JOURNAL OF FOOD BIOCHEMISTRY, Issue 3 2004
ANNA LISA BASSO
The aim of this study was to assess the proteolytic activity of Lactobacillus sakei (DSM 6333), L. plantarum (B21), and to a lesser extent, L. farciminis (DSM 20184) on meat sarcoplasmic proteins. The protein composition was assayed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and capillary electrophoresis after incubation of meat extract inoculated with bacteria. All strains showed proteolytic activity: a band about 94 kDa disappeared in samples inoculated with L. farciminis and L. plantarum and strongly decreased in those inoculated with L. sakei. The intensity of the bands with a molecular weight between 94 and 38 kDa decreased in all samples. Capillary electrophoresis analysis ascertained the disappearance of the fractions corresponding to 8.64 and 8.66 min retention time in all samples. The bands corresponding to 94 kDa and 38 kDa were, respectively, identified as glycogen phosphorylase muscle isoform and glyceraldehyde 3-phosphate dehydrogenase, by in situ digestion of protein gel bands and peptide map analysis using Matrix Assisted Laser Desorption/Ionization - Time of Flight Mass Spectrometry (MALDI-TOF MS). [source]

A Mass Spectrometry Plate Reader: Monitoring Enzyme Activity and Inhibition with a Desorption/Ionization on Silicon (DIOS) Platform

CHEMBIOCHEM, Issue 7 2004
Zhouxin Shen Dr.
Abstract A surface-based laser desorption/ionization mass spectrometry assay that makes use of Desorption/Ionization on Silicon Mass Spectrometry (DIOS-MS) has been developed to monitor enzyme activity and enzyme inhibition. DIOS-MS has been used to characterize inhibitors from a library and then to monitor their activity against selected enzyme targets, including proteases, glycotransferase, and acetylcholinesterase. An automated DIOS-MS system was also used as a high-throughput screen for the activity of novel enzymes and enzyme inhibitors. On two different commercially available instruments, a sampling rate of up to 38 inhibitors per minute was accomplished, with thousands of inhibitors being monitored. The ease of applying mass spectrometry toward developing enzyme assays and the speed of surface-based assays such as DIOS for monitoring inhibitor effectiveness and enzyme activity makes it attractive for a broad range of screening applications. [source]

Chemically modified porous silicon for laser desorption/ionization mass spectrometry of ionic dyes

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 8 2009
I. V. Shmigol
Abstract Desorption/ionization on silicon (DIOS) mass spectra of model ionic dyes methylene blue (MB+Cl,) and methyl orange (Na+MO,) were studied using p+ type-derived porous silicon (PS) free layers. As-prepared PS (PS-H), the PS thermally oxidized at 300 °C (PS-OX), PS with chemically grafted cation-exchanging alkylsulfonic acid (PS-SO3H) and anion-exchanging propyl-octadecyldimethylammonium chloride (PS-ODMA+Cl,) groups was tested as ionization platforms. Two mechanisms of the methylene blue desorption/ionization were found: (1) the formation of [MB + H]+, ion due to the reduction/protonation of MB+, which is predominant for PS-H and PS-OX platforms and (2) direct thermal desorption of the MB+ cation, prevailing for PS-SO3H. The fragmentation of the cation is significantly suppressed in the latter case. The samples of PS-SO3H and PS-ODMA+ Cl, efficiently adsorb the dyes of the opposite charge from their solutions via the ion-exchange. Consequent DIOS MS studies allow to detect only low fragmented ions (MB+ and MO,, respectively), demonstrating the potential of the ion-exchange adsorption combined with DIOS MS for the analysis of ionic organic compounds in solutions. Copyright © 2009 John Wiley & Sons, Ltd. [source]

Pressure Ionization and Transitions in Dense Hydrogen

CONTRIBUTIONS TO PLASMA PHYSICS, Issue 3-4 2005
W. Ebeling
Abstract Shock experiments with fluid hydrogen have shown that a transition from insulating behavior to metal-like conductivity occurs at pressures beyond 100 GPa. This requires the development of new methods to describe the transition region of dense plasmas. The traditional approach due to Saha is based on the assumption of chemical equilibrium between charged and neutral components. This is equivalent to minimizing the free energy with respect to the composition. Here we improve an expression for the free energy developed recently to determine Hugoniot curves and isentropes in dense hydrogen and deuterium plasma in the regions of partial dissociation and partial ionization. We show that at high pressures the influence of the excluded volume occupied by neutral species is crucial for the transition to full ionization. We present curves for several thermodynamic functions for the region 5000 K < T < 20000 K and 0.6 g/cm3 < , < 1 g/cm3. The influence of the effective radii of the neutral species is crucial in the transition region. (© 2005 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source]

Determination of the bacterial pathogen Edwardsiella tarda in fish species by capillary electrophoresis with blue light-emitting diode-induced fluorescence

ELECTROPHORESIS, Issue 18-19 2004
Lijun Yu
Abstract High-performance capillary electrophoresis (HPCE) has been applied to the identification, separation, and quantitation of intact bacteria. We demonstrate that a pathogen (Edwardsiella tarda) which causes systemic infection in commercially important fish species can be rapidly identified and determined (<10 min) after direct injection into fish fluid by CE blue light-emitting diode (LED)-induced fluorescence. SYTO 13 (488 nm/509 nm), a cell-permeable green nucleic acid stain, was used to stain the cells. Remarkably high efficiency (>1 200,000 theoretical plates/m) was achieved with this rapid and efficient CE method. It was found that proper sample vortexing (90 s) would be beneficial to disperse aggregated cells and facilitate the focusing of intact cells during electrophoresis. Ionization of the surface constituents of Edwardsiella tarda cells provided efficient surface charges for the intact cells to be separated from the EOF and damaged or lysed cells when the separation was performed in running buffer (3.94 mM Tris, 0.56 mM borate, 0.013 mM EDTA) at pH 10.5. The limit of detection (LOD) and recovery were found to be 4.2×104 cells/mL and 70.0%, respectively. This proposed CE method could become an effective tool for diagnosis and tracking of certain diseases caused by bacteria in fish species as well as in human beings. [source]

Aquatic toxicity of triclosan

ENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 7 2002
David R. Orvos
Abstract The aquatic toxicity of triclosan (TCS), a chlorinated biphenyl ether used as an antimicrobial in consumer products, was studied with activated-sludge microorganisms, algae, invertebrates, and fish. Triclosan, a compound used for inhibiting microbial growth, was not toxic to wastewater microorganisms at concentrations less than aqueous solubility. The 48-h Daphnia magna median effective concentration (EC50) was 390 ,g/L and the 96-h median lethal concentration values for Pimephales promelas and Lepomis macrochirus were 260 and 370 ,g/L, respectively. A no-observed-effect concentration (NOEC) and lowest-observed-effect concentration of 34.1 ,g/L and 71.3 ,g/L, respectively, were determined with an early life-stage toxicity test with Onco-rhynchus mykiss. During a 96-h Scenedesmus study, the 96-h biomass EC50 was 1.4 ,g/L and the 96-h NOEC was 0.69 ,g/L. Other algae and Lemna also were investigated. Bioconcentration was assessed with Danio rerio. The average TCS accumulation factor over the five-week test period was 4,157 at 3 ,g/L and 2,532 at 30 ,g/L. Algae were determined to be the most susceptible organisms. Toxicity of a TCS-containing wastewater secondary effluent to P. promelas and Ceriodaphnia was evaluated and no observed differences in toxicity between control and TCS-treated laboratory units were detected. The neutral form of TCS was determined to be associated with toxic effects. Ionization and sorption will mitigate those effects in the aquatic compartment. [source]

ESI+ MS/MS confirmation of canine ivermectin toxicity,

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 1 2009
A. F. Lehner
Abstract Ivermectin is a semisynthetic macrocyclic lactone anthelmintic of the avermectin family derived from Streptomyces fermentation products. Avermectins are used as antiparasitic agents in domestic animals; although considered relatively safe, one must consider animal species, breed, weight, and age in dosage determinations. In January 2006, two canines were presented to the UK Livestock Disease Diagnostic Center after dying from suspected ivermectin overdoses [30,50 mg/kg body weight]. To confirm this clinical diagnosis we developed a rapid, sensitive semiquantitative ElectroSpray Ionization,Mass Spectrometry (ESI/MS) method for ivermectin in canine tissue samples. Pharmaceutical ivermectin contains two ivermectins differing by a single methyl group, and each compound forms interpretation-confounding adducts with tissue Na+ and K+ ions. We now report that ivermectin administration was clearly confirmed by comparison with standard and dosage forms of ivermectin, and simple proportionalities based on mass spectral intensity of respective molecular ions allowed semiquantitative estimates of injection site tissue concentrations of 20 and 40 µg/g tissue (wet weight) in these animals, consistent with the history of ivermectin administration and the clinical signs observed. There is a distinct need for both rapid detection and confirmation of toxic exposures in veterinary diagnostics, whether for interpretation of clinical cases antemortem or for forensic reasons postmortem. It is vital that interpreters of analytical results have appropriate guidance in the scientific literature and elsewhere so as to enable clear-cut answers. The method presented here is suitable for routine diagnostic work in that it allows rapid extraction of ivermectin from tissue samples, avoids the need for high-performance liquid chromatography and allows ready interpretation of the multiple ivermectin species seen by ESI+ MS/MS in samples originating from veterinary dosage forms. Copyright © 2008 John Wiley & Sons, Ltd. [source]

Ionization, shocks and evolution of the emission-line gas of distant 3CR radio galaxies

MONTHLY NOTICES OF THE ROYAL ASTRONOMICAL SOCIETY, Issue 1 2000
P. N. Best
An analysis of the kinematics and ionization state of the emission-line gas of a sample of 14 3CR radio galaxies with redshifts z,1 is carried out. The data used for these studies, deep long-slit spectroscopic exposures from the William Herschel Telescope, are presented in an accompanying paper. It is found that radio sources with small linear sizes (,150 kpc) have lower ionization states, higher emission-line fluxes and broader line widths than larger radio sources. An analysis of the low-redshift sample of Baum et al. demonstrates that radio galaxies at low redshift show similar evolution in their velocity structures and emission-line ratios from small to large radio sources. The emission-line ratios of small radio sources are in agreement with theoretical shock ionization predictions, and their velocity profiles are distorted. Together with the other emission-line properties, this indicates that shocks associated with the radio source dominate the kinematics and ionization of the emission-line gas during the period that the radio source is expanding through the interstellar medium. Gas clouds are accelerated by the shocks, giving rise to the irregular velocity structures observed, whilst shock compression of emission-line gas clouds and the presence of the ionizing photons associated with the shocks combine to lower the ionization state of the emission-line gas. By contrast, in larger sources the shock fronts have passed well beyond the emission-line regions; the emission-line gas of these larger radio sources has much more settled kinematical properties, indicative of rotation, and emission-line ratios consistent with the dominant source of ionizing photons being the active galactic nucleus. This strong evolution with radio size of the emission-line gas properties of powerful radio galaxies mirrors the radio size evolution seen in the nature of the optical,ultraviolet continuum emission of these sources, implying that the continuum alignment effect is likely to be related to the same radio source shocks. [source]

Porous silicon surfaces for metabonomics: Detection and identification of nucleotides without matrix interference

PHYSICA STATUS SOLIDI (C) - CURRENT TOPICS IN SOLID STATE PHYSICS, Issue 6 2007
D. Gómez
Abstract In present work, porous silicon surfaces (PSS) have been developed for time of flight mass spectrometric experiments (TOF-MS) in the monitoring of nucleotides, commonly found as metabolites in the cell. The mass range of the studied molecules (, 400 amu) is common to several important messengers and other metabolites. Different porosified surfaces have been developed by means of electrochemical etching and different degree of porosity and pore size achieved as function of silicon dopant concentration, silicon resistivity, current density and the presence or absence of illumination along the process. As main conclusion, it can be said that an interesting commercial nucleotide (Cyclic adenosine monophosphate, c-AMP) has been detected on low concentrations (,hundreds of femtomols) for some of the fabricated porous surfaces. Taking into account that these concentrations are similar to the ones found in real samples, this result opens the possibility to the fabrication of DIOS (Desorption Ionization On Silicon) chips for the detection of nucleotides in biological fluids. (© 2007 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source]

Multiple Soft Ionization of Gas-Phase Proteins and Swift Backbone Dissociation in Collisions with ,99,eV Electrons,

ANGEWANDTE CHEMIE, Issue 8 2010
Roman
Unerwartet: Elektronen mit einer Energie über 20,eV können Polypeptide mehrmals ionisieren (siehe Bild), ohne dabei die Primär- oder sogar Elemente der Tertiärstruktur zu zerstören. Durch gleichzeitige elektronische Anregung lassen sich Rückgratbindungen rasch spalten, wobei schwache Molekül-Molekül-Bindungen intakt bleiben. Dieses ,sanfte Fragmentieren" sollte eine genaue Charakterisierung der Struktur von Proteinkomplexen ermöglichen. [source]

Evaluation of Intrinsic Ionization and Complexation Constants of TiO2 and Mg-Fe Hydrotalcite-like Compounds

CHINESE JOURNAL OF CHEMISTRY, Issue 10 2006
Wan-Guo Hou
Abstract The intrinsic surface reaction constants, pKinta1, pKinta2, p*KintC and p*KintA, were evaluated by a modified double extrapolation (MDE) for TiO2 without structural charge and Mg-Fe hydrotalcite-like compounds (HTlc) with structural charge, respectively. The results of intrinsic surface reaction constants for TiO2 were compared with those obtained by class double extrapolation (CDE) in literature. Furthermore, the values of intrinsic surface reaction constants obtained by MDE were used to simulate the charging behaviors of the materials. The following conclusions were obtained. For TiO2 without structural charge, the pKinta1 and pKinta2 evaluated by MDE are equal to those by CDE, however the p*KintC and p*KintA evaluated by MDE are much different from those by CDE. In principle, the results of the p*KintC and p*KintA evaluated by MDE are more accurate than those by CDE. The values of intrinsic surface reaction constants obtained by MDE can excellently simulate the charging curves for TiO2 with the triple layer model (TLM). For HTlc with positive structural charge, the results of *KintC=0 and *KintA,, were obtained by MDE, which means the inert electrolyte chemical binding does not exist; the point of zero net charge (PZNC) of c -independence also exist as the same as solid without structural charge, and the pHPZNC obtained by the acid-base titration can excellently be simulated and the surface charging tendency can be simulated to a great extent using the pKinta1 and pKinta2 evaluated by MDE and the diffuse layer model (DLM). [source]

Comparison of two glutaraldehyde immobilization techniques for solid-phase tryptic peptide mapping of human hemoglobin by capillary zone electrophoresis and mass spectrometry

ELECTROPHORESIS, Issue 9 2004
Isabelle Migneault
Abstract Stabilization of proteolytic enzymes, especially by immobilization, is of considerable interest because of their potential applications in medicine and the chemical and pharmaceutical industries. We report here a detailed comparison of two procedures for trypsin immobilization using the same homobifunctional agent, glutaraldehyde, for the purpose of peptide mapping. These methods include covalent coupling either to controlled pore glass (solid support) or via a cross-linking reaction (without any solid support). The immobilized trypsin preparations were characterized by the determination of immobilization efficiency, which ranged from 68 to > 95%, and measurement of apparent kinetic parameters toward a synthetic peptide-like substrate. Batch digestions of whole denaturated human normal adult hemoglobin (HbA) were performed to obtain peptide maps by capillary zone electrophoresis (CZE). Migration time reproducibility of the CZE maps was excellent, with a mean relative standard deviation of 1.5%. Moreover, the two immobilized enzyme preparations showed excellent reproducibility for repeated digestions. Matrix-assisted laser desorption/ionization (MALDI)-mass spectrometry was also used for peptide mass mapping of denaturated HbA digested using the two immobilized trypsin preparations. Even though the two immobilized trypsin preparations do not behave identically, similar sequence coverages of 57% and 61% (for the two HbA chains merged) were achieved for the support-based and cross-linked trypsin preparations, respectively. [source]

Actinide-Transition Metal Heteronuclear Ions and Their Oxides: {IrUO}+ as an Analogue to Uranyl

EUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 17 2006
Marta Santos
Abstract Recent theoretical calculations have shown that Ir should behave as a chemical analogue to N, with the result that IrUO+, like known NUO+, is predicted to be a stable species isoelectronic with UO22+, the uranyl dication. The target heterometallic analogue to uranyl has now been prepared by direct laser desorption/ionization of a U/Ir alloy, and by oxidation of UIr+ with N2O and C2H4O. Properties of UIr+, UPt+, and UAu+ bimetallic ions have been studied. They demonstrate direct actinide,transition metal bonding, and support the concept of "autogenic isolobality". (© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2006) [source]

Alterations in Barrett's-related adenocarcinomas: A proteomic approach

INTERNATIONAL JOURNAL OF CANCER, Issue 6 2008
DunFa Peng
Abstract In this study, we applied high-resolution, two-dimensional, gel electrophoresis and matrix-assisted laser desorption/ionization, time-of-flight and tandem mass spectrometry analysis (MALDI TOF MS) to identify novel proteins that are involved in Barrett's tumorigenesis. We analyzed 12 primary tissue samples that included 8 Barrett's-related adenocarcinomas (BA) and 4 normal mucosae samples. Twenty-three spots were consistently altered (,2-fold) in at least half of the tumors when compared with all normal samples and thus subjected to further analysis. The MALDI TOF MS analysis demonstrated biologically interesting upregulated proteins such as ErbB3, Dr5 and Cyclin D1 as well as several members of the zinc finger proteins (Znf146, Znf212 and Znf363). Examples of downregulated proteins included Lgi1 and Klf6. We selected four proteins (ErbB3, Dr5, Znf146 and Lgi1) that are novel for BAs for validation using quantitative real-time reverse-transcription PCR on 39 BA tissue samples when compared with normal samples. We demonstrated mRNA upregulation of ERBB3 (51.3%), DR5 (41%) and ZNF146 (30.7%) and downregulation of LGI1 (100%) in BA. We have further validated the protein overexpression of ErbB3, Dr5 and Znf146, using immunohistochemical (IHC) analysis on a tissue microarray that contained 75 BAs and normal gastric and esophageal mucosae samples. BA tissue samples demonstrated overexpression of ErbB3 (42%), Dr5 (90%) and Znf146 (30%) when compared with normal tissues. In conclusion, we have identified and validated several novel proteins that are involved in Barrett's carcinogenesis. © 2007 Wiley-Liss, Inc. [source]

The Detection of Multiply Charged Dyes Using Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry for the Forensic Examination of Pen Ink Dyes Directly from Paper

JOURNAL OF FORENSIC SCIENCES, Issue 5 2007
Jamie D. Dunn M.S.
Abstract:, Laser desorption mass spectrometry (LDMS) is emerging as a technique for questioned document examination. Its use is limited to detecting ink dyes that are neutral or singly charged. Several inks contain dyes that are multiply charged and LDMS cannot be employed for their identification. We have successfully detected >20 polyionic dyes that can be used in the manufacture of inks using matrix-assisted laser desorption/ionization (MALDI) MS, directly from paper, with the matrix, 2-(4-hydroxyphenylazo)benzoic acid (HABA), and the additive, diammonium hydrogen citrate (DAHC). For example, Acid Violet 49, a charged dye containing one positively-charged site and two negatively charged sulfonate groups, cannot be detected by LDMS, but forms intact, singly charged ions in the MALDI MS experiment. The method described is also useful for identifying multiply charged dye mixtures that are used in modern pen inks. [source]

Parental relationships among three grape varieties studied by MALDI of grape seed protein profiles,

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 9 2010
Antonella Bertazzo
Abstract Two Raboso cultivars, i.e. Raboso Veronese and Raboso Piave, are two black Vitis vinifera varieties. A genetic study suggested that Raboso Veronese is the progeny of a spontaneous cross between Raboso Piave and Marzemina Bianca cultivars. Parental relationships are usually investigated by genetic studies, which are effective to establish genetic links among different cultivars. Considering that proteome is the genome expression, in this article we evaluated the power of seed protein profiles obtained by matrix-assisted laser desorption/ionization (MALDI)/MS for parentage investigation. The three cultivars lead to very similar spectra with differences in the relative intensity of the most abundant species and the presence of very weak specific ions. In order to evaluate the analytical significance of these aspects, the variability due to instrumental factors and due to different harvesting areas and years of the same cultivars have been considered and measured by the calculation of discrepancy factor values. On one hand, the results obtained can be considered a valid confirmation of the genomic findings, whereas on the other hand, the results provide evidence for the ability of MALDI/MS to individuate minor differences in protein profiles of complex protein mixtures. Copyright © 2010 John Wiley & Sons, Ltd. [source]

Molecular dynamics simulations of MALDI: laser fluence and pulse width dependence of plume characteristics and consequences for matrix and analyte ionization

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 4 2010
Richard Knochenmuss
Abstract Molecular dynamics simulations of matrix-assisted laser desorption/ionization were carried out to investigate laser pulse width and fluence effects on primary and secondary ionization process. At the same fluence, short (35 or 350 ps) pulses lead to much higher initial pressures and ion concentrations than longer ones (3 ns), but these differences do not persist because the system relaxes toward local thermal equilibrium on a nanosecond timescale. Higher fluences accentuate the initial disparities, but downstream differences are not substantial. Axial velocities of ions and neutrals are found to span a wide range, and be fluence dependent. Total ion yield is only weakly dependent on pulse width, and consistent with experimental estimates. Secondary reactions of matrix cations with analyte neutrals are efficient even though analyte ions are ablated in clusters of matrix. Copyright © 2010 John Wiley & Sons, Ltd. [source]

Laser desorption postionization for imaging MS of biological material

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 2 2010
Artem Akhmetov
Abstract Vacuum ultraviolet single photon ionization (VUV SPI) is a soft ionization technique that has the potential to address many of the limitations of matrix-assisted laser desorption/ionization (MALDI) for imaging MS. Laser desorption postionization (LDPI) uses VUV SPI for postionization and is experimentally analogous to a MALDI instrument with the addition of a pulsed VUV light source. This review discusses progress in LDPI-MS over the last decade, with an emphasis on imaging MS of bacterial biofilms, analytes whose high salt environment make them particularly resistant to imaging by MALDI-MS. This review first considers fundamental aspects of VUV SPI including ionization mechanisms, cross sections, quantum yields of ionization, dissociation and potential mass limits. The most common sources of pulsed VUV radiation are then described along with a newly constructed LDPI-MS instrument with imaging capabilities. Next, the detection and imaging of small molecules within intact biofilms is demonstrated by LDPI-MS using 7.87 eV (157.6 nm) VUV photons from a molecular fluorine excimer laser, followed by the use of aromatic tags for detection of selected species within the biofilm. The final section considers the future prospects for imaging intact biological samples by LDPI-MS. Copyright © 2010 John Wiley & Sons, Ltd. [source]

Matrix-assisted laser desorption/ionization tissue profiling of secretoneurin in the nucleus accumbens shell from cocaine-sensitized rats

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 1 2010
Joachim D. Uys
Abstract Proteins in the nucleus accumbens mediate many cocaine-induced behaviors. In an effort to measure changes in nucleus accumbens protein expression as potential biomarkers for addiction, coronal tissue sections were obtained from rats that developed behavioral sensitization after daily administration of cocaine, or from daily saline-treated controls. The tissue sections were subjected to matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) profiling and tissue imaging. For profiling experiments, brain sections were manually spotted with matrix over the nucleus accumbens, a brain region known to regulate cocaine sensitization. Summed mass spectra (10 000 laser shots, grid) were acquired and spectra were aligned to reference peaks. Using bioinformatics tools, eight spectral features were found to be altered by cocaine treatment. Based on additional sequencing experiments with MALDI tandem MS and database searches of measured masses, secretoneurin (m/z 3653) was identified as having an increased expression. In addition, the distribution of m/z 3653 in the nucleus accumbens was determined by MALDI tissue imaging, and the increased expression of its precursor protein, secretogranin II, was verified by immunoblotting. Copyright © 2009 John Wiley & Sons, Ltd. [source]

A convenient purification and preconcentration of peptides with ,-cyano-4-hydroxycinnamic acid matrix crystals in a pipette tip for matrix-assisted laser desorption/ionization mass spectrometry,

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 1 2010
Helena, ehulková
Abstract Peptide samples derived from enzymatic in-gel digestion of proteins resolved by gel electrophoresis often contain high amount of salts originating from reaction and separation buffers. Different methods are used for desalting prior to matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS), e.g. reversed-phase pipette tip purification, on-target washing, adding co-matrices, etc. As a suitable matrix for MALDI MS of peptides, ,-cyano-4-hydroxycinnamic acid (CHCA) is frequently used. Crystalline CHCA shows the ability to bind peptides on its surface and because it is almost insoluble in acidic water solutions, the on-target washing of peptide samples can significantly improve MALDI MS signals. Although the common on-target washing represents a simple, cheap and fast procedure, only a small portion of the available peptide solution is efficiently used for the subsequent MS analysis. The present approach is a combination of the on-target washing principle carried out in a narrow-end pipette tip (e.g. GELoader tip) and preconcentration of peptides from acidified solution by passing it through small CHCA crystals captured inside the tip on a glass microfiber frit. The results of MALDI MS analysis using CHCA-tip peptide preconcentration are comparable with the use of homemade POROS R2 pipette tip microcolumns. Advantages and limitations of this approach are discussed. Copyright © 2009 John Wiley & Sons, Ltd. [source]

From large analogical instruments to small digital black boxes: 40 years of progress in mass spectrometry and its role in proteomics.

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 8 2009
Part II 198
Abstract This is the continuation of a personal retrospective on the developments that since 1965 have given shape to Mass Spectrometry (MS) and taken it from a position of simply playing a role in Protein Chemistry to becoming an indispensable tool in Proteomics, all within a 40-year span. Part I covered the period from 1965 to 1984. This second part reviews the Mass Spectrometry timeline of events from 1985 to 2000, stopping at various time points where MS made significant contributions to protein chemistry or where the development of new instrumentation for MS represented a major advance for peptide and protein work. Major highlights in the field and their significance for peptide and protein characterization such as the advent and practical consequences of electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI) are covered, including work done with triple quads, the development of time-of-flight (TOF) instruments and new ion traps and going on to the more recent work on the full characterization of the Proteome with ion traps, TOF instruments and new ionization and tagging techniques for protein sequencing. Copyright © 2009 John Wiley & Sons, Ltd. [source]

Chemically modified porous silicon for laser desorption/ionization mass spectrometry of ionic dyes

Microfluidic chips for mass spectrometry-based proteomics

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 5 2009
Jeonghoon Lee
Abstract Microfluidic devices coupled to mass spectrometers have emerged as excellent tools for solving the complex analytical challenges associated with the field of proteomics. Current proteome identification procedures are accomplished through a series of steps that require many hours of labor-intensive work. Microfluidics can play an important role in proteomic sample preparation steps prior to mass spectral identification such as sample cleanup, digestion, and separations due to its ability to handle small sample quantities with the potential for high-throughput parallel analysis. To utilize microfluidic devices for proteomic analysis, an efficient interface between the microchip and the mass spectrometer is required. This tutorial provides an overview of the technologies and applications of microfluidic chips coupled to mass spectrometry for proteome analysis. Various approaches for combining microfluidic devices with electrospray ionization (ESI) and matrix-assisted laser desorption/ionization (MALDI) are summarized and applications of chip-based separations and digestion technologies to proteomic analysis are presented. Copyright © 2009 John Wiley & Sons, Ltd. [source]

Modern MALDI time-of-flight mass spectrometry

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 3 2009
Marvin L. Vestal
Abstract This paper focuses on development of time-of-flight (TOF) mass spectrometry in response to the invention of matrix-assisted laser desorption/ionization (MALDI). Before this breakthrough ionization technique for nonvolatile molecules, TOF was generally considered as a useful tool for exotic studies of ion properties but was not widely applied to analytical problems. Improved TOF instruments and software that allow the full potential power of MALDI to be applied to difficult biological applications are described. A theoretical approach to the design and optimization of MALDI-TOF instruments for particular applications is presented. Experimental data are provided that are in excellent agreement with theoretical predictions of resolving power and mass accuracy. Data on sensitivity and dynamic range using kilohertz laser rates are also summarized. These results indicate that combinations of high-performance MALDI-TOF and TOF-TOF with off-line high-capacity separations may ultimately provide throughput and dynamic range several orders of magnitude greater than those currently available with electrospray LC-MS and MS-MS. Copyright © 2009 John Wiley & Sons, Ltd. [source]

Characterization of natural wax esters by MALDI-TOF mass spectrometry

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 1 2009
Vladimír Vrkoslav
Abstract The applicability of matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) to the analysis of wax esters (WEs) was investigated. A series of metal salts of 2,5-dihydroxybenzoic acid (DHB) was synthesized and tested as possible matrices. Alkali metal (Li, Na, K, Rb, Cs) and transition metal (Cu, Ag) salts were studied. The matrix properties were evaluated, including solubility in organic solvents, threshold laser power that should be applied for successful desorption/ionization of WEs, the nature of the matrix ions and the mass range occupied by them, and the complexity of the isotope clusters for individual metals. Lithium salt of dihydroxybenzoic acid (LiDHB) performed the best and matrices with purified lithium isotopes (6LiDHB or 7LiDHB) were recommended for WEs. Three sample preparation procedures were compared: (1) mixing the sample and matrix in a glass vial and deposition of the mixture on a MALDI plate (Mix), (2) deposition of sample followed by deposition of matrix (Sa/Ma), and (3) deposition of matrix followed by deposition of sample (Ma/Sa). Morphology of the samples was studied by scanning electron microscopy. The best sample preparation technique was Ma/Sa with the optimum sample to matrix molar ratio 1 : 100. Detection limit was in the low picomolar range. The relative response of WEs decreased with their molecular weight, and minor differences between signals of saturated and monounsaturated WEs were observed. MALDI spectra of WEs showed molecular adducts with lithium [M + Li]+. Fragments observed in postsource decay (PSD) spectra were related to the acidic part of WEs [RCOOH + Li]+ and they were used for structure assignment. MALDI with LiDHB was used for several samples of natural origin, including insect and plant WEs. A good agreement with GC/MS data was achieved. Moreover, MALDI allowed higher WEs to be analyzed, up to 64 carbon atoms in Ginkgo biloba leaves extract. Copyright © 2008 John Wiley & Sons, Ltd. [source]

Evaluation of a new matrix-free laser desorption/ionization method through statistic studies: comparison of the DIAMS (desorption/ionization on self-assembled monolayer surface) method with the MALDI and TGFA-LDI techniques

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 12 2008
Matthieu Bounichou
Abstract This work demonstrates that the desorption/ionization on self-assembled monolayer surface (DIAMS) mass spectrometry, a recent matrix-free laser desorption/ionization (LDI) method based on an organic target plate, is as statistically repeatable and reproducible as matrix assisted laser desorption ionization (MALDI) and thin gold film-assisted laser desorption/ionization (TGFA-LDI) mass spectrometries. On lipophilic DIAMS of target plates with a mixture of glycerides, repeatability/reproducibility has been estimated at 15 and 30% and the relative detection limit has been evaluated at 0.3 and 3 pmol, with and without NaI respectively. Salicylic acid and its d6 -isomer analysis confirm the applicability of the DIAMS method in the detection of compounds of low molecular weight. Copyright © 2008 John Wiley & Sons, Ltd. [source]

A sialylation study of mouse brain gangliosides by MALDI a-TOF and o-TOF mass spectrometry

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 6 2008
Mostafa Zarei
Abstract Matrix-assisted laser desorption/ionization (MALDI) process of sialoglycoconjugates is generally accompanied by different levels of cleavage of sialic acid residues and/or by dehydration, and decarboxylation reactions. Quantitative densitometry of the mouse brain ganglioside (MBG) components separated by high-performance thin layer chromatography (HPTLC) and evidenced by orcinol staining was a basis to verify the ganglioside composition pattern with respect to the relative abundances of individual components in the mixture. A systematic mass spectrometry (MS) sialylation analysis has been carried out to evaluate the feasibility of an axial time-of-flight (a-TOF) MS, equipped with a vacuum MALDI source and an orthogonal-TOF (o-TOF) instrument with an ion source operated at about 1 mbar of N2. Besides, the esterification by one methyl group of the carboxyl group in sialic acid to increase the stability of the ganglioside species for MALDI MS analysis has been tested and the yield of intact ganglioside species and of the neutral loss of water and carbon dioxide estimated. For the sialylation analysis of native ganglioside mixtures the MALDI o-TOF analysis with 6-azo-2-thiothymine/diammonium citrate (ATT/DAC) as a matrix appears as an optimal approach for ganglioside profiling. Copyright © 2008 John Wiley & Sons, Ltd. [source]

Identification of proteins directly from tissue: in situ tryptic digestions coupled with imaging mass spectrometry

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 2 2007
M. Reid Groseclose
Abstract A novel method for on-tissue identification of proteins in spatially discrete regions is described using tryptic digestion followed by matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) with MS/MS analysis. IMS is first used to reveal the protein and peptide spatial distribution in a tissue section and then a serial section is robotically spotted with small volumes of trypsin solution to carry out in situ protease digestion. After hydrolysis, 2,5-Dihydroxybenzoic acid (DHB) matrix solution is applied to the digested spots, with subsequent analysis by IMS to reveal the spatial distribution of the various tryptic fragments. Sequence determination of the tryptic fragments is performed using on-tissue MALDI MS/MS analysis directly from the individual digest spots. This protocol enables protein identification directly from tissue while preserving the spatial integrity of the tissue sample. The procedure is demonstrated with the identification of several proteins in the coronal sections of a rat brain. Copyright © 2007 John Wiley & Sons, Ltd. [source]

Direct laser desorption/ionization time-of-flight mass spectrometry of conjugated polymers

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 1 2007
Zhun Ma
Abstract Two conjugated polymers (CPs), poly(9,9-dioctylfluorene) (PF) and poly(3-octylthiophene) (PT) were analyzed by direct laser desorption/ionization time-of-flight mass spectrometry (LDI-ToF MS). Because of their strong absorption near the wavelength of the laser (337 nm), easy and transient energy transfer properties and sufficient thermal stability, CPs can be desorbed and ionized directly without a matrix. For comparison, these two polymers were also analyzed using matrix-assisted laser desorption/ionization (MALDI)-ToF MS in the positive reflectron mode. The results revealed that they are very similar in terms of quality and resolution. All results demonstrate that LDI-ToF MS is an alternative method for the mass characterization of some conjugated systems, thereby simplifying the process of sample preparation and result analysis. Copyright © 2006 John Wiley & Sons, Ltd. [source]

Effective detection of peptides containing cysteine sulfonic acid using matrix-assisted laser desorption/ionization and laser desorption/ionization on porous silicon mass spectrometry

JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 1 2006
Tomoya Kinumi
Abstract Cysteine sulfonic acid-containing peptides, being typical acidic peptides, exhibit low response in matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. In this study, matrix conditions and the effect of diammonium hydrogencitrate (DAHC) as additive were investigated for ionization of cysteine sulfonic acid-containing peptides in MALDI. A matrix-free ionization method, desorption/ionization on porous silicon (DIOS), was also utilized to evaluate the effect of DAHC. When equimolar three-component mixtures of peptides carrying free cysteine, cysteine sulfonic acid, and carbamidomethyl cysteine were measured by MALDI using a common matrix, ,-cyano-4-hydroxycinnamic acid (CHCA), no signal corresponding to cysteine sulfonic acid-containing peptide could be observed in the mass spectrum. However, by addition of DAHC to CHCA, the peaks of cysteine sulfonic acid-containing peptides were successfully observed, as well as when using 2,4,6-trihydroxyacetophenone (THAP) and 2,6-dihydroxyacetophenone with DAHC. In the DIOS mass spectra of these analytes, the use of DAHC also enhanced the peak intensity of the cysteine sulfonic acid-containing peptides. On the basis of studies with these model peptides, tryptic digests of oxidized peroxiredoxin 6 were examined as a complex peptide mixture by MALDI and DIOS. In MALDI, the peaks of cysteine sulfonic acid-containing peptides were observed when using THAP/DAHC as the matrix, but this was not so with CHCA. In DIOS, the signal from cysteine sulfonic acid-containing peptides was suppressed; however, the use of DAHC significantly enhanced the signal intensity with an increase in the number of observed peptides and increased signal-to-noise ratio in the DIOS spectra. The results show that DAHC in the matrix or on the DIOS chip decreases discrimination and suppression effects in addition to suppressing alkali-adduct ions, which leads to a beneficial effect on protonation of peptides containing cysteine sulfonic acid. Copyright © 2005 John Wiley & Sons, Ltd. [source]