. 21 is a regular issue with Emphasis on "Nucleic Acids". The first part has 7 articles on nucleic acids covering various topics, e.g., sequencing, genotyping, PCR, insertion, mutation, etc. The remaining 11 articles are concerned with monoliths, pseudo-phases, coating, and sample pretreatment such as derivatization and concentration. Selected articles are: Applications of MALDI-TOF MS to large-scale human mtDNA population-based studies ((10.1002/elps.200900294)) Visual DNA as a diagnostic tool ((10.1002/elps.200900273)) Preliminary results for two-dimensional separation with high performance thin-layer chromatography and pressurized planar electrochromatography ((10.1002/elps.200900471)) [source] A novel approach to tag and identify geranylgeranylated proteinsELECTROPHORESIS, Issue 20 2009Lai N. Chan Abstract A recently developed proteomic strategy, the "GG-azide"-labeling approach, is described for the detection and proteomic analysis of geranylgeranylated proteins. This approach involves metabolic incorporation of a synthetic azido-geranylgeranyl analog and chemoselective derivatization of azido-geranylgeranyl-modified proteins by the "click" chemistry, using a tetramethylrhodamine-alkyne. The resulting conjugated proteins can be separated by 1-D or 2-D and pH fractionation, and detected by fluorescence imaging. This method is compatible with downstream LC-MS/MS analysis. Proteomic analysis of conjugated proteins by this approach identified several known geranylgeranylated proteins as well as Rap2c, a novel member of the Ras family. Furthermore, prenylation of progerin in mouse embryonic fibroblast cells was examined using this approach, demonstrating that this strategy can be used to study prenylation of specific proteins. The "GG-azide"-labeling approach provides a new tool for the detection and proteomic analysis of geranylgeranylated proteins, and it can readily be extended to other post-translational modifications. [source] Fast derivatization of the non-protein amino acid ornithine with FITC using an ultrasound probe prior to enantiomeric determination in food supplements by EKCELECTROPHORESIS, Issue 6 2009Elena Domínguez-Vega Abstract An EKC method for the determination of ornithine (Orn) enantiomers has been developed after a fast pre-capillary derivatization with FITC. The derivatization step was needed to provide a chemical moiety to the Orn molecule, enabling a sensitive UV detection and the interaction with the CDs used as chiral selectors. To accelerate the derivatization reaction, an ultrasound probe was used. For the development of the chiral method, the influence of different experimental conditions (type and concentration of the chiral selector, temperature, and separation voltage) was investigated. Due to the anionic nature of the analyte (FITC-Orn), five neutral CDs were employed as chiral selectors. The native ,-CD showed the highest chiral separation power, observing that a low concentration of this CD (1,mM), using a working temperature of 25°C and a separation voltage of 20,kV, enabled to obtain the highest enantioresolution for Orn and its separation from other amino acids usually present in food supplements. After optimizing the method for the preconditioning of the capillary, the analytical characteristics of the chiral method were established. Linearity, LOD and LOQ, precision, and accuracy were evaluated previously to the determination of Orn enantiomers contained in ten commercial food supplements. No interferences from other amino acids present in these samples were observed. [source] Arsenic speciation in natural and contaminated waters using CZE with in situ derivatization by molybdate and direct UV-detectionELECTROPHORESIS, Issue 6 2009Olga S. Koshcheeva Abstract A simple and sensitive procedure for simultaneous determination of arsenate, arsenite, monomethylarsonate and dimethylarsinate (DMA) ions in waters using CZE with chemical derivatization in situ and UV-detection at 250,nm was developed. The separation was performed in a fused-silica capillary using solution containing sodium molybdate and sodium perchlorate as electrolyte. Molybdate forms heteropolycomplexes with arsenic species in low acidic media, while sodium perchlorate masks silicate ion. The analysis conditions were optimized; the best results were achieved with the electrolyte consisting of 10,mM Na2MoO4 and 10,mM NaClO4 at pH 3.0 using negative voltage and pneumatic injection of the sample. Nevertheless, the signal of arsenite ion was not detected, probably because of its instability. Arsenite ion was quantified as a difference between arsenate ion contents after and before oxidation by bromine water. The detection limits for the fresh water at the level of 5.0,,g/L for AsIII and AsV, 16,,g/L for DMA and 20,,g/L for MMA were achieved. The reproducibility varied in the range of 0.06,0.25 relative units. To reduce the interferences of the sample salinity an addition of organic substances and isotachophoretic effect were used. [source] Improved workup for glycosaminoglycan disaccharide analysis using CE with LIF detectionELECTROPHORESIS, Issue 22 2008Alicia M. Hitchcock Abstract This work describes improved workup and instrumental conditions to enable robust, sensitive glycosaminoglycan (GAG) disaccharide analysis from complex biological samples. In the process of applying CE with LIF to GAG disaccharide analysis in biological samples, we have made improvements to existing methods. These include (i) optimization of reductive amination conditions, (ii) improvement in sensitivity through the use of a cellulose cleanup procedure for the derivatization, and (iii) optimization of separation conditions for robustness and reproducibility. The improved method enables analysis of disaccharide quantities as low as 1,pmol prior to derivatization. Biological GAG samples were exhaustively digested using lyase enzymes, the disaccharide products and standards were derivatized with the fluorophore 2-aminoacridone and subjected to reversed polarity CE-LIF detection. These conditions resolved all known chondroitin sulfate (CS) disaccharides or 11 of 12 standard heparin/heparan sulfate disaccharides, using 50,mM phosphate buffer, pH 3.5, and reversed polarity at 30,kV with 0.3,psi pressure. Relative standard deviation in migration times of CS ranged from 0.1 to 2.0% over 60 days, and the relative standard deviations of peak areas were less than 3.2%, suggesting that the method is reproducible and precise. The CS disaccharide compositions are similar to those obtained by our group using tandem MS. The reversed polarity CE-LIF disaccharide analysis protocol yields baseline resolution and quantification of heparin/heparan sulfate and CS/dermatan sulfate disaccharides from both standard preparations and biologically relevant proteoglycan samples. The improved CE-LIF method enables disaccharide quantification of biologically relevant proteoglycans from small samples of intact tissue. [source] Comparative metabolite profiling of carboxylic acids in rat urine by CE-ESI MS/MS through positively pre-charged and 2H-coded derivatizationELECTROPHORESIS, Issue 22 2008Wen-Chu Yang Abstract A new approach to the selective comparative metabolite profiling of carboxylic acids in rat urine was established using CE-MS and a method for positively pre-charged and 2H-coded derivatization. Novel derivatizing reagents, N -alkyl-4-aminomethyl-pyridinum iodide (alkyl=butyl, butyl-d9 or hexyl), containing quaternary amine and stable-isotope atoms (deuterium), were introduced for the derivatization of carboxylic acids. CE separation in positive polarity showed high reproducibility (0.99,1.32% RSD of migration time) and eliminated problems with capillary coating known in CE-MS anion analyses. Essentially complete ionization and increased hydrophobicity after the derivatization also enhanced MS detection sensitivity (e.g. formic acid was detected at 0.5,pg). Simultaneous derivatization of one sample using two structurally similar reagents, N -butyl-4-aminomethyl-pyridinum iodide (BAMP) and N -hexyl-4-aminomethyl-pyridinum iodide, provided additional information for recognizing a carboxylic acid in an unknown sample. Moreover, characteristic fragmentation acquired by online CE-MS/MS allowed for identification and categorization of carboxylic acids. Applying this method on rat urine, we found 59 ions matching the characteristic patterns of carboxylic acids. From these 59, 32 ions were positively identified and confirmed with standards. For comparative analysis, 24 standard carboxylic acids were derivatized by chemically identical but isotopically distinct BAMP and N -butyl-d9-4-aminomethyl-pyridinium iodide, and their derivatization limits and linearity ranges were determined. Comparative analysis was also performed on two individual urine samples derivatized with BAMP and N -butyl-d9-4-aminomethyl-pyridinium iodide. The metabolite profiling variation between these two samples was clearly visualized. [source] Determination of gaseous and particulate carbonyls in air by gradient-elution micellar electrokinetic capillary chromatographyELECTROPHORESIS, Issue 19 2008Hui Sun Abstract A new continuous-flow gradient-elution micellar electrokinetic capillary chromatography method is developed for the determination of airborne carbonyls after derivatization with 2,4-dinitrophenylhydrazine. A total of 16 carbonyls can be determined with detection limits ranging from 0.94 to 8.50,mg/L, working range from 4.72 to 346,mg/L, and repeatabilities (relative standard deviation, n=5) from 1.23 to 4.6% or 3.93 to 7.6% for migration time and peak area, respectively. Coupling with denuder-filter sampling, a preliminary survey has been conducted to determine gaseous and particulate carbonyls from air sampled at a roadside station. The method is shown to have sufficient sensitivity for 1-h sampling of ambient carbonyls with detection limits ranging from 0.045 to 1.2,,g/m3 and working range from 0.11 to 43.3,,g/m3 at a flow rate of 10,Lpm. The method requires minimal modification of commercially available capillary electrophoresis equipment and can differentiate gaseous and particulate carbonyls to provide essential information and objective data for adopting effective measures to combat the discharge of carbonyl compounds to the atmosphere. [source] Assessment of protein-incorporated arginine methylation in biological specimens by CZE UV-detectionELECTROPHORESIS, Issue 23 2007Angelo Zinellu Dr. Abstract Protein arginine methyltransferases methylate post-translationally arginine residues in proteins to synthesize monomethylarginine (MMA), asymmetric dimethylarginine (ADMA), or symmetric dimethylarginine. Protein arginine methylation is involved in the regulation of signal transduction, RNA export, and cell proliferation. Moreover, upon proteolysis, arginines are released into the cytosol in which they exert important biological effects. Both MMA and ADMA are inhibitors of nitric oxide synthase and especially elevated levels of ADMA are associated with endothelial dysfunction and cardiovascular disease. Quantification of these analytes is commonly performed by HPLC after sample cleanup and derivatization. We propose a CE method in which these steps have been avoided and the procedure for sample preparation has been simplified. After acidic hydrolysis of proteins, samples were dried, resuspended in water, and directly injected in CE. A baseline separation of analytes was reached in a 60 cm×75,,m id uncoated silica capillary, by using a Tris-phosphate run buffer at pH,2.15. This method allows an accurate assessment of protein arginine methylation degree in different biological samples such as whole blood, plasma, red blood cells, cultured cells, and tissue. Moreover, its good sensitivity permits to evaluate the methylation of a single protein type after the opportune purification steps. A method applicability concerns both clinical laboratories, where the evaluation of blood protein from numerous samples could be rapidly performed, and research laboratories where the factors affecting the arginine protein methylation degree could be easily studied. [source] Analysis of dideoxyadenosine triphosphate by CE with fluorescence detection.ELECTROPHORESIS, Issue 21 2007Abstract A CZE method was developed, which separates 2,,3,-dideoxyadenosine-5,-triphosphate (ddATP) from other metabolites and endogenous nucleotides at high concentrations (20,200,,g/mL) to allow UV detection. To enhance sensitivity, fluorescence detection which requires prior derivatization of compounds was examined. Precapillary derivatization of ddATP in the presence of N -(3-dimethylaminopropyl)- N,-ethylcarbodiimide hydrochloride (EDAC) with dansyl ethylenediamine (dansyl EDA) was faster and stable compared to that of 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza- s -indacene-3-propionyl ethylenediamine (BODIPY FL EDA). Reaction conditions, reagent concentrations and detection parameters were optimized and highest derivatization efficiency was achieved in 0.1,M 1-methylimidazole buffer (pH,8.0) with 140,mM EDAC in 1-methylimidazole buffer and 30,mM dansyl EDA in DMF for 90,min at 60°C. Dansyl EDA derivatives of ddATP, 2,-deoxyadenosine-5,-triphosphate (dATP) and ATP were comigrating with the CZE method; therefore, a MEKC method was developed and optimized for repeatable separations. Upon dansylation, sensitivity of ddATP with fluorescence detection (LOQ,=,12,ng/mL) was 160 times higher than UV detection (LOQ,=,1.9,,g/mL). [source] Analysis of sub-ppb levels of Fe(II), Co(II), and Ni(II) by electrokinetic supercharging preconcentration, CZE separation, and in-capillary derivatizationELECTROPHORESIS, Issue 20 2007Marek Urbanek Abstract The analysis of sub-ppb levels of Fe(II), Co(II), and Ni(II) in heat exchanger fluids of nuclear power plants is needed to monitor corrosion. A method involving preconcentration with electrokinetic supercharging (electrokinetic injection with transient ITP), CZE separation, and in-capillary derivatization with ortho -phenanthroline (o -Phe) for direct UV detection was thus developed. First, a multizone BGE was loaded into the capillary by successive hydrodynamic introduction of zones of (i) o -Phe-containing BGE, (ii) BGE for the zonal separation, and (iii) ammonium-based leading electrolyte. Metal cations were electrokinetically injected and stacked at the capillary inlet behind this last leading zone. Finally, a terminating electrolyte zone was hydrodynamically introduced. When a constant voltage was applied, metal ions kept on concentrating isotachophoretically, then separated in CZE mode, were complexed by migrating through an o -Phe zone, and finally detected by direct absorbance. To detect extremely thin peaks, it was attempted for the first time to focus the derivatization reagent by inducing a second transient ITP, before labeling analytes, already separated in CZE mode. With this arrangement, LODs were about 30,ppt in pure water. In heat exchanger fluid matrices containing 1000,ppm bore and 2,ppm lithium, only Fe(II) cation was detected among the three cations of interest at the 1,ppb level using the present method, and its LOD was about ten times higher, due to the lower loading rate during electrokinetic injection. [source] Amino acid profiling in plant cell cultures: An inter-laboratory comparison of CE-MS and GC-MSELECTROPHORESIS, Issue 9 2007Brad J. Williams Abstract A CE-MS method for metabolic profiling of amino acids was developed and used in an integrated functional genomics project to study the response of Medicago truncatula liquid suspension cell cultures to stress. This project required the analysis of more than 500 root cell culture extracts. The CE-MS method profiled 20 biologically important amino acids. The CE-MS method required no sample derivatization prior to injection and used minimal sample preparation. The method is described in terms of CE and MS operational parameters, reproducibility of migration times and response ratios, sample preparation, sample throughput, and reliability. This method was then compared with a previously published report that used GC-MS metabolic profiling for the same tissues. The data reveal a high level of similarity between the CE-MS and GC-MS amino acid profiling methods, thus supporting these as complementary technologies for metabolomics. We conclude that CE-MS is a valid alternative to GC-MS for targeted profiling of metabolites, such as amino acids, and possesses some significant advantages over GC-MS. [source] Atmospheric molding of ionic copolymer MALDI-TOF/MS arrays: A new tool for protein identification/profilingELECTROPHORESIS, Issue 24 2006Alexander Muck Abstract An atmospheric molding protocol has been used to prepare an ionic methacrylate-based copolymer sample support chips for MALDI (pMALDI)-MS by targeting selected groups of various monomers copolymerized during molding, namely, carboxy, sulfo, dimethylalkyamino, and trimethylalkylammonium groups. The new disposable array chips provide analyte-oriented enhancement of protein adsorption to the modified substrates without requiring complicated surface coating or derivatization. The MALDI-MS performance of the new ionic copolymer chips was evaluated for lysozyme, ,-lactoglobulin,A, trypsinogen and carbonic anhydrase,I using washing with solutions prepared in pH or ionic strength steps. On cationic chips, the proteins are washed out at pH lower than their pI values, and on anionic chips at pH higher than their pI values. The ability of the microfabricated pMALDI chip set to selectively adsorb different proteins from real samples and to significantly increase their MS-signal was documented for the transmembrane photosystem,I protein complex from the green alga Chlamydomonas reinhardtii. The proteins were almost exclusively adsorbed according to calculated pI values and grand average of hydropathy (GRAVY) indexes. The new disposable chips reduce manipulation times and increase measurement sensitivity for real-world proteomic samples. The simple atmospheric molding procedure enables additional proteomic operations to be incorporated on disposable MALDI-MS integrated platforms. [source] A robust cross-linked polyacrylamide coating for microchip electrophoresis of dsDNA fragmentsELECTROPHORESIS, Issue 19 2006Joann J. Lu Abstract Surface derivatization plays an important role in microchip electrophoresis. It not only enhances the resolution, but also improves the reproducibility. So far, the most popularly used derivatization method for glass microchannels is to covalently attach a layer of linear polyacrylamide,(LPA) to the channel surfaces. However, LPA coating has two problems: incomplete coverage and limited lifetime. To address these issues, we have recently developed a cross-linked polyacrylamide,(CPA) derivatization protocol and demonstrated it for high-resolution protein separations by CIEF, CGE, and CZE. In this report, we used this protocol to coat microchip channels and exhibited the reliability and robustness of CPA coating for microchip electrophoresis of DNA molecules. dsDNA fragments were used as our test samples. High resolutions were obtained for fragments ranging from 100,bp to 10,kpb. After more than 800,runs, the CPA-coated microchannels still performed well and comparable resolutions were maintained throughout these runs. [source] Intracellular FITC-derivatization with PEGELECTROPHORESIS, Issue 21 2005Fanguo Chen Abstract In order to investigate the amino acids (AAs) in plant cells, we explore an avenue for intracellular derivatization with FITC. In this method, FITC was used to mark AAs in living protoplasts derived from embryogenic calli of common wheat (Triticum aestivum L.,c.v. Jinan,177) mediated by PEG. After FITC-derivatization, the AAs in the lysate were determined by CE. The result reveals that this PEG method can be used to transfer FITC into plant cells efficiently, which provides a good method for AA,analysis in plant cells. [source] On-line sample preconcentration with chemical derivatization of bacterial biomarkers by capillary electrophoresis: A dual strategy for integrating sample pretreatment with chemical analysisELECTROPHORESIS, Issue 21 2005Adam S. Ptolemy Abstract Simple, selective yet sensitive methods to quantify low-abundance bacterial biomarkers derived from complex samples are required in clinical, biological, and environmental applications. In this report, a new strategy to integrate sample pretreatment with chemical analysis is investigated using on-line preconcentration with chemical derivatization by CE and UV detection. Single-step enantioselective analysis of muramic acid (MA) and diaminopimelic acid (DAP) was achieved by CE via sample enrichment by dynamic pH junction with ortho -phthalaldehyde/N -acetyl- L -cysteine labeling directly in-capillary. The optimized method resulted in up to a 100-fold enhancement in concentration sensitivity compared to conventional off-line derivatization procedures. The method was also applied toward the detection of micromolar levels of MA and DAP excreted in the extracellular medium of Escherichia coli bacterial cell cultures. On-line preconcentration with chemical derivatization by CE represents a unique approach for conducting rapid, sensitive, and high-throughput analyses of other classes of amino acid and amino sugar metabolites with reduced sample handling, where the capillary functions simultaneously as a concentrator, microreactor, and chiral selector. [source] | ||||||||||||||||||||||||||||||||||||||||