Incorporation of either molybdenum or tungsten into formate dehydrogenase from Desulfovibrio alaskensis NCIMB 13491; EPR assignment of the proximal iron-sulfur cluster to the pterin cofactor in formate dehydrogenases from sulfate-reducing bacteria

J Biol Inorg Chem (2004) 9: 145–151 DOI 10.1007/s00775-003-0506-z

The role of electron transfer in DNA building blocks: evaluation of strand breaks and their implications

Dissertação para obtenção do Grau de Doutor em Engenharia Física

Compact ion-source based on superionic rubidium silver iodide (RbAg4I5) solid electrolyte

Dissertação para obtenção do Grau de Mestre em Engenharia Física

Analysis of the interaction of polycyclic aromatic compounds in a model organism: integration of genotoxic and histopathological effects

Due to their toxicity, especially their carcinogenic potential, polycyclic aromatic hydrocarbons (PAHs) became priority pollutants in biomonitoring programmes and environmental policy, such as the European Water Framework Directive. The model substances tested in this study, namely benzo[b]fluoranthene (B[b]F), considered potentially carcinogenic to humans and an effector carcinogenic PAH to wildlife, and phenanthrene (Phe), deemed a non-carcinogenic PAH, are common PAHs in coastal waters, owning distinct properties reflected in different, albeit overlapping, mechanisms of toxicity. Still, as for similar PAHs, their interaction effects remain largely unknown. In order to study the genotoxic effects of caused by the interaction of carcinogenic and non-carcinogenic PAHs, and their relation to histopathological alterations, juvenile sea basses, Dicentrarchus labrax, a highly ecologically- and economically-relevant marine fish, were injected with different doses (5 and 10 μg.g-1 fish ww) of the two PAHs, isolated or in mixture, and incubated for 48 h. Individuals injected with B[b]F and the PAH mixture exhibited higher clastogenic/aneugenic effects and DNA strand breakage in blood cells, determined through the erythrocytic nuclear abnormalities (ENA) and Comet assays, respectively. Also, hepatic histopathological alterations were found in all animals, especially those injected with B[b]F and the PAH mixture, relating especially to inflammation. Still, Phe also exhibited genotoxic effects in sea bass, especially in higher doses, revealing a very significant acute effect that was accordant with the Microtox test performed undergone in parallel. Overall, sea bass was sensitive to B[b]F (a higher molecular weight PAH), likely due to efficient bioactivation of the pollutant (yielding genotoxic metabolites and reactive oxygen species), when compared to Phe, the latter revealing a more significant acute effect. The results indicate no significant additive effect between the substances, under the current experimental conditions. The present study highlights the importance of understanding PAH interactions in aquatic organisms, since they are usually present in the aquatic environment in complex mixtures.

Characterization of molecular damage induced by UV photons and carbon ions on biomimetic heterostructures

The study of the effect of radiation on living tissues is a rather complex task to address mainly because they are made of a set of complex functional biological structures and interfaces. Particularly if one is looking for where damage is taking place in a first stage and what are the underlying reaction mechanisms. In this work a new approach is addressed to study the effect of radiation by making use of well identified molecular hetero-structures samples which mimic the biological environment. These were obtained by assembling onto a solid support deoxyribonucleic acid (DNA) and phospholipids together with a soft water-containing polyelectrolyte precursor in layered structures and by producing lipid layers at liquid/air interface with DNA as subphase. The effects of both ultraviolet (UV) radiation and carbon ions beams were systematically investigated in these heterostructures, namely damage on DNA by means vacuum ultraviolet (VUV), infrared (IR), X-Ray Photoelectron (XPS) and impedance spectroscopy. Experimental results revealed that UV affects furanose, PO2-, thymines, cytosines and adenines groups. The XPS spectrometry carried out on the samples allowed validate the VUV and IR results and to conclude that ionized phosphate groups, surrounded by the sodium counterions, congregate hydration water molecules which play a role of UV protection. The ac electrical conductivity measurements revealed that the DNA electrical conduction is arising from DNA chain electron hopping between base-pairs and phosphate groups, with the hopping distance equal to the distance between DNA base-pairs and is strongly dependent on UV radiation exposure, due loss of phosphate groups. Characterization of DNA samples exposed to a 4 keV C3+ ions beam revealed also carbon-oxygen bonds break, phosphate groups damage and formation of new species. Results from radiation induced damage carried out on biomimetic heterostructures having different compositions revealed that damage is dependent on sample composition, with respect to functional targeted groups and extent of damage. Conversely, LbL films of 1,2-dipalmitoyl-sn-Glycero-3-[Phospho-rac-(1-glycerol)] (Sodium Salt) (DPPG) liposomes, alternated with poly(allylamine hydrochloride) (PAH) revealed to be unaffected, even by prolonged UV irradiation exposure, in the absence of water molecules. However, DPPG molecules were damaged by the UV radiation in presence of water with cleavage of C-O, C=O and –PO2- bonds. Finally, the study of DNA interaction with the ionic lipids at liquid/air interfaces revealed that electrical charge of the lipid influences the interaction of phospholipid with DNA. In the presence of DNA in the subphase, the effects from UV irrladiation were seen to be smaller, which means that ionic products from biomolecules degradation stabilize the intact DPPG molecules. This mechanism may explain why UV irradiation does not cause immediate cell collapse, thus providing time for the cellular machinery to repair elements damaged by UV.

Integration of speech-processing technologies into Activobank's client interaction process

This dissertation analyzes the possibilities of utilizing speech-processing technologies to transform the user experience of ActivoBank’s customers while using remote banking solutions. The technologies are examined through different criteria to determine if they support the bank’s goals and strategy and whether they should be incorporated in the bank’s offering. These criteria include the alignment with ActivoBank’s values, the suitability of the technology providers, the benefits these technologies entail, potential risks, appeal to the customers and impact on customer satisfaction. The analysis suggests that ActivoBank might not be in a position to adopt these technologies at this point in time.

Design of bio-inspired ionic liquids for protein stabilisation

Ionic Liquids (ILs) consist in organic salts that are liquid at/or near room temperature. Since ILs are entirely composed of ions, the formation of ion pairs is expected to be one essential feature for describing solvation in ILs. In recent years, protein - ionic liquid (P-IL) interactions have been the subject of intensive studies mainly because of their capability to promote folding/unfolding of proteins. However, the ion pairs and their lifetimes in ILs in P-IL thematic is dismissed, since the action of ILs is therefore the result of a subtle equilibrium between anion-cation interaction, ion-solvent and ion-protein interaction. The work developed in this thesis innovates in this thematic, once the design of ILs for protein stabilisation was bio-inspired in the high concentration of organic charged metabolites found in cell milieu. Although this perception is overlooked, those combined concentrations have been estimated to be ~300 mM among the macromolecules at concentrations exceeding 300 g/L (macromolecular crowding) and transient ion-pair can naturally occur with a potential specific biological role. Hence the main objective of this work is to develop new bio-ILs with a detectable ion-pair and understand its effects on protein structure and stability, under crowding environment, using advanced NMR techniques and calorimetric techniques. The choline-glutamate ([Ch][Glu]) IL was synthesized and characterized. The ion-pair was detected in water solutions using mainly the selective NOE NMR technique. Through the same technique, it was possible to detect a similar ion-pair promotion under synthetic and natural crowding environments. Using NMR spectroscopy (protein diffusion, HSQC experiments, and hydrogen-deuterium exchange) and differential scanning calorimetry (DSC), the model protein GB1 (production and purification in isotopic enrichment media) it was studied in the presence of [Ch][Glu] under macromolecular crowding conditions (PEG, BSA, lysozyme). Under dilute condition, it is possible to assert that the [Ch][Glu] induces a preferential hydration by weak and non-specific interactions, which leads to a significant stabilisation. On the other hand, under crowding environment, the [Ch][Glu] ion pair is promoted, destabilising the protein by favourable weak hydrophobic interactions , which disrupt the hydration layer of the protein. However, this capability can mitigates the effect of protein crowders. Overall, this work explored the ion-pair existence and its consequences on proteins in conditions similar to cell milieu. In this way, the charged metabolites found in cell can be understood as key for protein stabilisation.

Generic system for human-computer gesture interaction: applications on sign language recognition and robotic soccer refereeing

Hand gestures are a powerful way for human communication, with lots of potential applications in the area of human computer interaction. Vision-based hand gesture recognition techniques have many proven advantages compared with traditional devices, giving users a simpler and more natural way to communicate with electronic devices. This work proposes a generic system architecture based in computer vision and machine learning, able to be used with any interface for human-computer interaction. The proposed solution is mainly composed of three modules: a pre-processing and hand segmentation module, a static gesture interface module and a dynamic gesture interface module. The experiments showed that the core of visionbased interaction systems could be the same for all applications and thus facilitate the implementation. For hand posture recognition, a SVM (Support Vector Machine) model was trained and used, able to achieve a final accuracy of 99.4%. For dynamic gestures, an HMM (Hidden Markov Model) model was trained for each gesture that the system could recognize with a final average accuracy of 93.7%. The proposed solution as the advantage of being generic enough with the trained models able to work in real-time, allowing its application in a wide range of human-machine applications. To validate the proposed framework two applications were implemented. The first one is a real-time system able to interpret the Portuguese Sign Language. The second one is an online system able to help a robotic soccer game referee judge a game in real time.

Generic system for human-computer gesture interaction

Hand gestures are a powerful way for human communication, with lots of potential applications in the area of human computer interaction. Vision-based hand gesture recognition techniques have many proven advantages compared with traditional devices, giving users a simpler and more natural way to communicate with electronic devices. This work proposes a generic system architecture based in computer vision and machine learning, able to be used with any interface for humancomputer interaction. The proposed solution is mainly composed of three modules: a pre-processing and hand segmentation module, a static gesture interface module and a dynamic gesture interface module. The experiments showed that the core of vision-based interaction systems can be the same for all applications and thus facilitate the implementation. In order to test the proposed solutions, three prototypes were implemented. For hand posture recognition, a SVM model was trained and used, able to achieve a final accuracy of 99.4%. For dynamic gestures, an HMM model was trained for each gesture that the system could recognize with a final average accuracy of 93.7%. The proposed solution as the advantage of being generic enough with the trained models able to work in real-time, allowing its application in a wide range of human-machine applications.

Leaf micronutrient concentrations and potential photosynthesis in Ochroma pyramidale established in a degraded land

In the Brazilian Amazon, large areas of abandoned lands may revert to secondary forest. In the process, pioneer tree species have an important role to restore productivity in old fields and improve environmental conditions. To determine potential photosynthesis (Apot), stomatal conductance (g), transpiration (E), and leaf micronutrient concentrations in Ochroma pyramidale (Cav. ex Lam.) Urban a study was carried out in the Brazilian Amazon (01o 51' S; 60o 04' W). Photosynthetic parameters were measured at increasing [CO2], saturating light intensity (1 mmol (photons) m-2 s-1), and ambient temperature. The rate of electron-transport (J), Apot,and water-use efficiency (WUE) increased consistently at increasing internal CO2 concentration (Ci). Conversely, increasing [CO2] decreased gs, E, and photorespiration (Pr). At the CO2-saturated region of the CO2 response curve (1.1 mmol (CO2) mol-1(air), J was 120 μmol (e-) m-2s-1 and Apot reached up to 24 μmol (CO2) m-2s-1. Likewise, at saturating C1 g and E were 30 and 1.4 mmol (H2O) m-2s-1, respectively, and P 2 r about 1.5 μmol (CO2) m-2s-1. Foliar nutrients were 185, 134, 50, and 10 μmol (element) m-2 (leaf area) for Fe, Mn, Zn, and Cu, respectively. It was concluded that [CO ] probably limits light saturated photosynthesis in this site. Furthermore, from a nutritional point of view, the low Fe to Cu ratio (15:1) may reflect nutritional imbalance in O. pyramidale at this site.

(Un)suitability of the use of pH buffers in biological, biochemical and environmental studies and their interaction with metal ions - a review

The use of buffers to maintain the pH within a desired range is a very common practice in chemical, biochemical and biological studies. Among them, zwitterionic N-substituted aminosulfonic acids, usually known as Good's buffers, although widely used, can complex metals and interact with biological systems. The present work reviews, discusses and updates the metal complexation characteristics of thirty one commercially available buffers. In addition, their impact on biological systems is also presented. The influences of these buffers on the results obtained in biological, biochemical and environmental studies, with special focus on their interaction with metal ions, are highlighted and critically reviewed. Using chemical speciation simulations, based on the current knowledge of the metal-buffer stability constants, a proposal of the most adequate buffer to employ for a given metal ion is presented.

Ion exchange dependent electroactive phase content and electrical properties of poly(vinylidene fluoride)/Na(M)Y composites

Different metal-ion exchanged NaY zeolite, Na(M)Y, were used to prepare poly(vinylidene fluoride) based composites by solvent casting and melting crystallization. The effect of different metal ion-exchanged zeolites on polymer crystallization and electrical properties was reported. Cation-framework interactions and hydration energy of the cations determined that K+ is the most efficient exchanged ion in NaY zeolite, followed by Cs+ and Li+. The electroactive phase crystallization strongly depends on the ions present in the zeolite, leading to variations of the surface energy characteristics of the Na(M)Y zeolites and the polymer chain ability of penetration in the zeolite. Thus, Na(Li)Y and NaY induces the complete electroactive -phase crystallization of the crystalline phase of PVDF, while Na(K)Y only induces it partly and Na(Cs)Y is not able to promote the crystallization of the electroactive phase. Furthermore, different ion size/weigh and different interaction with the zeolite framework results in significant variations in the electrical response of the composite. In this way, iinterfacial polarization effects in the zeolite cavities and zeolite-polymer interface, leads to strong increases of the dielectric constant on the composites with lightest ions weakly bound to the zeolite framework. Polymer composite with Na(Li)Y show the highest dielectric response, followed by NaY and Na(K)Y. Zeolite Na(Cs)Y contribute to a decrease of the dielectric constant of the composite. The results show the relevance of the materials for sensor development.

Estudo de validação do Partner Interaction Questionnaire para fumantes e ex-fumantes

O presente artigo tem por objetivo validar uma versão adaptada do Partner Interaction Questionnaire (PIQ), instrumento que avalia o suporte para deixar de fumar dado pelo parceiro. É apresentada uma versão adaptada do PIQ para fumantes e abstinentes. Participaram no estudo 224 fumantes (52.7% do sexo feminino e 47.3% do sexo masculino; média de idades=28.6 anos; DP=8.69) e 169 ex-fumantes (68.6% do sexo masculino e 31.4% do sexo feminino; média de idades= 43.1 anos; DP=13.02). O instrumento apresentou boa fidelidade (alfa de Cronbach: 0.73) para a subescala positiva e para a subescala negativa (alfa de Cronbach: 0.77). O estudo de validade revelou a presença de dois fatores que explicam 43.36% da variância. O instrumento apresenta boa validade de construto, correlacionando-se positivamente com o ajustamento de casais e a qualidade de vida mental, e negativamente com a depressão. Esta versão adaptada do PIQ reuniu as condições necessárias para a sua aplicação em fumantes e ex-fumantes, podendo ser uma escala útil em programas de intervenção de cessação tabágica e de manutenção da abstinência.

Carbon materials as new generation of new electron shuttles for the anaerobic degradation of environmental xenobiotics

Tese de Doutoramento em Engenharia Química e Biológica

Analysis of physicochemical interaction between the components of asphalt mixtures with rubber

Tese de Doutoramento em Engenharia Civil