Characterization of Alkali Induced Formation of Lanthionine, Trisulfides, and Tetrasulfides from Peptide Disulfides using Negative Ion Mass Spectrometry

Peptide disulfides are unstable under alkaline conditions, resulting in the formation of products containing lanthionine and polysulfied linkages. Electrospray ionization mass spectrometry has been used to characterize major species obtained when cyclic and acyclic peptide disulfides are exposed to alkaline media. Studies on a model cyclic peptide disulfide (Boc - Cys - Pro - Leu - Cys - NHMe) and an acyclic peptide, oxidized glutathione, bis ((gamma)Glu Cys - Gly - COOH), are described. Disulfide cleavage reactions are initiated by the abstraction of (CH)-H-alpha or (CH)-H-beta protons of Cys residues, with Subsequent elimination of H2S or H2S2. The buildup of reactive thiol species which act on intermediates containing dehydroalanine residues, rationalizes the formation of lanthionine and polysulfide products. In the case of the cyclic peptide disulfide, the formation of cyclic products is facilitated by the intramolecular nature of the Michael addition reaction of thiols to the dehydroalanine residue. Mass spectral evidence for the intermediate species is presented by using alkylation of thiol groups as a trapping method. Mass spectral fragmentation in the negative ion mode of the peptides derived from trisulfides and tetrasulfides results in elimination of S-2. (J Am Soc Mass Spectrom 2009, 20, 783-791) (C) 2009 American Society for Mass Spectrometry.

Simultaneous determination of microcystin-LR and its glutathione conjugate in fish tissues by liquid chromatography-tandem mass spectrometry

A sensitive and selective liquid chromatography-tandem mass spectrometry method was developed and validated for the simultaneous quantitative determination of microcystin-LR (MC-LR) and its glutathione conjugate (MC-LR-GSH) in fish tissues. The analytes were extracted from fish liver and kidney using 0.01 M EDTA-Na-2-5% acetic acid, followed by a solid-phase extraction (SPE) on Oasis HLB and silica cartridges. High-performance liquid chromatography (HPLC) with electrospray ionization mass spectrometry, operating in selected reaction monitoring (SRM) mode, was used to quantify MC-LR and its glutathione conjugate in fish liver and kidney. Recoveries of analytes were assessed at three concentrations (0.2, 1.0, and 5 mu g g(-1) dry weight [DW]) and ranged from 91 to 103% for MC-LR, and from 65.0 to 75.7% for MC-LR-GSH. The assay was linear within the range from 0.02 to 5.0 mu g g(-1) DW, with a limit of quantification (LOQ) of 0.02 mu g g(-1) DW. The limit of detection (LOD) of the method was 0.007 mu g g(-1) DW in both fish liver and kidney. The overall precision was determined on three different days. The values for within- and between-day precision in liver and kidney were within 15%. This method was applied to the identification and quantification of MC-LR and its glutathione conjugate in liver and kidney of fish with acute exposure of MC-LR. (c) 2007 Elsevier B.V. All rights reserved.

Qualitative and semi-quantitative analysis of quaternary alkaloids in Coptis-scute herb couple by Electrospray mass Spectrometry

A practical solution of qualitatively analyzing quaternary alkaloids in coptis-scute herb couple by electrospray ionization mass spectrometry(ESI-MS) was developed. Without the complicated pretreatment of sample, the active ingredients including berberine, palmatine, coptisine, jatrorrhizine, epiberberine, and columbamine were identified and some relative content changing rules of alkaloids in coptis-scute couple were summarized in this article. The overall profiles of the complex extracts were obtained.

New method of mass spectrometry for distinguishing ephedrine and isoephedrine

Ephedrine and isoephedrine were first distingshed by electrospy ionization mass spectrometry and in-sourice collision-induced dissociation technique. Based on this observation, a unkown sample was identified for ephedrine.

The application of matrix-assisted laser desorption jonization mass spectrometry in enzymes of Agkistrodon blomhoffii Ussurensis venom analysis

The matrix-assisted laser desorption ionization mass spectrometry (MALDI-TOF-MS) spectra of four enzymes (PLA, AEase, Fibrolase, L-a.a. oxidase) in Agkistrodon blomhoffii Ussurensis venom, were given and interpreted. The experiment data showed that MALDI-TOF-MS can be used directly in enzyme analysis with high sensitivity and rapidity. In addition, the results were better than those from sodium dodecyl sulfate-polyacrylamide.

Identification of phospholipid structures in human blood by direct-injection quadrupole-linear ion-trap mass spectrometry

Direct-injection electrospray ionization mass spectrometry in combination with information-dependent data acquisition (IDA), using a triple-quadrupole/linear ion trap combination, allows high-throughput qualitative analysis of complex phospholipid species from child whole blood. In the IDA experiments, scans to detect specific head groups (precursor ion or neutral loss scans) were used as survey scans to detect phospholipid classes. An enhanced resolution scan was then used to confirm the mass assignments, and the enhanced product ion scan was implemented as a dependent scan to determine the composition of each phospholipid class. These survey and dependent scans were performed sequentially and repeated for the entire duration of analysis, thus providing the maximum information from a single injection. In this way, 50 different phospholipids belonging to the phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, phosphatidylcholine and sphingomyelin classes were identified in child whole blood. Copyright (C) 2005 John Wiley & Sons, Ltd.

Indirect hydrogen analysis by gas chromatography coupled to mass spectrometry (GC-MS).

Gas chromatography (GC) is an analytical tool very useful to investigate the composition of gaseous mixtures. The different gases are separated by specific columns but, if hydrogen (H2 ) is present in the sample, its detection can be performed by a thermal conductivity detector or a helium ionization detector. Indeed, coupled to GC, no other detector can perform this detection except the expensive atomic emission detector. Based on the detection and analysis of H2 isotopes by low-pressure chemical ionization mass spectrometry (MS), a new method for H2 detection by GC coupled to MS with an electron ionization ion source and a quadrupole analyser is presented. The presence of H2 in a gaseous mixture could easily be put in evidence by the monitoring of the molecular ion of the protonated carrier gas. Copyright © 2013 John Wiley & Sons, Ltd.

Gas chromatographic electron capture negative ionization mass spectrometric (GC/ECNI/MS) determination of unique fluorinated compounds in the sediments of Lake Ontario and the effect of high-boiling alcohols (as injection solvents) on chromatographic behaviour of polycyclic aromatic hydrocarbons in gas chromatography

Part I - Fluorinated Compounds A method has been developed for the extraction, concentration, and determination of two unique fluorinated compounds from the sediments of Lake Ontario. These compounds originated from a common industrial landfill, and have been carried to Lake Ontario by the Niagara River. Sediment samples from the Mississauga basin of Lake Ontario have been evaluated for these compounds and a depositional trend was established. The sediments were extracted by accelerated solvent extraction (ASE) and then underwent clean-up, fractionation, solvent exchange, and were concentrated by reduction under nitrogen gas. The concentrated extracts were analyzed by gas chromatography - electron capture negative ionization - mass spectrometry. The depositional profile determined here is reflective of the operation of the landfill and shows that these compounds are still found at concentrations well above background levels. These increased levels have been attributed to physical disturbances of previously deposited contaminated sediments, and probable continued leaching from the dumpsite. Part II - Polycyclic Aromatic Hydrocarbons Gas chromatography/mass spectrometry is the most common method for the determination of polycyclic aromatic hydrocarbons (PAHs) from various matrices. Mass discrimination of high-boiling compounds in gas chromatographic methods is well known. The use of high-boiling injection solvents shows substantial increase in the response of late-eluting peaks. These solvents have an increased efficiently in the transfer of solutes from the injector to the analytical column. The effect of I-butanol, I-pentanol, cyclopentanol, I-hexanol, toluene and n-octane, as injection solvents, was studied. Higher-boiling solvents yield increased response for all PAHs. I -Hexanol is the best solvent, in terms of P AH response, but in this solvent P AHs were more susceptible to chromatographic problems such as peak splitting and tailing. Toluene was found to be the most forgiving solvent in terms of peak symmetry and response. It offered the smallest discrepancies in response, and symmetry over a wide range of initial column temperatures.

Single embryo and oocyte lipid fingerprinting by mass spectrometry

Methods used for lipid analysis in embryos and oocytes usually involve selective lipid extraction from a pool of many samples followed by chemical manipulation, separation and characterization of individual components by chromatographic techniques. Herein we report direct analysis by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of single and intact embryos or oocytes from various species. Biological samples were simply moisturized with the matrix solution and characteristic lipid ( represented by phosphatidylcholines, sphingomyelins and triacylglycerols) profiles were obtained via MALDI-MS. As representative examples, human, bovine, sheep and fish oocytes, as well as bovine and insect embryos were analyzed. MALDI-MS is shown to be capable of providing characteristic lipid profiles of gametes and embryos and also to respond to modifications due to developmental stages and in vitro culture conditions of bovine embryos. Investigation in developmental biology of the biological roles of structural and reserve lipids in embryos and oocytes should therefore benefit from these rapid MALDI-MS profiles from single and intact species.-Ferreira, C. R., S. A. Saraiva, R. R. Catharino, J. S. Garcia, F. C. Gozzo, G. B. Sanvido, L. F. A. Santos, E. G. Lo Turco, J. H. F. Pontes, A. C. Basso, R. P. Bertolla, R. Sartori, M. M. Guardieiro, F. Perecin, F. V. Meirelles, J. R. Sangalli, and M. N. Eberlin. Single embryo and oocyte lipid fingerprinting by mass spectrometry. J. Lipid Res. 2010. 51: 1218-1227.

Optimal single-embryo mass spectrometry fingerprinting

In pre-implantation embryos, lipids play key roles in determining viability, cryopreservation and implantation properties, but often their analysis is analytically challenging because of the few picograms of analytes present in each of them. Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) allows obtaining individual phospholipid profiles of these microscopic organisms. This technique is sensitive enough to enable analysis of individual intact embryos and monitoring the changes in membrane lipid composition in the early stages of development serving as screening method for studies of biology and biotechnologies of reproduction. This article introduces an improved, more comprehensive MALDI-MS lipid fingerprinting approach that considerably increases the lipid information obtained from a single embryo. Using bovine embryos as a biological model, we have also tested optimal sample storage and handling conditions before the MALDI-MS analysis. Improved information at the molecular level is provided by the use of a binary matrix that enables phosphatidylcholines, sphingomyelins, phosphatidylserines, phosphatidylinositols and phosphoethanolamines to be detected via MALDI(±)-MS in both the positive and negative ion modes. An optimal MALDI-MS protocol for lipidomic monitoring of a single intact embryo is therefore reported with potential applications in human and animal reproduction, cell development and stem cell research. Copyright © 2013 John Wiley & Sons, Ltd. Copyright © 2013 John Wiley & Sons, Ltd.

Membrane lipid profile monitored by mass spectrometry detected differences between fresh and vitrified in vitro-produced bovine embryos

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Mass spectrometry study of N-alkylbenzenesulfonamides with potential antagonist activity to potassium channels

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

STUDY BY MASS SPECTROMETRY OF SOLUTIONS OF [HYDROXY(TOSYLOXY)IODO]BENZENE: PROPOSED DISPROPORTIONATION MECHANISMS

STUDY BY MASS SPECTROMETRY OF SOLUTIONS OF [HYDROXY(TOSYLOXY)IODO]BENZENE: PROPOSED DISPROPORTIONATION MECHANISMS. Solutions of [hydroxy(tosyloxy)iodo]benzene (HTIB or Koser's reagent) in acetonitrile were analyzed using high resolution electrospray ionization mass spectrometry (ESI-MS) and electrospray ionization tandem mass spectrometry (ESI-MS/MS) under different conditions. Several species were characterized in these analyses. Based on these data, mechanisms were proposed for the disproportionation of the iodine(III) compounds in iodine(V) and iodine(I) species.

From MALDI-TOF mass spectrometry to soft-landing for electronics

Within this PhD thesis matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) has been used as a reliable tool for the quantitative characterization of giant molecules, such as alkyl substituted and unsubstituted large polycyclic aromatic hydrocarbons (PAH), which cannot be characterized by conventional analytic techniques due to their lack of solubility. The use of the MALDI solvent-free technique for the sample preparation and the application of the standard addition method have allowed the quantitative characterization of synthetic PAH mixtures. The knowledge, acquired by studying these representative systems, has been then transferred to the quantitative analyses of complex and slightly soluble natural PAH mixtures, such as mesophase pitch. Moreover, the possibility to ionize intractable and insoluble molecules via mass spectrometry has been recognized to be not only a powerful analytical method, but also to represent a unique change to handle giant aromatic systems and to deposit them on a surface for further investigations, in a process, which is defined as “soft-landing”. Within this novel deposition technique, ions of the desired analytes or analyte mixtures are generated by means of an MS ionization source, discriminated by their different mass to charge ratios via a mass analyzer and landed with retention of their structure on a desired surface. This soft-deposition is guaranteed by the use of decelerating potentials, which have in this work been recognized to influence the final packing of the analyte molecules reaching the landing surface. For a more detailed study of the electrical field action on disc-like and rod-like molecules, soft-landing-independent experiments have been additionally carried out. As a result unidirectionally ordered films of the analyte molecules have been obtained due to the application of an external electrical strength. This versatile alignment technique has then been used for obtaining ordered layers of semiconducting materials for the fabrication of organic field effect transistors (OFET) with improved performances.

Mass spectrometry-based analytical tools for the molecular protein characterization of human plasma lipoproteins

Lipoproteins are a heterogeneous population of blood plasma particles composed of apolipoproteins and lipids. Lipoproteins transport exogenous and endogenous triglycerides and cholesterol from sites of absorption and formation to sites of storage and usage. Three major classes of lipoproteins are distinguished according to their density: high-density (HDL), low-density (LDL) and very low-density lipoproteins (VLDL). While HDLs contain mainly apolipoproteins of lower molecular weight, the two other classes contain apolipoprotein B and apolipoprotein (a) together with triglycerides and cholesterol. HDL concentrations were found to be inversely related to coronary heart disease and LDL/VLDL concentrations directly related. Although many studies have been published in this area, few have concentrated on the exact protein composition of lipoprotein particles. Lipoproteins were separated by density gradient ultracentrifugation into different subclasses. Native gel electrophoresis revealed different gel migration behaviour of the particles, with less dense particles having higher apparent hydrodynamic radii than denser particles. Apolipoprotein composition profiles were measured by matrix-assisted laser desorption/ionization-mass spectrometry on a macromizer instrument, equipped with the recently introduced cryodetector technology, and revealed differences in apolipoprotein composition between HDL subclasses. By combining these profiles with protein identifications from native and denaturing polyacrylamide gels by liquid chromatography-tandem mass spectrometry, we characterized comprehensively the exact protein composition of different lipoprotein particles. We concluded that the differential display of protein weight information acquired by macromizer mass spectrometry is an excellent tool for revealing structural variations of different lipoprotein particles, and hence the foundation is laid for the screening of cardiovascular disease risk factors associated with lipoproteins.