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Separation Modes (separation + mode)
Selected AbstractsCo-electroosmotic capillary electrophoresis of basic proteins with 1-alkyl-3-methylimidazolium tetrafluoroborate ionic liquids as non-covalent coating agents of the fused-silica capillary and additives of the electrolyte solutionELECTROPHORESIS, Issue 11 2009Danilo Corradini Abstract The paper reports the results of a study carried out to evaluate the use of three 1-alkyl-3-methylimidazolium-based ionic liquids as non-covalent coating agents for bare fused-silica capillaries and additives of the electrolyte solutions (BGE) for CE of basic proteins in the co-EOF separation mode. The three ionic liquids are differentiated from each other by the length of the alkyl group on the imidazolium cation, consisting of either an ethyl, butyl or octyl substituent, whereas tetrafluoroborate is the common anionic component of the ionic liquids. Coating the capillary with the ionic liquid resulted in improved peak shape and protein separation, while the EOF was maintained cathodic. This indicates that each ionic liquid is effective at masking the protein interaction sites on the inner surface of the capillary, also when its adsorption onto the capillary wall has not completely neutralized all the negative charges arising from the ionization of the silanol groups and the ionic liquid is not incorporated into the BGE employed for separation. Using the coated capillaries with BGE containing the ionic liquid employed for the coating, at concentration low enough to maintaining the EOF cathodic, both peak shape and protein separation varied to different extents, based on the particular ionic liquid used and its concentration. Fast and efficient separation of the model basic protein mixture in co-electroosmotic CE is obtained with the 1-butyl-3-methylimidazolium tetrafluoroborate coated capillary and 100,mM acetate buffer (pH 4.0) containing 4.4,mM 1-butyl-3-methylimidazolium tetrafluoroborate as the BGE. [source] Simultaneous determination of melamine and related compounds by hydrophilic interaction liquid chromatography,electrospray mass spectrometryJOURNAL OF SEPARATION SCIENCE, JSS, Issue 17-18 2010Jingen Xia Abstract A hydrophilic interaction liquid chromatography coupled to ESI-MS (HILIC/ESI-MS) method for the simultaneous determination of melamine and related compounds, i.e. ammeline, ammelide and cyanuric acid in foods was developed and validated. The separation was accomplished on a Venusil HILIC column with a mobile phase of acetonitrile + 10,mM ammonium formate buffer solution at pH 3.5 (88:12, v/v) under isocratic elution mode. For the detection of the targets, the ESI probe worked in the positive and negative switching mode. For each compound, three ions were selected as qualitative ions to obtain high specificity, and the most abundant ion of each compound was selected for quantification to obtain high sensitivity. The target compounds were quantified using SIM with 15N3-melamine and 13C3-cyanuric acid being used as an internal standards in the positive and negative modes, respectively. Compared with RP separation mode, HILIC has merits such as high separation and anti-interference efficiency. The method validation including linearity, LOD, LOQ, precision and recovery proved that the method has merits such high sensitivity, specificity and simplicity versus the other methods reported in the literature. [source] The application of novel 1.7 ,m ethylene bridged hybrid particles for hydrophilic interaction chromatographyJOURNAL OF SEPARATION SCIENCE, JSS, Issue 9 2008Eric S. Grumbach Abstract An un-derivatized 1.7 ,m ethylene bridged hybrid (BEH) particle was evaluated for its utility in retaining polar species in hydrophilic interaction chromatography (HILIC), and was compared to a 3 ,m un-derivatized silica material. Retentivity as a function of mobile phase pH, polar modifier and ACN content was examined. Also, the efficiency of the two particle substrates was compared by plotting HETP vs. linear velocity. Improved chemical resistance of the un-derivatized BEH particle was compared to un-derivatized silica at pH 5, demonstrating no performance deterioration over the course of 2000 injections for the BEH particle, while the silica particle deteriorated rapidly after 800 injections. Lastly, ESI-MS sensitivity as a function of particle size and separation mode was demonstrated. A 2.2 to 4.7-times higher ESI-MS response was observed on the 1.7 ,m particle compared to the 3 ,m particle, whereas a 5.6 to 8.8-times higher ESI-MS response was observed using HILIC as when compared to traditional RP chromatography. [source] Capillary electrophoresis-electrospray-mass spectrometry in peptide analysis and peptidomicsELECTROPHORESIS, Issue 10 2008Miguel Herrero Abstract In the present work, an exhaustive review of the main developments and applications of CE-MS for peptide analysis is given. This review includes the use of different CE separation modes, MS analyzers, capillary coatings, preconcentration techniques, on-chip applications as well as other different multidimensional strategies for peptide analysis. Key applications are critically discussed and relevant works published from January 2000 to May 2007 are summarized including information concerning the type of sample, CE-MS parameters as well as some figures of merit of the different CE-MS procedures developed for peptide analysis and peptidomics. [source] Recent advances in capillary electrophoresis and capillary electrochromatography of peptidesELECTROPHORESIS, Issue 22-23 2003Václav Ka Abstract An overview of the recent developments in the applications of high-performance capillary electromigration methods, namely zone electrophoresis, isotachophoresis, isoelectric focusing, affinity electrophoresis, electrokinetic chromatography, and electrochromatography, to analysis, preparation, and physicochemical characterization of peptides is presented. New approaches to the theoretical description and experimental verification of the electromigration behavior of peptides and the methodological aspects of capillary electroseparations of peptides, such as rational selection of separation conditions, sample treatment, and suppression of adsorption, are discussed, and new developments in individual separation modes and new designs of detection systems applied to peptide separations are shown. Several types of applications of capillary electromigration methods to peptide analysis are presented: quality control and purity tests, determination in biomatrices, monitoring of physical and chemical changes and enzymatic conversions, amino acid and sequence analysis, and peptide mapping. The examples of micropreparative peptide separations are given and capabilities of capillary electromigration techniques to provide important physicochemical characteristics of peptides are demonstrated. [source] Comparison of HPLC enantioseparation of substituted binaphthyls on CD-, polysaccharide- and synthetic polymer-based chiral stationary phasesJOURNAL OF SEPARATION SCIENCE, JSS, Issue 9 2010Lucie Loukotková Abstract Retention and enantioseparation behavior of ten 2,2,-disubstituted or 2,3,2,-trisubstituted 1,1,-binaphthyls and 8,3,-disubstituted 1,2,-binaphthyls, which are used as catalysts in asymmetric synthesis, was investigated on eight chiral stationary phases (CSPs) based on ,-CD, polysaccharides (tris(3,5-dimethylphenylcarbamate) cellulose or amylose CSPs) and new synthetic polymers (trans -1,2-diamino-cyclohexane, trans -1,2-diphenylethylenediamine and trans -9,10-dihydro-9,10-ethanoanthracene-(11S,12S)-11,12-dicarboxylic acid CSPs). Normal-, reversed-phase and polar-organic separation modes were employed. The effect of the mobile phase composition was examined. The enantiomeric separation of binaphthyl derivatives, which possess quite similar structures, was possible in different enantioselective environments. The substituents and their positions on the binaphthyl skeleton affect their properties and, as a consequence, the separation system suitable for their enantioseparation. In general, the presence of ionizable groups on the binaphthyl skeleton, substitution with non-identical groups and a chiral axis in the 1,2, position had the greatest impact on the enantiomeric discrimination. The 8,3,-disubstituted 1,2,-binaphthyl derivatives were the most easily separated compounds in several separation systems. From all the chiral stationary phases tested, cellulose-based columns were shown to be the most convenient for enantioseparation of the studied analytes. However, the polymeric CSPs with their complementary behavior provided good enantioselective environments for some derivatives that could be hardly separated in any other chromatographic system. [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] Preparation and characterization of polymethacrylate monolithic capillary columns with dual hydrophilic interaction reversed-phase retention mechanism for polar compoundsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 15-16 2009í Urban Abstract Monolithic columns for capillary hydrophilic interaction liquid chromatography (HILIC) were prepared in fused-silica capillaries by radical co-polymerization of [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium hydroxide and ethylene dimethacrylate in various binary and ternary porogen solvent mixtures with azobisisobutyronitrile as the initiator of the polymerization reaction. Columns showed mixed separation modes: reversed-phase (RP) in water-rich mobile phases and HILIC at high concentrations of acetonitrile (>60,80%) in aqueous,organic mobile phases. A continuous change in retention was observed at increasing concentration of water in acetonitrile, giving rise to characteristic U-turn plots of retention factors versus the concentration of water in the mobile phase, with minima corresponding to the transition between the mechanisms controlling the retention. The selectivity of organic polymer monolithic columns for HILIC separations can be varied by adjusting the concentration of sulfobetaine monomer and the composition of the porogen solvent in the polymerization mixture. Under HILIC conditions, the monolithic capillary sulfobetaine columns show separation selectivities for polar phenolic acids similar to those of a commercial silica-based sulfobetaine ZIC-HILIC column, which, however, has limited selectivity in the RP mode due to lower retention. [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] Serially coupled microcolumn reversed phase liquid chromatography for shotgun proteomic analysisPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 7 2009Dingyin Tao Abstract Microcolumn RPLC (,RPLC) is one of the optimum separation modes for shotgun proteomic analysis. To identify as many proteins as possible by MS/MS, the improvement on separation efficiency and peak capacity of ,RPLC is indispensable. Although the increase in column length is one of the effective solutions, the preparation of a long microcolumn is rather difficult due to the high backpressure generated during the packing procedure. In our recent work, through connecting microcolumns of 5, 10, and 15,cm length via unions with minimal dead volume, long microcolumns with length up to 30,cm were obtained, with which 318 proteins were identified from proteins extracted from Escherichia coli by ,RPLC-ESI MS/MS, and similar distributions of Mw and pI were found with single and various coupled microcolumns. Furthermore, by using MS/MS with improved sensitivity, with such a serially coupled 30,cm long microcolumn, 1692 proteins were identified within 7,h from rat brain tissue, with false positive rate (FPR) <1%. All these results demonstrated that serially couple microcolumns might be of great promising to improve the separation capacity of ,RPLC in shotgun proteomic analysis. [source] Column selection and method development for the separation of nucleoside phosphotriester diastereoisomers, new potential anti-viral drugs.BIOMEDICAL CHROMATOGRAPHY, Issue 6 2005Application to cellular extract analysis Abstract Analytical HPLC methods using derivatized cellulose and amylose chiral stationary phases used in normal and reversed-phase modes were developed for the diastereoisomeric separation of mononucleotide prodrugs (pronucleotides) of 3,-azido-2,,3,-dideoxythymidine (AZT). The resolutions were performed with two silica-based celluloses using normal and reversed-phase methodologies: Tris-3,5-dimethylphenylcarbamate (Chiralcel OD-H and Chiracel OD-RH) and Tris-methylbenzoate (Chiralcel OJ and OJ-R). Two amyloses phases, Tris-3,5-dimethylphenylcarbamate (Chiralpak AD) and Tris-(S)-1-phenylethylcarbamate (Chiralpak AS), were used in normal-phase mode. Additionally, we developed separation using two stationary phases with immobilized cyclodextrins in reversed-phase and polar-organic modes. The mobile phase and the chiral stationary phase were varied to achieve the best resolution. Different types and concentration of aliphatic alcohols, acetonitrile or water in the mobile phase were also tested for the different separation modes. An optimal baseline separation (Rs > 1.5) was readily obtained with all silica-based celluloses and amyloses using a normal-phase methodology. The different columns gave complementary results in term of resolution. Limits of detection and quantification were 0.12,0.20 and 0.40,0.67 µm, respectively. This analytical method was applied in a preliminary study for the pronucleotide 2 quantification in cellular extract. Copyright © 2005 John Wiley & Sons, Ltd. [source] | ||||||||||