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Peak Capacity (peak + capacity)
Selected AbstractsPeak capacity of ion mobility mass spectrometry: the utility of varying drift gas polarizability for the separation of tryptic peptidesJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 4 2004Brandon T. Ruotolo Abstract Ion mobility mass spectrometry (IM-MS) peptide mass mapping experiments were performed using a variety of drift gases (He, N2, Ar and CH4). The drift gases studied cover a range of polarizabilities ((0.2,2.6) × 10,24 cm3) and the peak capacities obtained for tryptic peptides in each gas are compared. Although the different gases exhibit similar peak capacities (5430 (Ar) to 7580 (N2)) in some cases separation selectivity presumably based on peptide conformers (or conformer populations), is observed. For example the drift time profiles observed for some tryptic peptide ions from aldolase (rabbit muscle) show a dependence on drift gas. The transmission of high-mass ions (m/z > 2000) is also influenced by increased scattering cross-section of the more massive drift gases. Consequently the practical peak capacity for IM-MS separation cannot be assumed to be solely a function of resolution and the ability of a gas to distribute signals in two-dimensional space; rather, peak capacity estimates must account for the transmission losses experienced for peptide ions as the drift gas mass increases. Copyright © 2004 John Wiley & Sons, Ltd. [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] Peak capacity of ion mobility mass spectrometry: the utility of varying drift gas polarizability for the separation of tryptic peptidesJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 4 2004Brandon T. Ruotolo Abstract Ion mobility mass spectrometry (IM-MS) peptide mass mapping experiments were performed using a variety of drift gases (He, N2, Ar and CH4). The drift gases studied cover a range of polarizabilities ((0.2,2.6) × 10,24 cm3) and the peak capacities obtained for tryptic peptides in each gas are compared. Although the different gases exhibit similar peak capacities (5430 (Ar) to 7580 (N2)) in some cases separation selectivity presumably based on peptide conformers (or conformer populations), is observed. For example the drift time profiles observed for some tryptic peptide ions from aldolase (rabbit muscle) show a dependence on drift gas. The transmission of high-mass ions (m/z > 2000) is also influenced by increased scattering cross-section of the more massive drift gases. Consequently the practical peak capacity for IM-MS separation cannot be assumed to be solely a function of resolution and the ability of a gas to distribute signals in two-dimensional space; rather, peak capacity estimates must account for the transmission losses experienced for peptide ions as the drift gas mass increases. Copyright © 2004 John Wiley & Sons, Ltd. [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] The application of small porous particles, high temperatures, and high pressures to generate very high resolution LC and LC/MS separationsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 8 2007Robert Plumb Abstract The effect of combining sub-2 ,m porous particles with elevated operating temperatures on chromatographic performance has been investigated in terms of chromatographic efficiency, productivity, peak elution order, and observed operating pressure. The use of elevated temperature in LC does not increase the obtainable performance but allows the same performance to be obtained in less time. Increasing the column temperature did allow the use of longer columns, generating column efficiencies in excess of 100 000 plates and gradient peak capacities approaching 1000. Raising the temperature increased the optimal mobile phase linear velocity, negating somewhat the pressure benefits observed by reducing the solvent viscosity. When operating at higher temperature the analyte retention is not only reduced, but the order of elution will also often change. High temperature separations allowed exotic organic modifiers such as isopropanol to be exploited for alternative selectivity and faster analysis. Finally, care must be taken when using high temperature separations to ensure that the narrow peak widths produced do not compromise the quality of data obtained from detectors such as high resolution mass spectrometers. [source] Cationic and anionic lipid-based nanoparticles in CEC for protein separationELECTROPHORESIS, Issue 11 2010Christian Nilsson Abstract The development of new separation techniques is an important task in protein science. Herein, we describe how anionic and cationic lipid-based liquid crystalline nanoparticles can be used for protein separation. The potential of the suggested separation methods is demonstrated on green fluorescent protein (GFP) samples for future use on more complex samples. Three different CEC-LIF approaches for protein separation are described. (i) GFP and GFP N212Y, which are equally charged, were separated with high resolution by using anionic nanoparticles suspended in the electrolyte and adsorbed to the capillary wall. (ii) High efficiency (800,000 plates/m) and peak capacity were demonstrated separating GFP samples from Escherichia coli with cationic nanoparticles suspended in the electrolyte and adsorbed to the capillary wall. (iii) Three single amino-acid-substituted GFP variants were separated with high resolution using an approach based on a physical attached double-layer coating of cationic and anionic nanoparticles combined with anionic lipid nanoparticles suspended in the electrolyte. The soft and porous lipid-based nanoparticles were synthesized by a one-step procedure based on the self-assembly of lipids, and were biocompatible with a large surface-to-volume ratio. The methodology is still under development and the optimization of the nanoparticle chemistry and separation conditions can further improve the separation system. In contrast to conventional LC, a new interaction phase is introduced for every analysis, which minimizes carry-over and time-consuming column regeneration. [source] Microstructure of microemulsion in MEEKCELECTROPHORESIS, Issue 4 2010Yuhua Cao Abstract The influences of the composition of microemulsion on the microstructure including dimensions and , potentials of microdroplets were measured in details. The average dynamic dimension of microdroplets was measured by dynamic laser light scattering, and , potential was determined to characterize average surface charge density of microdroplets. The experiment results showed that increase of the amount of surfactant resulted in decrease of microdroplet size but almost invariant , potential, which would enlarge migration time of the microdroplet in MEEKC. With increment of cosurfactant concentration, the microdroplet size had an increasing trend, whereas the , potential decreased. Thus, observed migration velocity of microdroplets increased, which made the separation window in MEEKC shortened. Neither dimension nor , potential of microdroplets changed by varying both the type and the amount of the oil phase. Adding organic solvent as modifier to microemulsion did not change the microdroplet size, but lowered , potential. The migration time of microdroplet still became larger, since EOF slowed down owing to organic solvent in capillary. So, besides increment of surfactant concentration, organic additive could also enlarge the separation window. Increase of cosurfactant concentration was beneficial for separation efficiency thanks to the looser structure of swollen microdroplet, and the peak sharpening might compensate for the resolution and peak capacity owing to a narrow separation window. Except the oil phase, tuning the composition of microemulsion would change the microstructure, eventually could be exploited to optimize the resolution and save analysis time in MEEKC. [source] Accessible proteomics space and its implications for peak capacity for zero-, one- and two-dimensional separations coupled with FT-ICR and TOF mass spectrometryJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 3 2006Jennifer L. Frahm The number and wide dynamic range of components found in biological matrixes present several challenges for global proteomics. In this perspective, we will examine the potential of zero-dimensional (0D), one-dimensional (1D), and two-dimensional (2D) separations coupled with Fourier-transform ion cyclotron resonance (FT-ICR) and time-of-flight (TOF) mass spectrometry (MS) for the analysis of complex mixtures. We describe and further develop previous reports on the space occupied by peptides, to calculate the theoretical peak capacity available to each separations-mass spectrometry method examined. Briefly, the peak capacity attainable by each of the mass analyzers was determined from the mass resolving power (RP) and the m/z space occupied by peptides considered from the mass distribution of tryptic peptides from National Center for Biotechnology Information's (NCBI's) nonredundant database. Our results indicate that reverse-phase-nanoHPLC (RP-nHPLC) separation coupled with FT-ICR MS offers an order of magnitude improvement in peak capacity over RP-nHPLC separation coupled with TOF MS. The addition of an orthogonal separation method, strong cation exchange (SCX), for 2D LC-MS demonstrates an additional 10-fold improvement in peak capacity over 1D LC-MS methods. Peak capacity calculations for 0D LC, two different 1D RP-HPLC methods, and 2D LC (with various numbers of SCX fractions) for both RP-HPLC methods coupled to FT-ICR and TOF MS are examined in detail. Peak capacity production rates, which take into account the total analysis time, are also considered for each of the methods. Furthermore, the significance of the space occupied by peptides is discussed. Copyright © 2006 John Wiley & Sons, Ltd. [source] Peak capacity of ion mobility mass spectrometry: the utility of varying drift gas polarizability for the separation of tryptic peptidesJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 4 2004Brandon T. Ruotolo Abstract Ion mobility mass spectrometry (IM-MS) peptide mass mapping experiments were performed using a variety of drift gases (He, N2, Ar and CH4). The drift gases studied cover a range of polarizabilities ((0.2,2.6) × 10,24 cm3) and the peak capacities obtained for tryptic peptides in each gas are compared. Although the different gases exhibit similar peak capacities (5430 (Ar) to 7580 (N2)) in some cases separation selectivity presumably based on peptide conformers (or conformer populations), is observed. For example the drift time profiles observed for some tryptic peptide ions from aldolase (rabbit muscle) show a dependence on drift gas. The transmission of high-mass ions (m/z > 2000) is also influenced by increased scattering cross-section of the more massive drift gases. Consequently the practical peak capacity for IM-MS separation cannot be assumed to be solely a function of resolution and the ability of a gas to distribute signals in two-dimensional space; rather, peak capacity estimates must account for the transmission losses experienced for peptide ions as the drift gas mass increases. Copyright © 2004 John Wiley & Sons, Ltd. [source] Multidimensional chromatography coupled to mass spectrometry in analysing complex proteomics samplesJOURNAL OF SEPARATION SCIENCE, JSS, Issue 10 2010Péter Horvatovich Abstract Multidimensional chromatography coupled to mass spectrometry (LCn -MS) provides more separation power and an extended measured dynamic concentration range to analyse complex proteomics samples than one dimensional liquid chromatography coupled to mass spectrometry (1D-LC-MS). This review gives an overview of the most important aspects of LCn -MS with respect to optimizing peak capacity and evaluate orthogonality. We review recent developments in LCn -MS to analyse proteomics samples from the analyst point of view and give an overview over methods and future developments to process LCn -MS data for comprehensive differential protein expression profiling. Examples from our research, such as combining protein fractionation using high temperature reverse phase (RP) columns followed by analysis of the trypsin-digested fractions by RP LC-MS, serve to highlight possibilities and shortcomings of present-day approaches. Other LCn -MS systems that have been used to analyse highly complex shotgun proteomic samples, such as the combination of RP columns using low and high pH eluents or the combination of hydrophilic interaction liquid chromatography (HILIC) with RP-MS is discussed in detail. [source] Hydrophilic interaction LC of peptides: Columns comparison and clusteringJOURNAL OF SEPARATION SCIENCE, JSS, Issue 6-7 2010Sylvia Van Dorpe Abstract A wide variety of hydrophilic interaction chromatography (HILIC) stationary phase surface chemistries are currently available. Although their selectivity can be considerably different, column comparison or clustering using peptides is limited. In this study, ten pharmaceutically relevant model peptides are analyzed on seven different HILIC columns (bare silica, amide, poly-hydroxyethyl aspartamide, diol and zwitterionic) for the evaluation of their performance and classification. The responses examined include single and multiple responses: plate number, asymmetry factor, LOD, geometric mean resolution, resolution product, time corrected resolution product, peak capacity and chromatographic response function. Column classification was performed using hierarchical clustering and principal component analysis. Moreover, the overall performance quality of the HILIC columns was compared using a linear desirability function. Hierarchical clustering and principal component analysis showed consistent clusters. The zwitterionic phase was clustered apart from the other HILIC columns and both poly-aspartamide columns were clustered together. In addition, the two bare silica phases represent two different clusters, and thus different selectivities. Overall, the responses showed the best performance for one of the bare silica columns (Alltima-Alltech), followed by the zwitterionic phase (ZIC)-HILIC. Thus, these columns, belonging to different clusters, were found to be the best performing systems in pharmaceutical peptide analysis for the selected peptide set. [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] Elevated temperature,extended column length conventional liquid chromatography to increase peak capacity for the analysis of tryptic digestsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 2 2007Pat Sandra Abstract High efficiency separations (200 000 plates) were obtained on conventional LC equipment by coupling 8×25 cm×2.1 (or 4.6) mm id×5 ,m dp ODS columns (total length 2 m) and operation at 60°C using a dedicated LC oven. The peak capacity in this 1-D set-up was 900 for the separation of human serum tryptic peptides analyzed after depletion of six highly abundant proteins. The chromatographic performance of an elevated temperature,extended column length conventional LC is highlighted. [source] Simple 2D-HPLC using a monolithic silica column for peptide separationJOURNAL OF SEPARATION SCIENCE, JSS, Issue 10-11 2004Hiroshi Kimura Abstract Separation of peptides by fast and simple two-dimensional (2D)-HPLC was studied using a monolithic silica column as a second-dimension (2nd-D) column. Every fraction from the first column, 5 cm long (2.1 mm ID) packed with polymer-based cation exchange beads, was subjected to separation in the 2nd-D using an octadecylsilylated (C18) monolithic silica column (4.6 mm ID, 2.5 cm). A capillary-type monolithic silica C18 column (0.1 mm ID, 10 cm) was also employed as a 2nd-D column with split flow/injection. Effluent of the first dimension (1st-D) was directly loaded into an injector loop of 2nd-D HPLC. UV and MS detection were successfully carried out at high linear velocity of mobile phase at 2nd-D using flow splitting for the 4.6 mm ID 2nd-D column, or with direct connection of the capillary column to the MS interface. Two-minute fractionation in the 1st-D, 118-second loading, and 2-second injection by the 2nd-D injector, allowed one minute for gradient separation in the 2nd-D, resulting in a maximum peak capacity of about 700 within 40 min. The use of a capillary column in the 2nd-D led to less solvent consumption and better MS detectability compared to a larger-sized column. This kind of fast and simple 2D-HPLC utilizing monolithic silica columns will be useful for the separation of complex mixtures in a short time. [source] Application of comprehensive multidimensional gas chromatography combined with time-of-flight mass spectrometry (GC×GC-TOFMS) for high resolution analysis of hop essential oilJOURNAL OF SEPARATION SCIENCE, JSS, Issue 5-6 2004Mark T. Roberts Abstract The selection and quality of hops is a major determinant in beer flavour. Brewers acknowledge that distinctive characteristics of different hop varieties can be traced to the composition of their essential oils. The difficulty in characterising complex mixtures such as hop oil using 1-D chromatography is that many compounds co-elute. With the introduction of comprehensive multidimensional capillary gas chromatography (GC×GC), there is a tremendous improvement in the separation power or peak capacity. Recent work using GC×GC with flame ionisation detection has suggested that there may be over 1,000 compounds in hop oil. This work describes the use of GC×GC combined with TOFMS detection (Leco Pegasus 4D instrument) to analyse Target hop oil. The TOFMS spectral acquisition rate of 60 Hz provided sufficient spectra per peak (2-D peak base width of 0.1,0.2 s) for identification (119 components were identified with 45 previously unreported compounds). When analysing results, an advantage of GC×GC coupled to TOFMS is that 2-D chromatograms can be viewed for individual masses that are characteristic of particular functional groups. This allows the analyst to view the various homologous series of compounds although in certain cases coelution may still be present as shown by the esters with mass 75. [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] Increasing throughput and information content for in vitro drug metabolism experiments using ultra-performance liquid chromatography coupled to a quadrupole time-of-flight mass spectrometerRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 6 2005Jose Castro-Perez The field of drug metabolism has been revolutionized by liquid chromatography/mass spectrometry (LC/MS) applications with new technologies such as triple quadrupoles, ion traps and time-of-flight (ToF) instrumentation. Over the years, these developments have often relied on the improvements to the mass spectrometer hardware and software, which has allowed users to benefit from lower levels of detection and ease-of-use. One area in which the development pace has been slower is in high-performance liquid chromatography (HPLC). In the case of metabolite identification, where there are many challenges due to the complex nature of the biological matrices and the diversity of the metabolites produced, there is a need to obtain the most accurate data possible. Reactive or toxic metabolites need to be detected and identified as early as possible in the drug discovery process, in order to reduce the very costly attrition of compounds in late-phase development. High-resolution, exact mass measurement plays a very important role in metabolite identification because it allows the elimination of false positives and the determination of non-trivial metabolites in a much faster throughput environment than any other standard current methodology available to this field. By improving the chromatographic resolution, increased peak capacity can be achieved with a reduction in the number of co-eluting species leading to superior separations. The overall enhancement in the chromatographic resolution and peak capacity is transferred into a net reduction in ion suppression leading to an improvement in the MS sensitivity. To investigate this, a number of in vitro samples were analyzed using an ultra-performance liquid chromatography (UPLC) system, with columns packed with porous 1.7,,m particles, coupled to a hybrid quadrupole time-of-flight (ToF) mass spectrometer. This technique showed very clear examples for fundamental gains in sensitivity, chromatographic resolution and speed of analysis, which are all important factors for the demands of today's HTS in discovery. Copyright © 2005 John Wiley & Sons, Ltd. [source] | ||||||||||