Characterization of Pinus spp. needles by gas chromatography and mass spectrometry: Application to plant-insect interactions

Dissertação apresentada para obtenção do Grau de Doutor em Ciências do Ambiente pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecn

Valorização das águas corticeiras: extracção de antioxidantes

Dissertação para obtenção do Grau de Mestre em Engenharia Química e Bioquímica

Time-of-flight secondary ion mass spectrometry: new application for urinary stones analysis

Dissertação para obtenção do Grau de Doutor em Engenharia Física

Advances in microbial diagnosis: using mass spectrometry for proteomics and genomics applications

Since the last two decades mass spectrometry (MS) has been applied to analyse the chemical cellular components of microorganisms, providing rapid and discriminatory proteomic profiles for their species identification and, in some cases, subtyping. The application of MS for the microbial diagnosis is currently well-established. The remarkable reproducibility and objectivity of this method is based on the measurement of constantly expressed and highly abundant proteins, mainly important conservative ribosomal proteins, which are used as markers to generate a cellular fingerprint. Mass spectrometry based on matrix-assisted laser desorption ionization-time of flight (MALDI- TOF) technique has been an important tool for the microbial diagnostic. However, some technical limitation concerning both MALDI-TOF and its used protocols for sample preparation have fostered the research of new mass spectrometry systems (e.g. LC MS/MS). LC MS/MS is able to generate online mass spectra of specific ions with further online sequencing of these ions, which include both specific proteins and DNA fragments. In this work a set of data for yeasts and filamentous fungi diagnostic obtained through an international collaboration project involving partners from Argentina, Brazil, Chile and Portugal will be presented and discussed.

Aromatic amines sources, environmental impact and remediation

Aromatic amines are widely used industrial chemicals as their major sources in the environment include several chemical industry sectors such as oil refining, synthetic polymers, dyes, adhesives, rubbers, perfume, pharmaceuticals, pesticides and explosives. They result also from diesel exhaust, combustion of wood chips and rubber and tobacco smoke. Some types of aromatic amines are generated during cooking, special grilled meat and fish, as well. The intensive use and production of these compounds explains its occurrence in the environment such as in air, water and soil, thereby creating a potential for human exposure. Since aromatic amines are potential carcinogenic and toxic agents, they constitute an important class of environmental pollutants of enormous concern, which efficient removal is a crucial task for researchers, so several methods have been investigated and applied. In this chapter the types and general properties of aromatic amine compounds are reviewed. As aromatic amines are continuously entering the environment from various sources and have been designated as high priority pollutants, their presence in the environment must be monitored at concentration levels lower than 30 mg L1, compatible with the limits allowed by the regulations. Consequently, most relevant analytical methods to detect the aromatic amines composition in environmental matrices, and for monitoring their degradation, are essential and will be presented. Those include Spectroscopy, namely UV/visible and Fourier Transform Infrared Spectroscopy (FTIR); Chromatography, in particular Thin Layer (TLC), High Performance Liquid (HPLC) and Gas chromatography (GC); Capillary electrophoresis (CE); Mass spectrometry (MS) and combination of different methods including GC-MS, HPLC-MS and CE-MS. Choosing the best methods depend on their availability, costs, detection limit and sample concentration, which sometimes need to be concentrate or pretreated. However, combined methods may give more complete results based on the complementary information. The environmental impact, toxicity and carcinogenicity of many aromatic amines have been reported and are emphasized in this chapter too. Lately, the conventional aromatic amines degradation and the alternative biodegradation processes are highlighted. Parameters affecting biodegradation, role of different electron acceptors in aerobic and anaerobic biodegradation and kinetics are discussed. Conventional processes including extraction, adsorption onto activated carbon, chemical oxidation, advanced oxidation, electrochemical techniques and irradiation suffer from drawbacks including high costs, formation of hazardous by-products and low efficiency. Biological processes, taking advantage of the naturally processes occurring in environment, have been developed and tested, proved as an economic, energy efficient and environmentally feasible alternative. Aerobic biodegradation is one of the most promising techniques for aromatic amines remediation, but has the drawback of aromatic amines autooxidation once they are exposed to oxygen, instead of their degradation. Higher costs, especially due to power consumption for aeration, can also limit its application. Anaerobic degradation technology is the novel path for treatment of a wide variety of aromatic amines, including industrial wastewater, and will be discussed. However, some are difficult to degrade under anaerobic conditions and, thus, other electron acceptors such as nitrate, iron, sulphate, manganese and carbonate have, alternatively, been tested.

Análise de fragmentos arqueológicos: um diálogo entre a Química, Biologia e Arqueologia

Dissertação de mestrado em Técnicas de Caracterização e Análise Química

Caracterização de materiais plásticos usados na indústria têxtil e de vestuário por FTIR

Dissertação de mestrado em Técnicas de Caracterização e Análises Químicas

Enzymatic hydrophobic modification of jute fibers via grafting to reinforce composites

Horseradish peroxidase (HRP)/H2O2 system catalyzes the free-radical polymerization of aromatic compounds such as lignins and gallate esters. In this work, dodecyl gallate (DG) was grafted onto the surfaces of lignin-rich jute fabrics by HRP-mediated oxidative polymerization with an aim to enhance the hydrophobicity of the fibers. The DG-grafted jute fibers and reaction products of their model compounds were characterized by matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS), attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), thermogravimetric analysis (TGA), and differential scanning calorimetry (DSC). The results clearly indicated the grafting of DG to the jute fiber by HRP. Furthermore, the hydrophobicity of jute fabrics was determined by measuring the wetting time and static contact angle. Compared to the control sample, the wetting time and static contact angle of the grated fabrics changed from ~1 s to 1 h and from ~0° to 123.68°, respectively. This clearly proved that the hydrophobicity of jute fabrics improved considerably. Conditions of the HRP-catalyzed DG-grafting reactions were optimized in terms of the DG content of modified jute fabrics. Moreover, the results of breaking strength and elongation of DG-grafted jute/ polypropylene (PP) composites demonstrated improved reinforcement of the composite due to enzymatic hydrophobic modification of jute fibers.

Liquid chromatography-tandem mass spectrometry (LC/APCI-MS/MS) methods for the quantification of captan and folpet phthalimide metabolites in human plasma and urine.

Captan and folpet are fungicides largely used in agriculture. They have similar chemical structures, except that folpet has an aromatic ring unlike captan. Their half-lives in blood are very short, given that they are readily broken down to tetrahydrophthalimide (THPI) and phthalimide (PI), respectively. Few authors measured these biomarkers in plasma or urine, and analysis was conducted either by gas chromatography coupled to mass spectrometry or liquid chromatography with UV detection. The objective of this study was thus to develop simple, sensitive and specific liquid chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry (LC/APCI-MS/MS) methods to quantify both THPI and PI in human plasma and urine. Briefly, deuterated THPI was added as an internal standard and purification was performed by solid-phase extraction followed by LC/APCI-MS/MS analysis in negative ion mode for both compounds. Validation of the methods was conducted using spiked blank plasma and urine samples at concentrations ranging from 1 to 250 μg/L and 1 to 50 μg/L, respectively, along with samples of volunteers and workers exposed to captan or folpet. The methods showed a good linearity (R (2) > 0.99), recovery (on average 90% for THPI and 75% for PI), intra- and inter-day precision (RSD, <15%) and accuracy (<20%), and stability. The limit of detection was 0.58 μg/L in urine and 1.47 μg/L in plasma for THPI and 1.14 and 2.17 μg/L, respectively, for PI. The described methods proved to be accurate and suitable to determine the toxicokinetics of both metabolites in human plasma and urine.

Ultra high performance supercritical fluid chromatography coupled with tandem mass spectrometry for screening of doping agents. II: Analysis of biological samples.

The potential and applicability of UHPSFC-MS/MS for anti-doping screening in urine samples were tested for the first time. For this purpose, a group of 110 doping agents with diverse physicochemical properties was analyzed using two separation techniques, namely UHPLC-MS/MS and UHPSFC-MS/MS in both ESI+ and ESI- modes. The two approaches were compared in terms of selectivity, sensitivity, linearity and matrix effects. As expected, very diverse retentions and selectivities were obtained in UHPLC and UHPSFC, proving a good complementarity of these analytical strategies. In both conditions, acceptable peak shapes and MS detection capabilities were obtained within 7min analysis time, enabling the application of these two methods for screening purposes. Method sensitivity was found comparable for 46% of tested compounds, while higher sensitivity was observed for 21% of tested compounds in UHPLC-MS/MS and for 32% in UHPSFC-MS/MS. The latter demonstrated a lower susceptibility to matrix effects, which were mostly observed as signal suppression. In the case of UHPLC-MS/MS, more serious matrix effects were observed, leading typically to signal enhancement and the matrix effect was also concentration dependent, i.e., more significant matrix effects occurred at the lowest concentrations.

Performance of matrix-assisted laser desorption ionization-time of flight mass spectrometry for identification of bacterial strains routinely isolated in a clinical microbiology laboratory.

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has recently been introduced in diagnostic microbiology laboratories for the identification of bacterial and yeast strains isolated from clinical samples. In the present study, we prospectively compared MALDI-TOF MS to the conventional phenotypic method for the identification of routine isolates. Colonies were analyzed by MALDI-TOF MS either by direct deposition on the target plate or after a formic acid-acetonitrile extraction step if no valid result was initially obtained. Among 1,371 isolates identified by conventional methods, 1,278 (93.2%) were putatively identified to the species level by MALDI-TOF MS and 73 (5.3%) were identified to the genus level, but no reliable identification was obtained for 20 (1.5%). Among the 1,278 isolates identified to the species level by MALDI-TOF MS, 63 (4.9%) discordant results were initially identified. Most discordant results (42/63) were due to systematic database-related taxonomical differences, 14 were explained by poor discrimination of the MALDI-TOF MS spectra obtained, and 7 were due to errors in the initial conventional identification. An extraction step was required to obtain a valid MALDI-TOF MS identification for 25.6% of the 1,278 valid isolates. In conclusion, our results show that MALDI-TOF MS is a fast and reliable technique which has the potential to replace conventional phenotypic identification for most bacterial strains routinely isolated in clinical microbiology laboratories.

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.

Validation of an analytical method for nitrous oxide (N2O) laughing gas by headspace gas chromatography coupled to mass spectrometry (HS-GC-MS): Forensic application to a lethal intoxication.

Drug abuse is a widespread problem affecting both teenagers and adults. Nitrous oxide is becoming increasingly popular as an inhalation drug, causing harmful neurological and hematological effects. Some gas chromatography-mass spectrometry (GC-MS) methods for nitrous oxide measurement have been previously described. The main drawbacks of these methods include a lack of sensitivity for forensic applications; including an inability to quantitatively determine the concentration of gas present. The following study provides a validated method using HS-GC-MS which incorporates hydrogen sulfide as a suitable internal standard allowing the quantification of nitrous oxide. Upon analysis, sample and internal standard have similar retention times and are eluted quickly from the molecular sieve 5Å PLOT capillary column and the Porabond Q column therefore providing rapid data collection whilst preserving well defined peaks. After validation, the method has been applied to a real case of N2O intoxication indicating concentrations in a mono-intoxication.

Quantification of clenbuterol at trace level in human urine by ultra-high pressure liquid chromatography-tandem mass spectrometry.

Clenbuterol is a β2 agonist agent with anabolic properties given by the increase in the muscular mass in parallel to the decrease of the body fat. For this reason, the use of clenbuterol is forbidden by the World Anti-Doping Agency (WADA) in the practice of sport. This compound is of particular interest for anti-doping authorities and WADA-accredited laboratories due to the recent reporting of risk of unintentional doping following the eating of meat contaminated with traces of clenbuterol in some countries. In this work, the development and the validation of an ultra-high pressure liquid chromatography coupled to electrospray ionization tandem mass spectrometry (UHPLC-ESI-MS/MS) method for the quantification of clenbuterol in human urine is described. The analyte was extracted from urine samples by liquid-liquid extraction (LLE) in basic conditions using tert butyl-methyl ether (TBME) and analyzed by UHPLC-MS/MS with a linear gradient of acetonitrile in 9min only. The simple and rapid method presented here was validated in compliance with authority guidelines and showed a limit of quantification at 5pg/mL and a linearity range from 5pg/mL to 300pg/mL. Good trueness (85.8-105%), repeatability (5.7-10.6% RSD) and intermediate precision (5.9-14.9% RSD) results were obtained. The method was then applied to real samples from eighteen volunteers collecting urines after single oral doses administration (1, 5 and 10μg) of clenbuterol-enriched yogurts.