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Second Dimension (second + dimension)
Selected AbstractsHeart-cutting 2D-CE with on-line preconcentration for the chiral analysis of native amino acidsELECTROPHORESIS, Issue 6 2010Suzanne Anouti Abstract The use of transient moving chemical reaction boundary (tMCRB) was investigated for the on-line preconcentration of native amino acids in heart-cutting 2D-CE with multiple detection points using contactless conductivity detection. The tMCRB focusing was obtained by using ammonium formate (pH 8.56) as sample matrix and acetic acid (pH 2.3) as a BGE in the first dimension of the heart-cutting 2D-CE. Different experimental parameters such as the injected volume and the concentration in ammonium formate were optimized for improving the sensitivity of detection. A stacked fraction from the first dimension was selected, isolated in the capillary, and then separated in the second dimension in the presence of a chiral selector ((+)-(18-crown-6)-2,3,11,12-tetracarboxylic acid). This on-line tMCRB preconcentration coupled with heart-cutting 2D-CE was applied with success to the chiral separation of D,L -phenylalanine, and D,L -threonine in a mixture of 22 native amino acids. The sample mixture was diluted in 0.8,M of ammonium formate, and injected at a concentration of 2.5,,M for each enantiomer with a volume corresponding to 10% of the total capillary volume. An LOD (S/N=3) of 2,,M was determined for L -threonine. [source] Proteome analysis of human liver tumor tissue by two-dimensional gel electrophoresis and matrixassisted laser desorption/ionization-mass spectrometry for identification of disease-related proteinsELECTROPHORESIS, Issue 24 2002Jina Kim Abstract Hepatocellular carcinoma (HCC) is a common malignancy worldwide and is a leading cause of death. To contribute to the development and improvement of molecular markers for diagnostics and prognostics and of therapeutic targets for the disease, we have largely expanded the currently available human liver tissue maps and studied the differential expression of proteins in normal and cancer tissues. Reference two-dimensional electrophoresis (2-DE) maps of human liver tumor tissue include labeled 2-DE images for total homogenate and soluble fraction separated on pH 3,10 gels, and also images for soluble fraction separated on pH 4,7 and pH 6,9 gels for a more detailed map. Proteins were separated in the first dimension by isoelectric focusing on immobilized pH gradient (IPG) strips, and by 7.5,17.5% gradient sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gels in the second dimension. Protein identification was done by peptide mass fingerprinting with delayed extraction-matrix assisted laser desorption/ionization-time of flight-mass spectrometry (DE-MALDI-TOF-MS). In total, 212 protein spots (117 spots in pH 4,7 map and 95 spots in pH 6,9) corresponding to 127 different polypeptide chains were identified. In the next step, we analyzed the differential protein expression of liver tumor samples, to find out candidates for liver cancer-associated proteins. Matched pairs of tissues from 11 liver cancer patients were analyzed for their 2-DE profiles. Protein expression was comparatively analyzed by use of image analysis software. Proteins whose expression levels were different by more than three-fold in at least 30% (four) of the patients were further analyzed. Numbers of protein spots overexpressed or underexpressed in tumor tissues as compared with nontumorous regions were 9 and 28, respectively. Among these 37 spots, 1 overexpressed and 15 underexpressed spots, corresponding to 11 proteins, were identified. The physiological significance of the differential expressions is discussed. [source] The development of an emotion model based on colour combinationsINTERNATIONAL JOURNAL OF CONSUMER STUDIES, Issue 2 2006Young-Jin Lee Abstract The purpose of this study was to develop an emotion model based on the colour combinations popularly used for interior coordination in Korea. To summarize, the emotion model had the following features: (1) It consisted of three axes named as ,soft,hard' (first dimension), ,light,heavy' (second dimension) and ,splendid,sober' (third dimension). (2) The emotion descriptors were categorized into nine emotion groups and matched with the representing colour combinations. (3) This emotion model had a one-to-multiplicity correspondence structure between the colour combination and the emotion descriptor, whereas most of the previously developed models included only one-to-one correspondence. (4) It was observed that the emotion variable only showed a relatively consistent tendency within the space of the emotion model as the difference in the tone of colour combinations increased or decreased. The emotion model developed in this study can be used as a basis for the determination of local consumers' emotion on colour combinations to support colour planning in the industrial design field relevant to interior coordination. [source] Immunoisolation of two synaptic vesicle pools from synaptosomes: a proteomics analysisJOURNAL OF NEUROCHEMISTRY, Issue 6 2005Marco Morciano Abstract The nerve terminal proteome governs neurotransmitter release as well as the structural and functional dynamics of the presynaptic compartment. In order to further define specific presynaptic subproteomes we used subcellular fractionation and a monoclonal antibody against the synaptic vesicle protein SV2 for immunoaffinity purification of two major synaptosome-derived synaptic vesicle-containing fractions: one sedimenting at lower and one sedimenting at higher sucrose density. The less dense fraction contains free synaptic vesicles, the denser fraction synaptic vesicles as well as components of the presynaptic membrane compartment. These immunoisolated fractions were analyzed using the cationic benzyldimethyl- n -hexadecylammonium chloride (BAC) polyacrylamide gel system in the first and sodium dodecyl sulfate,polyacrylamide gel electrophoresis in the second dimension. Protein spots were subjected to analysis by matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI TOF MS). We identified 72 proteins in the free vesicle fraction and 81 proteins in the plasma membrane-containing denser fraction. Synaptic vesicles contain a considerably larger number of protein constituents than previously anticipated. The plasma membrane-containing fraction contains synaptic vesicle proteins, components of the presynaptic fusion and retrieval machinery and numerous other proteins potentially involved in regulating the functional and structural dynamics of the nerve terminal. [source] THE CONSUMER SENSORY PERCEPTION OF PASSION-FRUIT JUICE USING FREE-CHOICE PROFILINGJOURNAL OF SENSORY STUDIES, Issue 1 2005ROSIRES DELIZA ABSTRACT Free-choice profiling (FCP) was carried out in order to investigate how naive consumers (who had never tried the product before) described and perceived passion-fruit juice. This method allows participants to use their own attributes to describe and quantify food products and beverages. The study used four different samples of passion-fruit juice, analyzed by 10 consumers in three replicates. The data were analyzed by using generalized Procrustes analysis. The first and second dimension accounted for 78.7% of the variance. The product consensus configuration revealed that assessors were able to reproduce samples' description, and also to differentiate samples. Free-choice profiling is a useful method for describing consumer perception of passion-fruit juice. [source] Orthogonality of silver-ion and non-aqueous reversed-phase HPLC/MS in the analysis of complex natural mixtures of triacylglycerolsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 21 2009Michal Hol Abstract The goal of this work is the study of possibilities of two basic separation modes used in the analysis of complex triacylglycerol (TG) samples of plant oils and animal fats, i.e. non-aqueous reversed-phase (NARP) and silver-ion HPLC coupled with atmospheric pressure chemical ionization mass spectrometry (APCI-MS). The orthogonality of both separation modes is tested for complex TG mixtures containing fatty acids (FAs) with different acyl chain lengths, different number, positions and geometry of double bonds (DBs) and different regioisomeric positions of FAs on the glycerol skeleton. The retention in NARP mode is governed by the equivalent carbon number, while the retention in silver-ion chromatography increases with the increasing number of DBs with a clear differentiation between cis - and trans- FAs. Moreover, silver-ion mode enables at least the partial resolution of regioisomeric TG mixtures including cis -/trans -regioisomers, as illustrated on two examples of randomization mixtures. Off-line 2D coupling of both complementary modes (NARP in the first dimension and silver-ion in the second dimension) yields the superior chromatographic selectivity resulting in the highest number of identified TGs ever reported for studied samples. Off-line 2D chromatograms are processed with the home-made software providing various ways of data visualization. [source] Purification of alkaloids from Corydalis yanhusuo W. T. Wang using preparative 2-D HPLCJOURNAL OF SEPARATION SCIENCE, JSS, Issue 9 2009Jing Zhang Abstract Two-dimensional preparative multi-channel parallel high performance liquid chromatography was successfully applied for the first time to isolate and purify alkaloids from Corydalis yanhusuo. The experiments were performed in off-line mode using the same preparative chromatographic column with pH 3.5 in the first and pH 10.0 in the second separation dimension. In the preparative process, UV-triggered fraction collection was used in the first dimension while UV and MS-triggered collection were used in the second dimension for reasons of sensitivity and complementarity. Two pure compounds and nine fractions were obtained in the first dimension. Then two representative fractions were further purified in the second dimension and six pure compounds were obtained. The results demonstrated that this procedure is an effective approach for the preparative isolation and purification of alkaloids from Corydalis yanhusuo. Based on the different pH values of the mobile phase in this method, it is also suitable for the preparative isolation and purification of other compounds from TCMs which are sensitive to the pH of the solutions. Moreover, this method will be a promising tool for the purification of low content compounds from natural products. [source] Linear peak capacity of a comprehensive multi-dimensional separationJOURNAL OF SEPARATION SCIENCE, JSS, Issue 19 2008Leonid M. Blumberg Abstract In order to resolve (quantifiably and identifiably separate) the same number of peaks in the analysis of the same mixture yielding statistically uniform peak distribution, a comprehensive 2-D separation needs a two times larger peak capacity than a 1-D separation does. Each additional dimension further reduces the utilization of the peak capacity of comprehensive multi-dimensional (MD) separation by a factor of two per dimension. As a result, the same peak capacity means different things for separations with different dimensionalities. This complicates the use of the peak capacity for comparison of the potential separation performance of the separations with different dimensionalities. To facilitate the comparison, a concept of a linear peak capacity has been proposed. The linear peak capacity of an MD separation is the peak capacity of a 1-D separation that, in the analysis of the same mixture, is statistically expected to resolve the same number of peaks as the MD separation is. There are other factors that differently affect the performance of the separations that have different dimensionalities. Peak capacity of a 2-D separation with a rectangular separation space is 27% larger than the product of the peak capacities of its first and second dimension. This advantage of a 2-D separation is essentially nullified by the fact that the peak capacity of the first dimension of an optimized 2-D separation cannot be higher than 80% of the peak capacity of its first dimension standing alone. All in all, the incremental peak capacity gained from addition of a second dimension will not exceed 50% of the peak capacity of the added second dimension. All results are valid for arbitrarily shaped (not necessarily Gaussian) peaks. [source] Characterization of biodiesel and biodiesel blends using comprehensive two-dimensional gas chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 14 2008Warawut Tiyapongpattana Abstract In this work the development of a comprehensive 2-D GC flame ionization detection (GC×GC FID) method for biodiesel fuels is reported. This method is used for the analysis of fatty acid methyl esters (FAMEs) in both biodiesel (B100) and biodiesel blend (B5) samples. The separation of FAME was based on component boiling point in the first dimension and polarity in the second dimension by using a BPX5/BP20 column set to provide a measure of ,orthogonality' in the 2-D space. Here the columns are coupled with a cryogenic modulator operating in a novel temperature programmed mode (TM) whereby the cryotrap is progressively incremented in temperature as the oven temperature is increased. The final method employs eight cryotrap temperature settings. The developed GC×GC method is able to successfully characterize and identify both B100 and B5 FAME components, which are produced from a variety of vegetable oils, animal fats and waste cooking oils, with high precision. The method is capable of analysing FAME with carbon numbers C4,C24, and is particularly suitable to characterize various types of biodiesel, making it possible to differentiate the origin and type of FAME used in the biodiesel samples. [source] The evolution of comprehensive two-dimensional gas chromatography (GC×GC)JOURNAL OF SEPARATION SCIENCE, JSS, Issue 5-6 2004Tadeusz Górecki Abstract For a technology little over a decade old, comprehensive two-dimensional gas chromatography (GC×GC) has quickly reached the status of one of the most powerful analytical tools for volatile organic compounds. At the heart of any GC×GC system is an interface, which physically connects the primary and the secondary columns and acts to preserve the separation obtained in the first dimension (first column) while allowing additional separation in the second dimension. The paper presents a review of the technology, including fundamental principles of the technique, data processing and interpretation and a timeline of inventive contributions to interface design. In addition, applications of the technique are presented, with a more detailed discussion of selected examples. [source] A portable multi-dimensional gas chromatographic system for field applicationsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 12-13 2003Jon H. Wahl Abstract We have constructed and tested a multi-dimensional gas chromatographic system that can be utilized for field portable applications. The chromatographic system is capable of one-dimensional separations and multi-dimensional gas chromatographic (MDGC) separations in a single compact package. Three different general multi-dimensional separation approaches are possible: column switching; traditional heart-cutting; and comprehensive analyses. The MDGC system utilizes a simple 10-port valving approach to accomplish these separations to a single point detector. Because of this valving scheme no hardware change is required to switch between the heart-cut and the comprehensive separation modes, only a software methodology change is required. An additional advantage of this valving approach is that 100% of the first-dimensional effluent is sampled to the second dimension for separation. The system is capable of rapid column heating (room temperature to 250°C in approximately 10 s) and rapid column cooling (250°C to room temperature within approximately 30 s). Preliminary results for heart-cut and comprehensive separations that target five compounds against high concentration levels of complex background are illustrated. [source] Two-Dimensional Chromatography of Complex Polymers, 7 , Detailed Study of Polystyrene- block -Polyisoprene Diblock Copolymers Prepared by Sequential Anionic Polymerization and Coupling ChemistryMACROMOLECULAR CHEMISTRY AND PHYSICS, Issue 19 2008Valentina Mass Abstract Two-dimensional chromatographic methods were developed using LC-CC in the first and SEC in the second dimension. These methods were applied for the investigation of PS- b -PI diblock copolymers synthesized by different approaches: sequential living anionic polymerization and coupling of living precursor blocks. The first dimension separates according to the individual block length of PS or PI blocks, whereas the second dimension separates with respect to the total molar masses of components. 2D-LC analysis provides information on the purity of the reaction products, the presence of by-products, the chemical compositions and the molar masses of all product components. The accuracy and selectivity of 2D-LC is discussed. [source] Fast 1H,13C correlation data for use in automatic structure confirmation of small organic compoundsMAGNETIC RESONANCE IN CHEMISTRY, Issue 2 2005Adrian J. Dunn Abstract A method of speeding up the acquisition of 1H,13C correlation data has been developed. It is applicable in situations where the experiment time is determined by the need to sample the second dimension adequately rather than by signal-to-noise ratio requirements. Two spectra with different, reduced, 13C sweep widths are measured, time being saved by reducing the number of increments in line with the reduction in the sweep width. Rules are presented for the selection of the two reduced sweep widths so that the correct 13C chemical shifts can be easily and unambiguously calculated. The benefits and limitations of this approach, in the context of the structure confirmation of small (MW , 450) organic compounds, is discussed. The use of a third spectrum to resolve problems that may be encountered when proton signals overlap is demonstrated. Copyright © 2004 John Wiley & Sons, Ltd. [source] Dealcrafting: The Substance of Three-Dimensional NegotiationsNEGOTIATION JOURNAL, Issue 1 2002David A. Lax Much of our understanding of negotiation focuses on the process at the table involving a complicated set of interpersonal dynamics and strategies, or a "one-dimensional" approach to the subject. Conceptually independent of one-dimensional process factors is a second dimension of negotiation, "dealcrafting," which focuses on substance in the effort to create joint value. A third dimension of negotiation, involving entrepreneurial moves "away from the table," includes the first two dimensions but offers ways in which negotiators can change the game advantageously. Within this overall 3-D perspective, the second dimension (dealcrafting) calls for a relentless focus on creating maximum value and an equally relentless focus on differences as means to create joint gains. Following their description of the overall 3-D approach, the authors use numerous case examples to illustrate how principles of dealcrafting work in practice. [source] Two-dimensional reference map for the basic proteome of the human dorsolateral prefrontal cortex (dlPFC) of the prefrontal lobe region of the brainPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 13 2010Ciara A. McManus Abstract We describe a 2-DE proteomic reference map containing 227 basic proteins in the dorsolateral prefrontal cortex region of the human brain. Proteins were separated in the first dimension on pH 6,11 IPG strips using paper-bridge loading and on 12% SDS-PAGE in the second dimension. Proteins were subsequently identified by MS and spectra were analyzed using an in-house proteomics data analysis platform, Proline. The 2-DE reference map is available via the UCD 2-DE Proteome Database (http://proteomics-portal.ucd.ie:8082) and can also be accessed via the WORLD-2DPAGE Portal (http://www.expasy.ch/world-2dpage/). The associated protein identification data have been submitted to the PRIDE database (accession numbers 10018,10033). Separation of proteins in the basic region resolves more membrane associated proteins relevant to the synaptic pathology central to many neurological disorders. The 2-DE reference map will aid with further characterisation of neurological disorders such as bipolar and schizophrenia. [source] Purification and identification of a transcription factor, USF-2, binding to E-box element in the promoter of human telomerase reverse transcriptase (hTERT)PROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 2 2010Shoulei Jiang Abstract Controversy remains about the identity of the transcription factor(s) (TFs), which bind to the two E-box elements (CACGTG, proximal and distal) of the human telomerase (hTERT) gene promoter, the essential elements in the regulation of telomerase. Here, systematic oligonucleotide trapping supplemented with 2-DE and proteomic methods was used to identify E-box binding TFs. Although insufficient purity was obtained from the proximal E-box element trapping, further fractionation provided by 2-DE and specific identification from Southwestern blotting analysis allow us to clearly identify an E-box binding TF. The protein spot was cut from 2-DE and in-gel digested with trypsin for LC-nanospray ESI-MS/MS analysis. This identified upstream stimulatory factor 2 (USF2). Western blotting analysis with specific antibodies clearly shows USF2 present in the purified fraction and USF2 antibody supershifts the specific DNA-binding complex on non-denaturing gels. Furthermore, a novel method was developed in which the specific DNA-TF complex was separated on a non-denaturing gel, the band was cut and applied to SDS-PAGE for a second dimension. Western blots of this second gel also confirmed the presence of USF2. [source] Proteomic and functional alterations in brain mitochondria from Tg2576 mice occur before amyloid plaque depositionPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 4 2007Frank Gillardon Dr. Abstract Synaptic dysfunction is an early event in Alzheimer's disease patients and has also been detected in transgenic mouse models. In the present study, we analyzed proteomic changes in synaptosomal fractions from Tg2576 mice that overexpress mutant human amyloid precursor protein (K670N, M671L) and from their nontransgenic littermates. Cortical and hippocampal tissue was microdissected at the onset of cognitive impairment, but before deposition of amyloid plaques. Crude synaptosomal fractions were prepared by differential centrifugation, proteins were separated by 2-D DIGE and identified by MS/MS. Significant alterations were detected in mitochondrial heat shock protein 70 pointing to a mitochondrial stress response. Subsequently, synaptosomal versus nonsynaptic mitochondria were purified from Tg2576 mice brains by density gradient centrifugation. Mitochondrial proteins were separated by IEF or Blue-native gel electrophoresis in the first dimension and SDS-PAGE in the second dimension. Numerous changes in the protein subunit composition of the respiratory chain complexes I and III were identified. Levels of corresponding mRNAs remain unchanged as shown by Affymetrix oligonucleotide array analysis. Functional examination revealed impaired state 3 respiration and uncoupled respiration in brain mitochondria from young Tg2576 mice. By immunoblotting, amyloid-beta oligomers were detected in synaptosomal fractions from Tg2576 mice and reduced glucose metabolism was observed in Tg2576 mice brains by [14C]-2-deoxyglucose infusion. Taken together, we demonstrate alterations in the mitochondrial proteome and function that occur in Tg2576 mice brains before amyloid plaque deposition suggesting that mitochondria are early targets of amyloid-beta aggregates. [source] Pilot study of the Human Proteome Organisation Brain Proteome Project: Applying different 2-DE techniques to monitor proteomic changes during murine brain developmentPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 18 2006Kai Stühler Dr. Abstract The Human Proteome Organisation Brain Proteome Project aims at coordinating neuroproteomic activities with respect to analysis of development, aging, and evolution in human and mice and at analysing normal aging processes as well as neurodegenerative diseases. Our group participated in the mouse pilot study of this project using two different 2-DE systems, to find out the optimal conditions for comprehensive gel-based differential proteome analysis. Besides the assessment of the best methodical conditions the question of "How many biological replicate analyses have to be performed to get reliable statistically validated results?" was addressed. In total 420 differences were detected in all analyses. Both 2-DE methods were found to be suitable for comprehensive differential proteome analysis. Nevertheless, each of the methods showed substantial advantages and disadvantages resulting in the fact that modification of both systems is essential. From our results we can draw the conclusions that for the future optimal quantitative differential gel-based brain proteome analyses the sample preparation has to be slightly changed, the resolution of the first as well as the second dimension has to be advanced, the number of experiments has to be increased and that the 2D-DIGE system should be applied. [source] On-line 2D-LC-ESI/MS/MS determination of rifaximin in rat serumBIOMEDICAL CHROMATOGRAPHY, Issue 11 2009R. Nageswara Rao Abstract A highly sensitive and selective on-line two-dimensional reversed-phase liquid chromatography/electrospray ionization,tandem mass spectrometry (2D-LC-ESI/MS/MS) method was developed and validated to determine rifaximin in rat serum by direct injection. The 2D-LC-ESI/MS/MS system consisted of a restricted access media column for trapping proteins as the first dimension and a Waters C18 column as second dimension using 0.1% aqueous acetic acid:acetonitrile as mobile phase in a gradient elution mode. Rifampacin was used as an internal standard. The linear dynamic range was 0.5,10 ng/mL (r2 > 0.998). Acceptable precision and accuracy were obtained over the calibration range. The assay was successfully used in analysis of rat serum to support pharmacokinetic studies. Copyright © 2009 John Wiley & Sons, Ltd. [source] | ||||||||||||||||