bluuebunny/crossref_metadata_2025_split huggingface dataset:
The effect of cyclodextrin modifiers on electrophoretic separation of aromatic hydrocarbons
Authors: Nguyen An‐Lac , Luong John H. T. 
Published: May 2005

AbstractCyclodextrin‐modified capillary electrophoresis has been developed for separation and analysis of benzene and its derivatives. The procedure used a mixture of negatively charged sulfobutyl ether‐β‐cyclodextrin (SBβCD) and neutral hydroxypropyl‐β‐cyclodextrin (HPβCD) to effect differential distribution (partitioning) of the aromatic hydrocarbons between the buffer and CD phases. In 80 mM phosphate buffer, containing 15 mM SBβCD and 5 mM HPβCD, benzene, toluene, ethyl benzene and three xylene isomers (BTEXs) were well resolved with a number of theoretical plates well above 100 000, for 50 cm of effective length. Some halogenated benzenes were also observed to separate well from the BTEX components to indicate their suitability as an internal standard for BTEX analyses. Equilibrium complexation models were used for investigating the effect of the cyclodextrin(s) at different concentrations and temperature on the electrophoretic mobility.
Analysis of brevetoxins by micellar electrokinetic capillary chromatography and laser‐induced fluorescence detection
Authors: Shea Damian 
Published: May 2005

AbstractMicellar electrokinetic capillary chromatography (MEKC) with laser‐induced fluorescence (LIF) detection was used to measure four red tide brevetoxins at sub‐attomole levels. The separation of four brevetoxins by MEKC was achieved with a sodium borate/sodium dodecyl sulfate buffer at pH 9.3. Brevetoxins with a terminal alcohol group were derivatized with an acyl azide coumarin to form stable, highly fluorescent products. Brevetoxins with a terminal aldehyde group were reduced to the alcohol with sodium borohydride prior to derivatization with the coumarin. Three derivatized brevetoxins (PbTx‐3, PbTx‐5, and Pb Tx‐9) were separated by MEKC and detected using He/Cd laser excitation at 354 nm and fluorescence emission at 410 nm. A fourth brevetoxin (Pb Tx‐2) was converted to Pb Tx‐3 prior to derivatization and was then determined by subtraction. Instrumental detection limits for all four toxins were approximately 0.10 fg or about 106‐fold more sensitive than existing liquid chromatographic methods. Brevetoxins were isolated from cell cultures and fish tissue using an alumina column/gel‐permeation chromatography procedure. Method detection limits for the brevetoxins in fish tissue were approximately 4 pg/g. These method detection limits are at least 100‐fold better than previous chromatographic and/or electrophoretic methods. The MEKC‐LIF method reported here allows measurement of brevetoxins at the trace levels considered critical for understanding toxin metabolism and mode of action.
Analysis of toxic drugs by capillary electrophoresis using polyacrylamide‐coated columns
Authors: Jinno Kiyokatsu , Han Yinghong , Sawada Hirokazu 
Published: May 2005

AbstractToxic drugs, including barbiturates and benzodiazepines, were analyzed using polyacrylamide‐coated columns in capillary electrophoresis (CE). The separations were carried out in absence of electroosmotic flow. Seven kinds of barbiturates were successfully separated with the coated columns in free solution without further additives. Benzodiazepines, the electrically neutral solutes, were introduced onto the coated column, and separated in presence of sodium dodecyl sulfate above its critical micelle concentration in the running buffer. This CE method offered fast and efficient separation of more hydrophobic solutes, such as benzodiazepines. The separation of seven barbiturates was studied in linear (noncross‐linked) polyacrylamide solutions and in acrylamide / N‐isopropylacrylamide (AA/IPAA) copolymer solutions to explore the effect of isopropyl groups in the AA/IPAA copolymer chain.
Improvement of the solubilization of proteins in two‐dimensional electrophoresis with immobilized pH gradients
Authors: Rabilloud Thierry , Adessi Céline , Giraudel Anne , Lunardi Joël 
Published: May 2005

AbstractMembrane and nuclear proteins of poor solubility have been separated by high resolution two‐dimensional (2‐D) gel electrophoresis. Isoelectric focusing with immobilized pH gradients leads to severe quantitative losses of proteins in the resulting 2‐D map, although the resolution is usually high. Protein solubility could be improved by using denaturing solutions containing various detergents and chaotropes. Best results were obtained with a denaturing solution containing urea, thiourea, and detergents (both nonionic and zwitterionic). The usefulness of thiourea‐containing denaturing mixtures is shown for microsomal and nuclear proteins as well as for tubulin, a protein highly prone to aggregation.
Sensitivity and mass accuracy for proteins analyzed directly from polyacrylamide gels: Implications for proteome mapping
Authors: Loo Rachel R. Ogorzalek , Mitchell Charles , Stevenson Tracy I. , Martin Stephen A. , Hines Wade M. , Juhasz Peter , Patterson Dale H. , Peltier John M. , Loo Joseph A. , Andrews Philip C. 
Published: May 2005

AbstractMatrix‐assisted laser desorption ionization (MALDI) mass spectra have been obtained directly from thin‐layer isoelectric focusing (IEF) gels with as little as 700 femtomoles of α‐ and β‐chain bovine hemoglobin and bovine carbonic anhydrase, and 2 picomoles of bovine trypsinogen, soybean trypsin inhibitor, and bovine serum albumin all loaded onto a single lane. By soaking the gel in a matrix solution, matrix was deposited over the entire gel surface, allowing MALDI scanning down complete lanes of the one‐dimensional gel. As long as matrix crystals were deposited finely on the surface of the gel, time‐lag focusing techniques were capable of ameliorating some of the mass accuracy limitations inherent in desorbing from uneven insulator surfaces with external calibration. Eleven measurements on the 5 kDa α‐subunit proteins of lentil lectin measured over the course of 1 h and referenced to a single calibration yielded a standard deviation of 0.025%. Colloidal gold staining was found to be compatible with desorption directly from IEF and sodium dodecyl sulfate (SDS)‐polyacrylamide gels. This direct approach simplifies the interface between gel electrophoresis and mass spectrometry dramatically, making the process more amenable to automation.
A comparison of selected mRNA and protein abundances in human liver
Authors: Anderson Leigh , Seilhamer Jeff 
Published: May 2005

AbstractIn order to obtain an estimate of the overall level of correlation between mRNA and protein abundances for a well‐characterized pharmaceutically relevant biological system, we have analyzed human liver by quantitative two‐dimensional electrophoresis (for protein abundances) and by Transcript Image methodology (for mRNA abundances). Incyte's LifeSeq™ database was searched for expressed sequence tag (EST) sequences corresponding to a series of 23 proteins identified on 2‐D maps in the Large Scale Biology (LSB) Molecular Anatomy™ database, resulting in estimated abundances for 19 messages (4 were undetected) among 7926 liver clones sequenced. A correlation coefficient of 0.48 was obtained between the mRNA and protein abundances determined by the two approaches, suggesting that post‐transcriptional regulation of gene expression is a frequent phenomenon in higher organisms. A comparison with published data (Kawamoto, S., et al., Gene 1996, 174, 151–158) on the abundances of liver mRNAs for plasma proteins (secreted by the liver) suggests that higher abundance messages are strongly enriched in secreted sequences. Our data confirms this: of the 50 most abundant liver mRNAs, 29 coded for secreted proteins, while none of the 50 most abundant proteins appeared to be secreted products (although four plasma and red blood cell proteins were presents in this group as contaminants from tissue blood).
Characterisation of wool intermediate filament proteins separated by micropreparative two‐dimensional electrophoresis
Authors: Herbert Ben R. , Molloy Mark P. , Yan Jun X. , Gooley Andrew A. , Bryson Warren G. , Williams Keith L. 
Published: May 2005

AbstractWool intermediate filament proteins (IFP) are a subclass of the cytokeratins, a group of structural proteins which form intermediate filaments in many cell types. Post‐translational modifications, such as phosphorylation, play an important role in the control of intermediate filament assembly. Two‐dimensional electrophoresis has previously been used to study the IFP distribution in wools with different physical characteristics. Charge heterogeneity has been observed in Type I and Type II IFP. In a previous study, two‐dimensional electrophoresis of alkaline phosphatase‐treated wool protein extracts was used to show that Type II IFP are phosphorylated. To facilitate post‐separation analysis, micropreparative two‐dimensional electrophoresis was used to separate milligram quantities of wool protein. Direct phosphoamino acid analysis has confirmed the presence of phosphorylation on serine residues on Type II IFP, whose identity was confirmed by amino acid compositional analysis. The isoelectric points of Type I IFP are very similar and they do not separate completely on the commercially available pH 4–7 immobilized pH gradients (IPG) used in this study. In situ tryptic digestion followed by automated Edman sequencing of the high performance liquid chromatography (HPLC)‐separated peptides was used to confirm the identity of this group as Type I IFP. To improve the separation of the Type I IFP it will be necessary to use narrow range IPGs such as Immobiline DryPlates which are available from Pharmacia Biotech, in the pH ranges 4.2–4.9, 4.5–5.4, 5.0–6.0 and 5.6–6.6.
Protein expression changes associated with radiation‐induced neoplastic progression of human prostate epithelial cells
Authors: Prasad Sarada C. , Thraves Peter J. , Dritschilo Anatoly , Kuettel Michael R. 
Published: May 2005

AbstractCarcinogenic progression in most epithelial systems is a multistep process and presents as numerous (un)stable intermediate stages prior to the development of a fully malignant phenotype. Recently, we reported the neoplastic transformation of an SV40 immortalized, neonatal human prostate epithelial cell line (267B1) by multiple exposures to X‐rays [1, 2]. The parental 267B1 cells acquired anchorage‐independence and exhibited morphological transformation following exposure to two consecutive doses of 2 Gy. Exposure of either the parental 267B1 cells or the anchorage‐independent derivatives (F3‐SAC) to a total dose of 30 Gy of X‐rays yielded tumorigenic transformants (267B1‐XR and 267B1‐SXR, respectively). All of these radiation‐treated derivatives (F3‐SAC, 267B1‐XR, and 267B1‐SXR) were characterized by reduced cell size and poorly organized actin stress fibers [2, 3]. The present study examines the protein expression changes associated with cytoskeletal alterations during the different steps of neoplastic progression induced by X‐rays in the in vitro human prostate cell system. This analysis was achieved by using the high resolving power of two‐dimensional polyacrylamide gel electrophoresis (2‐D PAGE) in the 267B1, F3‐SAC, 267B1‐XR, and 267B1‐SXR cells. We report changes in the expression of gelsolin in the partially transformed, anchorage‐independent, nontumorigenic (F3‐SAC) cells and a progressive loss of expression of tropomyosin isoforms (TM‐1 and TM‐3), and myosin light chain‐2 (MLC‐2) in the tumorigenic (267B1‐XR; 267B1‐SXR) cells, respectively. In contrast, our results demonstrate that the levels of the small GTP‐binding protein Rho‐A, an active participant in the actin stress fiber organization, are not altered during neoplastic progression of these 267B1 cells. Thus the changes in synthesis of gelsolin, tropomyosins, and MLC‐2 provide a rationale for the alterations in the actin stress fiber formation and reduction in cell size during the exposure of prostate epithelial cells to multiple doses of X‐rays.
Capillary electrophoretic separation of tricyclic antidepressants using charged carboxymethyl‐β‐cyclodextrin as a buffer additive
Authors: Spencer Brian J. , Zhang Wenbin , Purdy William C. 
Published: May 2005

AbstractCharged carboxymethyl‐β‐cyclodextrin was successful in the capillary electrophoretic separation of a series of tricyclic antidepressants. The cyclodextrin alone was successful in the separation of carbamazepine, protriptyline, desipramine, clomipramine, and opipramol using a 3‐(trimethoxysilyl)propyl methacrylate capillary coating to reduce the electroosmotic flow. The ideal buffer pH was found to be in the range of 6–7 and the ideal cyclodextrin concentration to be 10 mM. All nine antidepressants were resolved using the charged cyclodextrin in the micellar electrokinetic chromatography (MEKC) mode with sodium dodecyl sulfate as the surfactant. Neither the cyclodextrin nor the surfactant alone were successful in resolving the whole series of compounds under investigation but a combination of both produced the separation. Separations were performed on a linear polyacrylamide coated capillary. The ideal pH of the buffer was in the range of 5–7.
Isolation of subcellular‐sized particles separated by electrophoresis in dilute polymer solution, using commercial electrophoresis apparatus with intermittent scanning of fluorescence
Authors: Chen Nong , Chrambach Andreas 
Published: May 2005

AbstractResolution of subcellular‐sized particles in electrophoresis employing semidilute polymer solutions as “sieving media” improves as the polymer concentration is decreased. Therefore, the previously reported conditions of preparative electrophoresis of microsomes, using concentrated (12%) polyvinylpyrrolidone (PVP) solutions, while solving the problem of non‐entrance of large particles into “sieving media”, do not provide adequate resolving capacity, as exemplified by failure of the microsome preparation used, to resolve in the manner of gels or dilute solutions. The present report provides the conditions under which the HPGE‐1000 apparatus can be preparatively applied when the electrophoretic separation is effectively conducted in a dilute polymer solution. The isolation of three microsome components under those conditions constitutes the first application of “particle sieving”, i.e., a separation due preponderantly to size and shape differences, at a preparative scale.
Alternative method of subtyping the A subunit of coagulation factor XIII (FXIIIA)
Authors: Kido Akira , Oya Masakazu , Jin Zeming 
Published: May 2005

AbstractAn alternative isoelectric focusing method for the A subunit of coagulation factor XIII (FXIIIA) subtyping is described which employs three different carrier ampholytes pH 4–6.5, pH 5–6 and pH 5–8 (2:2:1), and sample application at the anode. With this technique ten common subtypes, including a variant, were clearly and reliably identified in Japanese and Chinese population samples.
Dilated cardiomyopathy‐associated proteins and their presentation in a WWW‐accessible two‐dimensional gel protein database
Authors: Plei Klaus‐Peter , Söding Peter , Sander Stefan , Oswald Helmut , Neuß Michael , Regitz‐Zagrosek Vera , Fleck Eckart 
Published: May 2005

AbstractHigh resolution two‐dimensional electrophoresis (2‐DE) and computer‐assisted image analysis were used to screen 13 patients suffering from dilated cardiomyopathy (DCM) versus 15 control patients for quantitative and qualitative differences in their myocardial protein expression. Right atrial tissue samples were obtained from end‐stage failing explanted hearts and control hearts. Fifty‐two spots differed significantly in average intensity between the DCM and the control groups. Myosin light chain 2, ventricular (MLC2) and heat shock protein HSP 27 were identified by protein microsequencing and gel map comparison with other databases. These proteins were found to be characteristic protein markers for DCM in the right atrium. In DCM patients, the spot intensity (protein abundance) of MLC2 is increased to 336% and HSP 27 is decreased to 59%, compared to the control group. The HEART‐2DPAGE, a World Wide Web‐accessible 2‐DE database, was used and extended for the presentation of these disease‐associated proteins. Retrievable via Internet we present a list of disease‐associated proteins, their altered level of expression in DCM, and their position on a right atrial protein pattern. The accession number to protein sequence databases confers a connection to databases like SWISS‐PROT to obtain a detailed functional and structural description of disease‐associated proteins. New DCM‐associated proteins are detected and their presentation in a 2‐DE gel protein database is described.
High resolution two‐dimensional electrophoretic analysis of urinary proteins of patients with prostatic cancer
Authors: Grover Phulwinder K. , Resnick Martin I. 
Published: May 2005

AbstractIn an attempt to identify marker(s) for prostatic cancer, proteins in urines of normal and prostatectomized males and in men with cancerous prostate were analyzed. Only urines collected with protease inhibitors were examined. Two‐dimensional (2‐D) gel electrophoresis was used for high resolution separation of proteins and the electrophoretograms were either developed by double stain or the proteins were electrophoretically transferred onto nitrocellulose for immunological identification. The pool of each group exhibited similar relative positions of major protein spots. The study of normal and prostatectomized men identified two proteins denoted as A (36 kDa, pI 6–6.5) and B (23 kDa, pI 6.6), which were undetectable in the latter group. A visual comparison of the patterns of normals and patients with cancerous prostate revealed that both these proteins were undetectable in urines of men with malignant prostatic carcinoma (PCA) and benign prostatic hyperplasia (BPH) and hence may be useful in identifying prostatic carcinoma. Also, while protein C 43.5 kDA, pI 6–6.6) was discerned in normals its abundance, along with those of proteins D (40 kDa, pI 6–6.4) and E (26.5 kDa, pI 6–6.7), appeared to be higher in BPA than in PCA. Protein F (18–28 kDa, pI 4–5.5) was found in patients with BPH but was undetected in normals and men with PCA. Hence, it may become useful in distinguishing BPH from PCA. All the proteins, A to F, appear to be previously unidentified. Their further characterization is warranted.
Analysis of proteins from different phase variants of the entomopathogenic bacteria Photorhabdus luminescens by two‐dimensional zymography
Authors: Ong Kevin L. , Chang Frank N. 
Published: May 2005

AbstractTwo‐dimensional zymography which combines two‐dimensional electrophoresis with zymography was used to analyze proteases and other proteins produced by different phase variants of two strains of Photorhabdus luminescens. Both the primary and secondary phases of P. luminescens strains Hp and Hm secreted proteases. The protease in P. luminescens Hp has a molecular weight (Mr) of 57 000 and an isoelectric point (pI) of 4.4 whereas that in P. luminescens Hm has an Mr of 59 000 and pI of 4.9. Several putative protease degradation products were clearly visible in the zymograms from both bacterial strains. Two‐dimensional zymography also showed that several secretory proteins were present only in particular phase variants and therefore could be used as specific markers. Unexpectedly, the two‐dimensional zymography revealed that a nonsecretory protease with an Mr of 47 000 and a pI of 4.0 was present in the cell extracts of all phases of both P. luminescens Hp and Hm. The application of the two‐dimensional zymography for the identification of other enzymes was also discussed.
Use of a zwitterionic cyclodextrin as a chiral agent for the separation of enantiomers by capillary electrophoresis
Authors: Lelièvre Frédéric , Gueit Carole , Gareil Pierre , Bahaddi Youssef , Galons Hervé 
Published: May 2005

AbstractThe purity and enantioselectivity of a novel chiral agent, the zwitterionic mono‐(6‐δ‐glutamylamino‐6‐deoxy)‐β‐cyclodextrin (β‐CD‐Glu), were studied by capillary electrophoresis. Chiral separation of the enantiomers of chlorthalidone was obtained at pH 2.3, a pH at which β‐CD‐Glu is partially protonated. Comparison with the cationic mono‐(6‐amino‐6‐deoxy)‐β‐cyclodextrin (β‐CD‐NH2) enantioselectivity clearly shows that the greater the difference in mobility between the free analyte and the analyte‐cyclodextrin complex, the better the resolution. Hydrobenzoin enantiomers were separated at pH 11.2, a pH at which β‐CD‐Glu is anionic. Under these conditions, the migration order was opposite to that observed in the presence of β‐CD‐NH2 at pH 2.3. When no separation was obtained directly with β‐CD‐Glu, a dual cyclodextrin system was developed. Carprofen enantiomers were resolved at pH 2.3 in the presence of a β‐CD‐Glu/trimethyl‐β‐cyclodextrin (TM‐β‐CD) system in which the charged CD confers a non‐zero mobility to the analyte, while the neutral CD allows chiral recognition.
Chiral separation of DL‐peptides and enantioselective interactions between teicoplanin and D‐peptides in capillary electrophoresis
Authors: Wan Hong , Blomberg Lars G. 
Published: May 2005

AbstractTeicoplanin has been evaluated as a selector for enantioseparation of di‐ and tripeptide derivatives in capillary electrophoresis. Separation variables such as type of buffer, pH, concentrations of teicoplanin and organic modifier were examined. Optimal separation conditions were obtained by means of factorial design experiments. The effects of teicoplanin concentrations below and above its critical micellar concentration (CMC) and of acetonitrile (ACN) on the separation were demonstrated. The use of a high concentration of ACN resulted not only in increased selectivity, but also in improved separation efficiency. Electroosmotic flow was observed to be largely independent of the concentrations of teicoplanin under the optimized conditions. Good repeatability of migration times was obtained. The interactions between teicoplanin and D and L peptides were studied, and it was found that, for some peptides, teicoplanin exhibited enantioselective interaction only with the D‐form. Somewhat lower separation efficiencies were thus observed for the strongest interacting (D‐form) peptides. Chiral separation of 15 DL‐peptide derivatives was achieved in less than 10 min.
A quantitative relationship between capacity factor and selector concentration in capillary electrophoresis and high performance liquid chromatography: Evidence from the enantioselective resolution of benzoin using human serum albumin as a chiral selector
Authors: Lloyd David K. , Ahmed Abdelwab , Pastoré Florent 
Published: May 2005

AbstractMany selectors are used both in pressure‐driven liquid chromatography (LC) and in electrokinetic chromatography (EKC), particularly chiral species such as cyclodextrins and proteins. It should be possible to readily apply information gleaned using one technique to the other, since in both techniques the underlying molecular interactions which lead to separations are expected to be the same. Superficially this may be the case, but an exact transfer of operating conditions, i.e., background electrolyte (BGE) composition/mobile phase composition, assuming that these meet certain minimum requirements for each technique, is not often possible. To investigate the reason for this we have measured retention (K′) of a neutral solute (racemic benzoin) in HPLC and EKC using an identical range of BGE/mobile phase conditions in both techniques. The selector used was human serum albumin. The K′ measurements obtained for each benzoin enantiomer were consistently higher in HPLC than in EKC. This can be explained very simply if one considers that retention in both systems is related to the selector concentration [S], by the expression K′=K[S], where K is the affinity constant. In EKC, [S] is simply the concentration of free selector in the BGE, while in LC, [S] = mp/Vo, where mp is the number of moles of accessible selector, and Vo is the column void volume. In LC, [S] is generally considerably higher than in EKC, leading to larger values of K′.