Biblioteca Digital

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq); High-performance liquid chromatography/electrospray ionization mass spectrometry (HPLC/ESI-MS) and high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) techniques were applied for the detection, purification, monitoring, and sequencing of two novel and biologically active peptides occurring at very low levels in the venom of the wasp Agelaia pallipes pallipes. These peptides were sequenced under LC/ESI-MS/MS conditions and designated as Agelaia-CP (I/L-L-G-T-I-L-G-L-L-K-G-I/L-NH2, MW 1207.8Da) and Agelaia-MP (I/L-N-W-L-K-L-G-K-A-I-I-D-A-I/L-NH2, MW 1565.0Da). The peptide Agelaia-CP showed no hemolytic activity, but it behaved as a mast cell degranulator and induced a potent chemotaxis in polymorphonucleated leukocyte (PMNL) cells, typical of a wasp chemotactic peptide. The peptide Agelaia-MP showed both powerful mast cell degranulation and hemolysis of washed rat red blood cells, and is thus assigned as a new member of the mastoparan family of peptides. Both peptides seem to be directly involved in the strong inflammatory reactions associated with wasp stings. © Copyright 2004 John Wiley & Sons, Ltd.

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...

Este trabalho direcionou-se ao controle de qualidade de óleos vegetais e gorduras animais pelo emprego de análise de baixa massa molecular pela técnica EASI-MS utilizando a ferramenta metabolômica. As matérias-primas graxas (óleos e gorduras vegetais e animais) e os produtos sintetizados a partir destes insumos são misturas complexas, com aplicações e valores agregados variados como por exemplo para a nutrição humana, aplicações industriais na produção de lubrificantes, biodiesel, plasticidas, surfactantes, entre outros. As técnicas disponíveis como Cromatografia gasosa (GC) com detector de ionização de chama (FID) ou acoplada a um espectrômetro de massas (GC-MS) tem sido as técnicas mais utilizadas para caracterizar óleos e gorduras, através da determinação da composição de ácidos graxos. Como a caracterização da composição graxa destas matrizes são muitas vezes restritas, exigindo a necessidade de procedimentos laboriosos como extrações, purificações que requerem muito tempo utilizamos neste trabalho uma recente técnica de ionização ambiente de espectrometria de massas: EASI-MS (easy ambient sonic-spray ionization mass spectrometry) na caracterização de óleos vegetais e gorduras animais que dispensa o emprego de processos de derivatização química e a separação cromatográfica. Neste trabalho aplicamos a metabolômica para elucidação de um conjugado de marcadores taxonômicos de óleos nunca observados ou analisados em conjunto antes...

Embora seja um tema que desperte bastante interesse perante a sociedade científica, a aplicação da química no campo da criminalística ainda constitui uma nova linha de pesquisa no Brasil. Neste trabalho, o perfil químico de várias drogas de abuso (anfetaminas, piperazinas, cocaína e dietilamida do ácido lisérgico (LSD)), documentos e derivados de petróleo foram investigados. Para análise de drogas de abuso, a easy ambient sonic-spray ionization (EASI-MS) é uma poderosa ferramenta na caracterização de amostras na sua forma original, como é o caso de amostras de ecstasy e LSD, que são vendidos como comprimidos e selos, respectivamente. A espectrometria de massas foi utilizada também para estudar a mobilidade iônica de isômeros da clorofenilpiperazina (o-CPP, m-CPP e p-CPP), onde o m-CPP se enquadra na lista de substâncias proscritas. Foi também demonstrado que a cromatografia em camada delgada (CCD) continua sendo uma técnica confiável para a identificação do 3,4-metilenodimetoxianfetamina (MDMA) e outros adulterantes em comprimidos de ecstasy, mas ela pode fornecer resultados falso-negativos. A associação da CCD com EASI-MS potencializa a identificação de todas as anfetaminas que poderiam ser usadas na fabricação de drogas de rua...

Com o objetivo de entender as propriedades intrínsecas dos líquidos iônicos imidazólios, foram desenvolvidos trabalhos de investigação dos mecanismos de dissociação e destilação, força intrínseca de ligações de hidrogênio e formação e caracterização de carbenos a partir destes compostos. Tais estudos foram realizados através da espectrometria de massas com técnicas API (Atmospheric Pressure Ionization) como ESI-MS(/MS) (Eletrospray Ionization Mass Spectrometry) e APCI-MS(/MS) (Atmospheric Pressure Chemical Ionization Mass Spectrometry) que são técincas de ionização branda que permitem a análise dos compostos por infusão direta da solução de interesse. Para estes estudos foram escolhidos cátions imidazólios com diferentes substituintes nas posições N-1, N-3 e C-2, além de diferentes ânions como contra-íons. O objetivo do trabalho foi testar como essas modificações na cadeia iriam afetar as porpriedades intrínsecas estudadas. A partir dos resultados obtidos nos trabalhos realizados foi possível propor mecanismos de fragmentação para uma vasta gama de líquidos iônicos imidazólios, e propor um mecanismo de destilação para alguns destes compostos. Foram realizadas medidas onde foi possível ordenar e medir a magnitude intrínseca das ligações de hidrogênio para diferentes cátions. Além disso foram obtidas informações a respeito da formação e caracterização de carbenos a partir de líquidos iônicos di...

O petróleo é uma mistura complexa, constituída de milhares de compostos e sua caracterização é fundamental em diversos processos da indústria petrolífera. A abordagem moderna para a caracterização total dos compostos polares (N, O, S) do petróleo é através da petroleômica, utilizando a espectrometria de massas de altíssima resolução e exatidão obtida em espectrômetros de ressonância ciclotrônica de íons com análise por transformada de Fourier (FT-ICR MS). FT-ICR MS é o equipamento ideal para estas análises, no entanto a literatura carece de informações quanto a otimização do número de scans e do efeito da resolução na análise petroleômica. Determinamos como condições ideais para a análise destas amostras de altíssima complexidade a obtenção de espectros de MS com poder de resolução de 400.000 em m/z 400 e acúmulo de 100 mscans. Foram analisados também os compostos polares presentes em outras fontes de combustíveis fósseis, como carvão mineral e folhelho. Os resultados mostraram padrões distintos em relação à origem do material e possivelmente ao grau de evolução térmica. Comparamos a análise por FT-ICR MS dos ácidos carboxílicos de amostras de petróleo com os resultados das análise de frações ácidas por FAB MS obtidas no CENPES . PETROBRAS...

Essa dissertação de Mestrado visa estudar o mecanismo da reação de Morita-Baylis-Hillman (MBH) em três versões (Clássica, Aza e Assimétrica) utilizando a técnica de espectrometria de massas com ionização por eletrospray (ESI-MS). Para o monitoramento mecanístico das versões clássica e Aza (MBH/Aza-MBH) foi introduzida uma nova abordagem, que é a utilização de reagentes marcados com etiquetas de carga (líquidos iônicos), a fim de facilitar a detecção dos intermediários e maximizar a aquisição de dados obtidos pelo monitoramento destas reações. Entre os reagentes de partida, (eletrófilo, catalisador e alceno ativado) o alceno ativado foi o reagente marcado, utilizou-se o íon metil imidazólio como etiqueta de carga. Além do uso de um reagente marcado, ambas as versões foram monitoradas utilizando reagentes neutros, com o propósito de avaliar em nível de comparação as metodologias abordadas, marcada e não-marcada. Para versão assimétrica da reação de MBH, foram utilizadas duas aminas hidroxiladas quirais derivadas de alcalóides cinchona, quinidina e β-isocupreidina. O monitoramento mecanístico da versão assimétrica foi realizado utilizando um espectrômetro de massas com mobilidade iônica IM-MS. O tipo de mobilidade utilizada nesse estudo foi a Traveling Wave...

Proceedings of the Chemistry and Conservation Science; In the last decade the great development of mass spectrometry techniques made them ideal tools for the characterization of many materials containing either inorganic or organic compounds. Pigments in paints, main constituents of glass and ceramic objects, enamels and glazes can be characterized by inorganic mass spectrometry. Temperas, varnishes and adhesives can be studied by organic mass spectrometry; compounds as glycerides, proteins and sugars can also be easily analysed. Due to the diversity of mass spectrometry techniques almost any compound has a special method that can be used to characterize it. A brief introduction to mass spectrometry will be given, in particular a reference to the ion generation methods and their application to the different types of samples that a conservator can have in his/her hands when studying an artwork. A few applications in cultural heritage will be given.

In this paper, we report the use of mass spectrometry imaging and structural analysis of lipids directly on a tissue specimen, carried out by means of matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry, using a combination of spiral orbit-type and reflectron-type time-of-flight mass spectrometers. The most intense peak observed in the mass spectrum from a brain tissue specimen was confirmed as phosphatidylcholine (34 : 1) [M+K]+, using tandem mass spectrometry. The charge remote fragmentation channels, which are characteristically observed using high-energy collision-induced dissociation, contributed significantly to this confirmation. Accurate mass analysis was further facilitated by mass correction using the confirmed peak. In mass spectrometry imaging, the high resolving power of our system could separate doublet peak of less than 0.1 u difference, which would otherwise be problematic when using a low-resolution reflectron type time-of-flight mass spectrometer. Two compounds, observed at m/z 848.56 and 848.65, were found to be located in complementary positions on a brain tissue specimen. These results demonstrate the importance of a high-performance tandem time-of-flight mass spectrometer for mass spectrometry imaging and analysis of observed compounds...

The paper has three parts, (i) a brief overview of the main achievements made using mass spectrometry across all the fields of science, (ii) a survey of some of the topics currently being pursued most activity, including both applications and fundamental studies, and (iii) some hints as to what the future of mass spectrometry might hold with particular emphasis on revolutionary changes in the subject. Emphasis is given to ambient methods of ionization and their use in disease diagnosis and to their use in combination with miniature mass spectrometers for in-situ measurements. Special attention goes to the chemical aspects of mass spectrometry, including its emerging role as a preparative method based on accelerated bimolecular reaction rates in solution and on ion soft landing as a means of surface tailoring. In summary, the paper covers the proud history, vibrant present and expansive future of mass spectrometry.

Mass spectrometry-based investigation of clinical samples enables the high-throughput identification of protein biomarkers. We provide an overview of mass spectrometry-based proteomic techniques that are applicable to the investigation of clinical samples. We address sample collection, protein extraction and fractionation, mass spectrometry modalities, and quantitative proteomics. Finally, we examine the limitations and further potential of such technologies. Liquid chromatography fractionation coupled with tandem mass spectrometry is well suited to handle mixtures of hundreds or thousands of proteins. Mass spectrometry-based proteome elucidation can reveal potential biomarkers and aid in the development of hypotheses for downstream investigation of the molecular mechanisms of disease.

Tremendous technology improvements of the last decades has given mass spectrometry a more and more expanding role in the study of a wide range of molecules: from the identification and quantification of small molecular weight molecules to the structural determination of biomacromolecules. Many are the fields of application for this technique and the various versions of it. In the present study three different applications have been explored. The first application is a pharmacokinetics study of anticancer drug Gemcitabine and its principal metabolite, where the role of the LC-MS/MS is essential both for the selectivity of the detection of the small analytes and the sensitivity enhanced by multi-reaction monitoring experiments. The design of the study involved the collection of several blood samples at selected times and from patients that would have met certain eligibility criteria. The ESI demonstrated to be the most suitable approach and it provided the necessary data to conclude that toxicity of Gemcitabine did not increase when administered at FDR (Fixed Dose Rate) infusion in patients with impaired hepatic function. The second application describes an example of how MS represents a powerful tool in cancer research, from serum profiling study with high resolution MALDITOF and bioinformatic analysis...

Mass spectrometry coupled to liquid chromatography (LC-MS) is an analytical technique becoming increasingly popular in biomedical research. Especially in high-throughput proteomics and metabolomics mass spectrometry is widely used because it provides both qualitative and quantitative information about analytes. The standard protocol is that complex analyte mixtures are first separated in liquid chromatography and then analyzed using mass spectrometry. Finally, computational tools extract all relevant information from the large amounts of data produced. This thesis aims at improving computational analysis of LC-MS data|we present two novel computational methods and a software framework for the development of LC-MS data analysis tools. In the first part of this thesis we present a quantitation algorithm for peptide signals in isotope-resolved LC-MS data. Exact quantitation of all peptide signals (so-called peptide features) is an essential step in most LC-MS data analysis pipelines. Our algorithm detects and quantifies peptide features in centroided peak maps using a multi-phase approach: First, putative feature centroid peaks, so-called seeds, are determined based on signal properties that are typical for peptide features. In the second phase...

The aim of this study was to investigate the utility of direct analysis in real time quadrupole time-of-flight mass spectrometry and gas chromatography quadrupole time-of-flight mass spectrometry in the analysis of 162 different synthetic cannabinoids. Direct analysis in real time quadrupole time-of-flight mass spectrometry is shown to be a rapid and accurate analytical method for synthetic cannabinoids. Spectra can be generated with less than 1.5 ng of the drug in under a minute and be successfully searched against previously generated ESI-QTOF libraries in most cases (118/130 drugs tested) as well as can also be applied to the identification of synthetic cannabinoids in a mixture. Gas chromatography quadrupole time-of-flight mass spectrometry, while requiring a much longer analysis time, is shown to accurately distinguish all but 19 compounds (140/159). These two instruments have proven to be viable alternatives in synthetic cannabinoid analysis and will greatly benefit forensic laboratories.

11 pages, 10 figures.-- PMID: 16841894 [PubMed].-- Printed version published Jul 15, 2006.; Mass spectrometry has recently become one of the major analytical tools to study biomolecular structure and function. Ionization techniques, such as electrospray ionization (ESI), desorb biomolecules from solution to the gas phase keeping practically intact their natural structure. ESI applied to a protein solution produces a mixture of multiply charged ions, the ion charge distribution of which depends on the oligomeric form (mass) and on the protein surface exposed (amount of accommodated charges) of the related protein conformation. ESI-MS provides an efficient way to monitor protein processes; however, the ionic contributions of the different protein conformations involved usually overlap, and the use of chemometric tools is necessary to unravel the information related to the pure conformations that the biomolecule adopts along the process. Multivariate curve resolution-alternating least squares applied to MS-monitored protein processes provides the concentration profiles associated with the different protein conformations occurring during the process and the related pure mass spectra. The concentration profiles, in this context, the ionic contributions...

10 pages, 5 tables.-- PMID: 17540393 [PubMed].-- Printed version published on Aug 10, 2207.-- Issue title: 23rd Montreux Symposium on Liquid Chromatography-Mass Spectrometry, Supercritical Fluid Chromatography-Mass Spectrometry, Capillary Electrophoresis-Mass Spectrometry and Tandem Mass Spectrometry (Montreaux, Switzerland, Nov 8-10, 2006).; Presented at the 2nd International Workshop on Liquid-Chromatography-Tandem Mass Spectrometry for Screening and Trace Level Quantitation in Environmental and Food Samples, Barcelona, Spain, September 18–19, 2006.; àáEstrogens have been often identified as the major contributors to the endocrine-disrupting activity observed in environmental waters. However, their analysis in these, sometimes very complex, matrices is still challenging due to the very low detection limits and the selectivity required for their reliable determination at the very low concentrations at which they are physiologically active. In this work, a polyclonal enzyme-linked immunosorbent assay (ELISA) kit for 17-prueba-estradiol analysis, high-performance liquid chromatography-tandem mass spectrometry (HPLC–MS/MS) based on triple-quadrupole analyzer (QqQ), and a newly developed method based on ultra performance liquid chromatography–quadrupole time of flight mass spectrometry (UPLC–Q-TOF-MS) have been evaluated in terms of performance for the rapid screening...

12 pages, 5 tables, 6 figures.-- PMID: 18021786 [PubMed].-- Printed version published on Dec 28, 2007.; Ultra-high-performance liquid chromatography–quadrupole time-of-flight mass spectrometry (UPLC-QqTOF-MS) is an emerging technique offering more rapid and efficient separation, as well as the possibility to obtain accurate mass measurement and tandem mass spectrometry (MS/MS). This paper deals with the use of UPLC-QqTOF-MS to identify the pesticide residues present in complex pear extracts. Carbendazim, imazalil, and ethoxyquin were successfully identified because of the accurate mass determination of their protonated molecule and their major fragments in the product ion mass spectra. A few plastic and latex additives were also found, most of them probably coming from the packaging transfer to the fruits. The potential of the UPLC-QqTOF-MS and UPLC-QqTOF-MS/MS techniques as a quantification tool is also discussed taking imazalil as example. For quantification, calibration curves were linear over a dynamic range of 2 orders of magnitude, whereas higher calibration ranges are better adjusted to polynomial curves of second and third order. Quantification using different mass windows was also assessed. Accurate quantification required mass windows as wide as 20 mDa...

Generally low levels of plutonium in environmental samples, often combined with limited sample sizes, necessitate reliable low-level techniques for determination of Pu isotopes. Accelerator mass spectrometry (AMS) has proved to be a powerful method for measuring low-level Pu activity concentrations and Pu isotope ratios. Based on procedural blanks, detection limits for AMS were below 1fg Pu (equivalent to ca. 2μBq 139Pu), which can compete with both TIMS, high sensitivity ICP-MS, and certainly alpha-spectrometry, while showing less interference, memory and matrix effects as compared to routine ICP-MS techniques. In addition to low detection limits, the technique offers the advantage of giving information on Pu isotope ratios. Measurements of sediments collected from dumping sites at Novaya Zemlya showed deviation from global fallout 240Pu/239Pu ratios.

The small doubly-charged molecular anion SiF6 2- was studied by two distinct approaches, one experimental the other theoretical. The dianion was produced in the gas phase by sputtering a Li2 SiF6 specimen with Cs+ ions and was detected by means of accelerator mass spectrometry. The identification was via the Si29 F6 2- 19 isotopomer; it has an odd total mass and therefore the dianion shows up at a half-integral mass-to-charge ratio (M q=71.5 amu) in the mass spectrum, facilitating a positive identification. The flight time through the mass spectrometer of 10 μs establishes a lower limit with respect to the intrinsic lifetime of this species. Attempts to detect the SiF6 2- dianion also by secondary-ion mass spectrometry failed, but provided an upper limit in terms of its formation probability with respect to the F- ion of Si29 F6 2- 19 F-19 <2× 10-9. Furthermore, theoretical calculations of the photoelectron spectrum by means of the relativistic one-particle propagator predict considerable stability of the dianion against autodetachment. The first ionization potential of SiF6 2- was determined as 2.79 eV at the optimized bond length of 1.718 in the gas phase.