bluuebunny/crossref_metadata_2025_split huggingface dataset:
Synthesis of zwitterionic acrylic acid buffers for isoelectric focusing in immobilized pH gradients
Authors: Bellini Marco P. , Manchester Keith L. 
Published: May 2005

AbstractA range of zwitterionic acrylic acid derivatives, buffering in the neutral and basic pH ranges, have been synthesized by the Mannich reaction of malonic acid, formaldehyde and a secondary amine. These compounds include 2‐(4‐morpholinomethyl)propenoic acid pK2 7.59 ± 0.03 (23°C), 2‐[bis(2‐hy‐droxyethyl)aminomethyl]propenoic acid pK2 ≈ 8.6 (20°C), 2‐[bis(2‐hydroxy‐propyl) aminomethyl]propenoic acid pK2 ≈ 8.7 (20°C), 2‐[N‐(2‐hydroxyethyl)‐N‐methylaminomethyl]propenoic acid pK2 9.22 ± 0.08 (22°C), 2‐[N‐ethyl‐N‐(2‐hydroxyethyl)aminomethyl]‐propenoic acid pK2 ≈ 9.6 (20°C), and 2‐[4‐(2‐carboxyprop‐2‐enyl)piperazinylmethyl]propenoic acid, which has a sigmoidal buffering profile over the pH range 3–10. These zwitterionic acrylic acid buffers were successfully copolymerized with acrylamide to prepare immobilized pH gradients (IPGs) in the neutral to alkaline portion of the pH range. Bovine erythrocyte carbonic anhydrase isozymes were resolved on a pH 5–8 IPG prepared using 2‐[4‐(2‐carboxyprop‐2‐enyl)piperazinylmethyl]propenoic acid as the immobilized buffer and horse heart myoglobin was focused on pH 7.1–8.1 and pH 7.5–7.7 IPGs, using 2‐(4‐morpholinomethyl)propenoic acid as the immobilized buffer. In both cases the pK 9.3 Immobiline compound was used as the strongly basic titrant. These new compounds, besides possessing more hydrophilic residues than the corresponding commercial basic acrylamido buffers (Immobilines), resist hydrolysis at alkaline pH values.
Determination of plasmid‐encoded functions in Rhizobium leguminosarum biovar trifolii using proteome analysis of plasmid‐cured derivatives
Authors: Guerreiro Nelson , Stepkowski Tomasz , Rolfe Barry G. , Djordjevic Michael A. 
Published: May 2005

AbstractWe have used proteome analysis of derivatives of R. leguminosarum biovar trifolii strain ANU843, cured of indigenous plasmids by a direct selection system, to investigate plasmid‐encoded functions. Under the conditions used, the plasmid‐encoded gene products contributed to only a small proportion of the 2000 proteins visualised in the two‐dimensional (2‐D) protein map of strain ANU843. The level of synthesis of thirty‐nine proteins was affected after curing of either plasmid a, c or e. The differences observed upon plasmid curing included: protein loss, up/down‐regulation of specific proteins and novel synthesis of some proteins. This suggests that a complex interplay between the cured plasmid and the remaining replicons is occurring. Twenty‐two proteins appeared to be absent in the cured strains and these presumably are encoded by plasmid genes. Of these, a small heat shock protein, a cold shock protein, a hypothetical YTFG‐29.7 kDa protein, and the alpha and beta sub‐units of the electron transfer flavoprotein were identified by N‐terminal micro‐sequencing and predicted to be encoded by plasmid e. Four of the sequenced proteins putatively encoded on plasmid e and two encoded on plasmid c were novel. In addition, curing of plasmid e and c consistently decreased the levels of 3‐isopropylmalate dehydratase and malate dehydrogenase, respectively, suggesting that levels of these proteins may be influenced by plasmid‐encoded functions. A protein with homology to 4‐oxalocrotonate tautomerase, which is involved in the biodegradation of phenolic compounds, was found to be newly synthesised in the strain cured of plasmid e. Proteome analysis provides a sensitive tool to examine the functional organisation of the Rhizobium genome and the global gene interactions which occur between the different replicons.
Cardiac protein abnormalities in dilated cardiomyopathy detected by two‐dimensional polyacrylamide gel electrophoresis
Authors: Corbett Joseph M. , Why Howard J. , Wheeler Colin H. , Richardson Peter J. , Archard Leonard C. , Yacoub Magdi H. , Dunn Michael J. 
Published: May 2005

AbstractThe aim of the investigation was to determine whether there are specific global quantitative and qualitative changes in protein expression in heart tissue from patients with dilated cardiomyopathy (DCM) compared with ischaemic heart disease and undiseased tissue. Two‐dimensional (2‐D) polyacrylamide gel electrophoresis and computer analysis was used to study protein alteration in DCM biopsy material (n=28) compared with donor heart biopsy samples (n=9) and explanted hearts from individuals suffering from ischaemic heart disease (IHD; n = 21). A total of 88 proteins displayed decreased abundance in DCM versus IHD material while five proteins had elevated levels in the DCM group (p<0.01). The most prominent changes occurred in the contractile protein myosin light chain 2 and in a group of proteins identified as desmin. These changes do not appear to be artefactual degradation events occurring during sample processing. These proteins are not apparent in electrophoretic separations of vascular tissue or cultured endothelial cells, mesothelial cells or cardiac fibroblasts, which are clearly distinguishable from the 2‐D protein patterns of whole heart and of isolated cardiac myocytes and do not appear to reflect variations in the cellular composition of biopsy samples. The different protein patterns observed in cardiomyopathy showed no obvious relationship with New York Heart Association (NYHA) functional class or haemodynamic parameters. The study has demonstrated significant alterations in quantitative protein expression in the DCM heart which would have serious implications for myocyte function. These changes might be explained by altered protease activity in DCM which could exacerbate contractile dysfunction in the failing heart.
Capillary electrophoresis, nuclear magnetic resonance and mass spectrometry studies of opposite chiral recognition of chlorpheniramine enantiomers with various cyclodextrins
Authors: Chankvetadze Bezhan , Burjanadze Naira , Bergenthal Dieter , Strickmann Dirk , Blaschke Gottfried , Pintore Giorgio , Cerri Riccardo 
Published: May 2005

AbstractMarkedly different chiral separation abilities were observed for native β‐cyclodextrin (β‐CD), carboxymethyl‐β‐CD (CM‐β‐CD) and heptakis (2,3,6‐tri‐O‐methyl)‐β‐CD (TM‐β‐CD) towards the enantiomers of (±)‐chlorpheniramine ((±)‐CHL) in capillary electrophoresis (CE). Native β‐CD afforded almost baseline enantioseparation at a concentration of 18 mg/mL, whereas only 1 mg/mL solution of CM‐β‐CD was required for adequate enantioseparation. TM‐β‐CD allowed the nearly baseline enantioseparation only at a concentration as high as 80 mg/mL. Moreover, the migration order of (±)‐CHL in the presence of TM‐β‐CD was opposite to that with β‐CD and CM‐β‐CD. 1H and 13C‐NMR spectroscopy and electrospray ionization mass spectrometry (ESI‐MS) have been used in order to obtain preliminary information about the stoichiometry and the binding constants in the intermolecular diastereomeric complexes of (±)‐CHL with these CDs.
Capillary electrophoresis of anions at high salt concentrations
Authors: Ding Weiliang , Thornton Michelle J. , Fritz James S. 
Published: May 2005

AbstractIt is commonly thought that even a moderately high ionic concentration in the background electrolyte (BGE) would leaad to Joule heating and serious peak distortion. However, we obtained very satisfactory separations of both inorganic and organic anions in electrolyte solutions as high as 5 M sodium chloride using direct photometric detection. Samples containing a 0.5 M concentration of a salt can be analyzed directly by making the BGE concentration of the same salt even higher to obtain electrostacking. The temperature in the center of the capillary was calculated to be 49°C when the current is at its maximum of 280 μA. The effect of various salts on electrophoretic and electroosmotic mobility is discussed. Several examples are given of capillary electrophoresis under high‐salt conditions.
Characterization of micelles by capillary electrophoresis
Authors: Schwarz Maria A. , Raith Klaus , Neubert Reinhard H. H. 
Published: May 2005

AbstractA practical approach for the characterization of pure micelles, and binary or ternary mixed micelles by capillary electrophoretic methods is presented. Interest is focused on the determination of mobility and composition of the micelles. The investigations are performed with bile salts, phosphatidylcho‐lines and fatty acids. Pure bile salt micelles are characterized with the help of marking and displacement methods. Binary bile salt/phospholipid and ternary bile salt/phospholipid/fatty acid micelles are analyzed using capillary electrophoresis (CE) techniques with standard and improved UV detection, laser‐induced fluorescence and electrospray ionization ‐ mass spectrometry (ESI‐MS). For both the binary and the ternary systems, only one stable mixed micellar phase is found with a high negative mobility.
On‐line capillary electrophoresis‐electrospray ionization mass spectrometry using a polymerized anionic surfactant
Authors: Lu Wenzhe , Shamsi Shahab A. , McCarley Tracy D. , Warner Isiah M. 
Published: May 2005

AbstractOn‐line capillary electrophoresis ‐ electrospray ionization ‐ mass spectrometry (CE‐ESI‐MS) has been used to determine the tricyclic antidepressant drugs (imipramine, doxepin, and amitriptyline) as well as the β‐adrenergic blocker drugs (propranolol and alprenolol). A CE‐ESI‐MS interface linking a manually operated CE system and a Finnigan MAT‐900 sector mass spectrometer (with an Analytica electrospray ionization source) has been constructed in‐house and employed for this study. Although a water/methanol based capillary zone electrophoresis (CZE) buffer was initially used to determine these analytes, enhanced resolution was obtained by addition of a polymerized surfactant, i.e., poly‐sodium undecylenic sulfate (poly‐SUS), into the electrokinetic chromatography (EKC) buffer. When a low concentration of this poly‐SUS surfactant was added to a volatile EKC buffer, these structurally similar cationic drugs were EKC separated and on‐line detected by ESI‐MS.
Capillary electrophoresis and capillary electrophoresis ‐ electrospray ‐ mass spectroscopy for the analysis of heterocyclic amines
Authors: Zhao Yining , Schelfaut Marc , Sandra Pat , Banks Fred 
Published: May 2005

AbstractFourteen heterocyclic amines (HAs) were analyzed by capillary electrophoresis (CE) on a polyvinyl alcohol (PVA)‐coated capillary column. The optimized electrolyte is composed of 20 mM ammonium acetate, pH 3.0, and 20% methanol. Similar conditions were applied in electrospray ‐ mass spectrometry (CE‐ES‐MS). The CE‐ES‐MS optimization procedure includes the position of the capillary in the stainless steel interface, the flow of the sheath liquid and its composition.
Simultaneous measurement of α‐amylase and glucoamylase activities in sake rice koji by capillary electrophoresis of sodium dodecyl sulfate‐protein complexes and activity measurement of glucoamylase by in‐capillary enzyme reaction method
Authors: Watanabe Toshiro , Yamamoto Akira , Nagai Shiro , Terabe Shigeru 
Published: May 2005

AbstractCapillary electrophoresis (CE) of sodium dodecyl sulfate (SDS)‐protein complexes using a nongel sieving matrix (CE‐SDS) has been applied to the simultaneous analysis of α‐amylase and glucoamylase activity in sake rice koji which is employed for the brewing of sake. α‐Amylase and glucoamylase in sake rice koji extracts were successfully analyzed by CE‐SDS. α‐Amylase and glucoamylase were found to have molecular masses of 53 000 and 63 000 Da, respectively, as determined by the migration times of eight standard proteins. These values agree with those determined by SDS‐polyacrylamide gel electrophoresis (PAGE). The results of CE‐SDS method were compared with those achieved by the official method. The relative standard deviations (RSD) of the α‐amylase and glucoamylase activities by CE‐SDS were less than 5.0% in both intra‐day and inter‐day experiments. An electrophoretic analysis of products of an enzyme reaction of a substrate by in‐capillary reaction was also useful for the activity measurement of glucoamylase in sake rice koji. p‐Nitrophenyl‐β‐D‐maltoside (PNP‐Mal) was employed as a substrate and p‐nitrophenyl‐β‐D‐glucopyranoside (PNP‐Glu) was the product of the enzyme reaction. The glucoamylase activity of sake rice koji samples gave the good linear relationship with the peak area observed in the in‐capillary enzyme reaction method. The glucoamylase activity in sake rice koji was measured by either CE‐SDS or the in‐capillary enzyme reaction more easily than by the official method. Both methods can be applied to the routine quality control of α‐amylase and glucoamylase activities in sake rice koji.
High resolution analysis of protein phosphorylation using capillary isoelectric focusing ‐ electrospray ionization ‐ mass spectrometry
Authors: Wei Jing , Yang Liyu , Harrata A. Kamel , Lee Cheng S. 
Published: May 2005

AbstractOn‐line capillary isoelectric focusing (CIEF)‐electrospray ionization ‐ mass spectrometry (ESI‐MS) as a two‐dimensional separation system is employed for high resolution analysis of ovalbumin phosphorylation. On the basis of their differences in isoelectric point (pI), the mono‐ and diphosphoovalbumins are separated and resolved in CIEF. The focused protein zones of mono‐ and diphosphoovalbumins are eluted by combining gravity with cathodic mobilization. At the end of the CIEF capillary, the mobilized ovalbumin zones are analyzed by mass spectrometry coupled on‐line to an electrospray interface with a coaxial sheath flow configuration. Additional ovalbumin variants within each of the mono‐ and diphosphoovalbumins, differing in their molecular masses due to glycosylation microheterogeneity, are easily distinguished by ESI‐MS.
Genotyping metabolic polymorphisms in a cohort of Caucasians and single strand conformation polymorphism analysis of point mutations in human hprt exons 7 and 8
Authors: Krause Günter , Garganta Fátima , Kosytorz Patrick , Scherer Gerhard 
Published: May 2005

AbstractIn genetic toxicology, the main fields of applications of the polymerase chain reaction (PCR) with subsequent electrophoretic characterization of amplificates include genotyping polymorphisms in the xenobiotic metabolism and mutant analysis. To assess the role of the individual sets of biotransformation enzymes for the internal dose resulting from xenobiotic exposure, we investigated blood samples from 69 healthy donors for the occurrence of known genetic polymorphisms in the xenobiotic metabolizing enzymes N‐acetylaminotransfrase II (NAT2), glutathione‐S‐transferase (GST) μ and θ, and several cytochromes P450 (CYP), namely CYP1A1, CYP2E1 and CYP2A6. Using single strand conformation polymorphism (SSCP) analysis, five known single base substitutions located in the middle portion of 144 bp amplificates comprising exons 7 and 8 of the human hypoxanthine guanine phosphoribosyl transferase (hprt) cDNA, were clearly distinguished from wild type and from each other. Biomagnetic strand separation assigned the slower migrating single strand bands to the biotinylated sense strands.
Separation of inorganic and organic anions by capillary zone electrophoresis with simultaneous indirect UV and conductivity detection
Authors: Klampfl Christian W. , Katzmayr Martin U. , Buchberger Wolfgang 
Published: May 2005

AbstractThe analysis of low‐molecular‐mass anionic compounds using capillary zone electrophoresis (CZE) with simultaneous direct conductivity and indirect UV detection is described. A number of carrier electrolyte compositions has been investigated with respect to the crucial parameters for their compatibility with both detection principles discussed in this paper (UV absorptivity and electrophoretic mobility of the buffer coion). Additionally, parameters including pH and buffer concentration affecting both peak symmetry and signal‐to‐noise ratio obtained for the analytes of interest had to be optimized. Best results could be achieved with a carrier electrolyte consisting of 4‐aminobenzoic acid, bis(2‐hydroxyethyl)imino‐tris(hydroxymethyl)aminomethane (BIS‐Tris) and a pH of 7.0 adjusted with LiOH. This running buffer was used for the analysis of low‐molecular‐mass anionic compounds in a sample of white wine.
Off‐line quantitative monitoring of plasmid copy number in bacterial fermentation by capillary electrophoresis
Authors: Breuer Susanne , Marzban Gorji , Cserjan‐Puschman Monika , Dürrschmid Eberhard , Bayer Karl 
Published: May 2005

AbstractCapillary electrophoresis (CE) is an effective instrumental alternative to conventional slab gel electrophoresis in the determination of plasmid copy number during recombinant protein formation processes. This analytical setup provides efficient separation of different species of linearized plasmid molecules and quantification by UV detection. Both fused silica and gel‐filled capillaries are assessed with respect to peak resolution and reproducibility. The application of coated capillaries eliminates the electroosmatic flow to a large extent, resulting in excellent separation of DNA fragments. The application of UV detection enables the analysis of linearized plasmid DNA with a conventional laboratory CE device. All investigated plasmids show good peak resolution due to their significant differences in molecular size, which is essential for sufficient separation of individual DNA molecules.
Differential expression of cytosolic proteins in the rat kidney cortex and medulla: Preliminary proteomics
Authors: Witzmann Frank A. , Fultz Carla D. , Grant Raymond A. , Wright Lynda S. , Kornguth Steven E. , Siegel Frank L. 
Published: May 2005

AbstractThe rodent kidney is a target of many xenobiotics and is typified by regionally specific structure and function. This renders distinct regions of the kidney differentially susceptible to toxic exposure and effect. To characterize these differences at the proteome level, protein patterns from male rat kidney cortex and medulla cytosols were examined by two‐dimensional electrophoresis (2‐DE) and image analysis and prominent proteins identified immunologically or by matrix‐assisted laser desorption/ionization ‐ mass spectrometry (MALDI‐MS) and electrospray/ionization ‐ tandem mass spectrometry (ESI‐MS/MS) sequence tag identification. An average of 727 protein spots were resolved and matched to the cortex cytosol reference pattern, and 716 in the medulla. Of this total, 127 proteins were found to differ in abundance (86 higher in cortex; 41 higher in medulla) (P < 0.001). Of those proteins that were detectable in both cortex and medulla, the abundance of 97 differed significantly while 30 proteins were found to be unique to one region or the other (26 in cortex, 4 in medulla). Twenty protein spots were identified and their regional differences are discussed. These results both confirm and expand our understanding of the molecular heterogeneity characterizing structurally and functionally distinct regions of the kidney and serve as a useful foundation for future nephrotoxicologic studies.
Identification of minor proteins of human colostrum and mature milk by two‐dimensional electrophoresis
Authors: Murakami Kouki , Lagarde Margaret , Yuki Yoshikazu 
Published: May 2005

AbstractTwo‐dimensional electrophoresis (2‐DE) followed by electroblotting and microsequencing is considered to be the most powerful method for the isolation and characterization of proteins. In this paper, we report the separation and determination of the N‐terminal and/or internal amino acid sequences of the minor proteins of human colostral and mature milk by 2‐DE and microsequencing. In order to analyze the minor proteins of human milk, we use immunoabsorbents to remove three major proteins, α‐lactalbumin, lactoferrin and secretory immunoglobulin A. The major proteins removed by this process accounted for about 79 and 93% of the total whey proteins of mature and colostral milk, respectively. The remaining milk proteins were then separated by isoelectric focusing gel electrophoresis between pH 3 and 10, and subjected to 12.5% sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE). Approximately 400 spots were detected in both colostral and mature milk by silver staining after 2‐DE. Twenty‐two major, well‐resolved proteins (out of 400) were microsequenced (N‐termini as well as internal). These include fatty acid binding protein, β2‐microglobulin, complement C4, clusterin, α1‐antritrypsin, lysozyme C, α‐ and β‐casein, prealbumin, serotransferrin, fructose‐bisphosphate aldolase A, and β‐casein fragments. No major differences in the protein patterns were observed between the minor proteins of colostrum and mature milk, indicating that the minor proteins remained relatively constant during lactation. These results suggest that the minor milk proteins are important for the health and development of breast‐fed infants throughout lactation.
Electrophoretic methods for process monitoring and the quality assessment of recombinant glycoproteins
Authors: Taverna Myriam , Tran Nguyet Thuy , Merry Tony , Horvath Eszter , Ferrier Danielle 
Published: May 2005

AbstractIn many ways electrophoretic techniques appear ideal for quality monitoring of proteins and are thus well suited for the analysis of recombinant glycoproteins. The requirements of high throughput, comparative analysis and resolution of many variants are met by several electrophoretic techniques. A wide variety of such techniques are available to biotechnologists in the rapidly developing area of recombinant glycoproteins. It is the aim of this review to specifically cover recent work which has been applied to the analysis of DNA‐derived glycoproteins, both from a process control standpoint and final product validation. All major areas of electrophoresis including sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE), isoelectric focusing and techniques utilizing capillary electrophoresis are covered, with emphasis on analysis of glycoforms and oligosaccharide profiles of recombinant glycoproteins. As illustration, actual examples rather than standard glycoproteins are given to indicate the potential and limitations which may be encountered. It is anticipated that this review will prove a useful and practical guide to the latest developments by indicating the relevant merits of different methods.
Capillary affinity chromatography and affinity capillary electrophoresis of heparin binding proteins
Authors: Wu Xiaojun , Linhardt Robert J. 
Published: May 2005

AbstractA new approach for separation, capillary affinity chromatography, is introduced for studying the interaction of heparin with antithrombin III and secretory leukocyte proteinase inhibitor. Heparin is covalently immobilized on the surface of an etched capillary through a silane spacer. The proteins are injected into the heparinized capillary, bound to the heparin, washed with buffer, eluted with sodium chloride in the same buffer using a pressure injection mode and eluting protein detected by absorbance. The resulting affinity separation is similar to that obtained from traditional affinity chromatography. The quantity of loaded protein in capillary affinity chromatography is at the nanogram level, offering an improvement over the milligram levels required for standard affinity chromatographic methods.