Heavy-metal resistance in Marinobacter aquaeolei 617 insights into copper resistance

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do grau de Mestre em Biotecnologia

(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.

Use of a fluorescence-based approach to assess short-term responses of the alga Pseudokirchneriella subcapitata to metal stress

This work explores the use of fluorescent probes to evaluate the responses of the green alga Pseudokirchneriella subcapitata to the action of three nominal concentrations of Cd(II), Cr(VI), Cu(II) and Zn(II) for a short time (6 h). The toxic effect of the metals on algal cells was monitored using the fluorochromes SYTOX Green (SG, membrane integrity), fluorescein diacetate (FDA, esterase activity) and rhodamine 123 (Rh123, mitochondrial membrane potential). The impact of metals on chlorophyll a (Chl a) autofluorescence was also evaluated. Esterase activity was the most sensitive parameter. At the concentrations studied, all metals induced the loss of esterase activity. SG could be used to effectively detect the loss of membrane integrity in algal cells exposed to 0.32 or 1.3 μmol L−1 Cu(II). Rh123 revealed a decrease in the mitochondrial membrane potential of algal cells exposed to 0.32 and 1.3 μmol L−1 Cu(II), indicating that mitochondrial activity was compromised. Chl a autofluorescence was also affected by the presence of Cr(VI) and Cu(II), suggesting perturbation of photosynthesis. In conclusion, the fluorescence-based approach was useful for detecting the disturbance of specific cellular characteristics. Fluorescent probes are a useful diagnostic tool for the assessment of the impact of toxicants on specific targets of P. subcapitata algal cells.

Responses of the alga Pseudokirchneriella subcapitata to long-term exposure to metal stress

The green alga Pseudokirchneriella subcapitata has been widely used in ecological risk assessment, usually based on the impact of the toxicants in the alga growth. However, the physiological causes that lead algal growth inhibition are not completely understood. This work aimed to evaluate the biochemical and structural modifications in P. subcapitata after exposure, for 72 h, to three nominal concentrations of Cd(II), Cr(VI), Cu(II) and Zn(II), corresponding approximately to 72 h-EC10 and 72 h-EC50 values and a high concentration (above 72 h-EC90 values). The incubation of algal cells with the highest concentration of Cd(II), Cr(VI) or Cu(II) resulted in a loss of membrane integrity of ~16, 38 and 55%, respectively. For all metals tested, an inhibition of esterase activity, in a dose-dependent manner, was observed. Reduction of chlorophyll a content, decrease of maximum quantum yield of photosystem II and modification of mitochondrial membrane potential was also verified. In conclusion, the exposure of P. subcapitata to metals resulted in a perturbation of the cell physiological status. Principal component analysis revealed that the impairment of esterase activity combined with the reduction of chlorophyll a content were related with the inhibition of growth caused by a prolonged exposure to the heavy metals.

Crystal structure of the zinc-, cobalt-, and iron-containing adenylate kinase from Desulfovibrio gigas: a novel metal-containing adenylate kinase from Gram-negative bacteria

J Biol Inorg Chem (2011) 16:51–61 DOI 10.1007/s00775-010-0700-8

Synthesis, characterization and applications of new schiff base fluorescent chemosensors for metal and DNA interactions: conventional and "green" approaches

Dissertação apresentada para a obtenção do Grau de Doutor em Química Sustentável, especialidade de Química-Física Inorgânica, pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia

Portuguese banks and the economic adjustment programme: balance sheet adjustments and impact on profitability

Double Degree. A Work Project presented as part of the requirements for the Award of a Masters Degree in Management from the NOVA – School of Business and Economics and a Masters Degree in Finance from Louvain School of Management

Engineering of metal oxide nanoparticles for application in electrochemical devices

The growing demand for materials and devices with new functionalities led to the increased inter-est in the field of nanomaterials and nanotechnologies. Nanoparticles, not only present a reduced size as well as high reactivity, which allows the development of electronic and electrochemical devices with exclusive properties, when compared with thin films. This dissertation aims to explore the development of several nanostructured metal oxides by sol-vothermal synthesis and its application in different electrochemical devices. Within this broad theme, this study has a specific number of objectives: a) research of the influence of the synthesis parameters to the structure and morphology of the nanoparticles; b) improvement of the perfor-mance of the electrochromic devices with the application of the nanoparticles as electrode; c) application of the nanoparticles as probes to sensing devices; and d) production of solution-pro-cessed transistors with a nanostructured metal oxide semiconductor. Regarding the results, several conclusions can be exposed. Solvothermal synthesis shows to be a very versatile method to control the growth and morphology of the nanoparticles. The electrochromic device performance is influenced by the different structures and morphologies of WO3 nanoparticles, mainly due to the surface area and conductivity of the materials. The dep-osition of the electrochromic layer by inkjet printing allows the patterning of the electrodes without wasting material and without any additional steps. Nanostructured WO3 probes were produced by electrodeposition and drop casting and applied as pH sensor and biosensor, respectively. The good performance and sensitivity of the devices is explained by the high number of electrochemical reactions occurring at the surface of the na-noparticles. GIZO nanoparticles were deposited by spin coating and used in electrolyte-gated transistors, which promotes a good interface between the semiconductor and the dielectric. The produced transistors work at low potential and with improved ON-OFF current ratio, up to 6 orders of mag-nitude. To summarize, the low temperatures used in the production of the devices are compatible with flexible substrates and additionally, the low cost of the techniques involved can be adapted for disposable devices.

Development of an experimental setup for metal cutting research

Analytical, numerical and experimental models have been developed over time to try to characterize and understand the metal cutting process by chip removal. A true knowledge of the cutting process by chip removal is required by the increasing production, by the quality requirements of the product and by the reduced production time, in the industries in which it is employed. In this thesis an experimental setup is developed to evaluate the forces and the temperature distribution in the tool according to the orthogonal cutting model conditions, in order to evaluate its performance and its possible adoption in future works. The experimental setup is developed in a CNC lathe and uses an orthogonal cutting configuration, in which thin discs fixed onto a mandrel are cut by the cutting insert. In this experimental setup, the forces are measured by a piezoelectric dynamometer while temperatures are measured by thermocouples placed juxtaposed to the side face of the cutting insert. Three different solutions are implemented and evaluated for the thermocouples attachment in the cutting insert: thermocouples embedded in thermal paste, thermocouples embedded in copper plate and thermocouples brazed in the cutting insert. From the tests performed in the experimental setup it is concluded that the adopted forces measurement technique shows a good performance. Regarding to the adopted temperatures measurement techniques, only the thermocouples brazed in the cutting insert solution shows a good performance for temperature measurement. The remaining solutions show contact problems between the thermocouple and the side face of the cutting insert, especially when the vibration phenomenon intensifies during the cut. It is concluded that the experimental setup does not present a sufficiently robust and reliable performance, and that it can only be used in future work after making improvements in the assembly of the thermocouples.

An investigation of safety design practices of metal machines

BACKGROUND: Machinery safety issues are a challenge facing manufacturers who are supposed to create and provide products in a better and faster way. In spite of their construction and technological advance, they still contribute to many potential hazards for operators and those nearby. OBJECTIVE: The aim of this study is to investigate safety aspects of metal machinery offered for sale on Internet market according to compliance with minimum and fundamental requirements. METHODS: The study was carried out with the application of a checklist prepared on the basis of Directive 2006/42/EC and Directive 2009/104/EC and regulations enforcing them into Polish law. RESULTS: On the basis of the study it was possible to reveal the safety aspects that were not met in practice. It appeared that in the case of minimum requirements the most relevant problems concerned information, signal and control elements, technology and machinery operations, whereas as far as fundamental aspects are concerned it was hard to assure safe work process. CONCLUSIONS: In spite of the fact that more and more legal acts binding in the Member Countries of the European Union are being introduced to alleviate the phenomenon, these regulations are often not fulfilled.

(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.

Electronic structure and optical signatures of semiconducting transition metal dichalcogenide nanosheets

CONSPECTUS: Two-dimensional (2D) crystals derived from transition metal dichalcogenides (TMDs) are intriguing materials that offer a unique platform to study fundamental physical phenomena as well as to explore development of novel devices. Semiconducting group 6 TMDs such as MoS2 and WSe2 are known for their large optical absorption coefficient and their potential for high efficiency photovoltaics and photodetectors. Monolayer sheets of these compounds are flexible, stretchable, and soft semiconductors with a direct band gap in contrast to their well-known bulk crystals that are rigid and hard indirect gap semiconductors. Recent intense research has been motivated by the distinct electrical, optical, and mechanical properties of these TMD crystals in the ultimate thickness regime. As a semiconductor with a band gap in the visible to near-IR frequencies, these 2D MX2 materials (M = Mo, W; X = S, Se) exhibit distinct excitonic absorption and emission features. In this Account, we discuss how optical spectroscopy of these materials allows investigation of their electronic properties and the relaxation dynamics of excitons. We first discuss the basic electronic structure of 2D TMDs highlighting the key features of the dispersion relation. With the help of theoretical calculations, we further discuss how photoluminescence energy of direct and indirect excitons provide a guide to understanding the evolution of the electronic structure as a function of the number of layers. We also highlight the behavior of the two competing conduction valleys and their role in the optical processes. Intercalation of group 6 TMDs by alkali metals results in the structural phase transformation with corresponding semiconductor-to-metal transition. Monolayer TMDs obtained by intercalation-assisted exfoliation retains the metastable metallic phase. Mild annealing, however, destabilizes the metastable phase and gradually restores the original semiconducting phase. Interestingly, the semiconducting 2H phase, metallic 1T phase, and a charge-density-wave-like 1T' phase can coexist within a single crystalline monolayer sheet. We further discuss the electronic properties of the restacked films of chemically exfoliated MoS2. Finally, we focus on the strong optical absorption and related exciton relaxation in monolayer and bilayer MX2. Monolayer MX2 absorbs as much as 30% of incident photons in the blue region of the visible light despite being atomically thin. This giant absorption is attributed to nesting of the conduction and valence bands, which leads to diversion of optical conductivity. We describe how the relaxation pathway of excitons depends strongly on the excitation energy. Excitation at the band nesting region is of unique significance because it leads to relaxation of electrons and holes with opposite momentum and spontaneous formation of indirect excitons.

Toxicity induced by a metal mixture (Cd, Pb and Zn) in the yeast Pichia kudriavzevii: The role of oxidative stress

The present work aims to contribute for the elucidation of the role of oxidative stress in the toxicity associated with the exposure of Pichia kudriavzevii to multi-metals (Cd, Pb and Zn). Cells of the non-conventional yeast P. kudriavzevii exposed for 6 h to the action of multi-metals accumulated intracellular reactive oxygen species (ROS), evaluated through the oxidation of the probe 2,7-dichlorodihydrofluorescein diacetate. A progressive loss of membrane integrity (monitored using propidium iodide) was observed in multi-metal-treated cells. The triggering of intracellular ROS accumulation preceded the loss of membrane integrity. These results suggest that the disruption of membrane integrity can be attributed to the oxidative stress. The exposure of yeast cells to single metal showed that, under the concentrations tested, Pb was the metal responsible for the induction of the oxidative stress. Yeast cells coexposed to an antioxidant (ascorbic acid) and multi-metals did not accumulate intracellular ROS, but loss proliferation capacity. Together, the data obtained indicated that intracellular ROS accumulation contributed to metal toxicity, namely for the disruption of membrane integrity of the yeast P. kudriavzevii. It was proposed that Pb toxicity (the metal responsible for the toxic symptoms under the conditions tested) result from the combination of an ionic mechanism and the intracellular ROS accumulation.

Cytochemical characterization of gill and hepatopancreatic cells of the crab Ucides cordatus (Crustacea, Brachyura) validated by cell metal transport

Ucides cordatus (Linnaeus, 1763) is a hypo-hyper-regulating mangrove crab possessing gills for respiratory and osmoregulatory processes, separated in anterior and posterior sections. They also have hepatopancreas, which is responsible for digestion and absorption of nutrients and detoxification of toxic metals. Each of these organs has specific cells that are important for in vitro studies in cell biology, ion and toxic metals transport. In order to study and characterize cells from gills and hepatopancreas, both were separated using a Sucrose Gradient (SG) from 10 to 40% and cells in each gradient were characterized using the vital mitochondrial dye DASPEI (2-(4-dimethylaminostyryl)-N- ethylpyridinium iodide) and Trichrome Mallory's stain. Both in 20 and 40% SG for gill cells and 30% SG for hepatopancreatic cells, a greater number of cells were colored with DASPEI, indicating a larger number of mitochondria in these cells. It is concluded that the gill cells present in 20% and 40% SG are Thin cells, responsible for respiratory processes and Ionocytes responsible for ion transport, respectively. For hepatopancreatic cells, the 30% SG is composed of Fibrillar cells that possess larger number of membrane ion and nutrient transporters. Moreover, the transport of toxic metal cadmium (Cd) by isolated hepatopancreatic cells was performed as a way of following cell physiological integrity after cell separation and to study differences in transport among the cells. All hepatopancreatic cells were able to transport Cd. These findings are the first step for further work on isolated cells of these important exchange epithelia of crabs, using a simple separation method and to further develop successful in vitro cell culture in crabs.

Full Exploration of the Diels–Alder Cycloaddition on Metallofullerenes M3N@C80 (M=Sc, Lu, Gd): The D5h versus Ih Isomer and the Influence of the Metal Cluster

In this work a detailed investigation of the exohedral reactivity of the most important and abundant endohedral metallofullerene (EMF) is provided, that is, Sc3N@Ih-C80 and its D5h counterpart Sc3N@D5h-C80, and the (bio)chemically relevant lutetium- and gadolinium-based M3N@Ih/D5h-C80 EMFs (M=Sc, Lu, Gd). In particular, we analyze the thermodynamics and kinetics of the Diels–Alder cycloaddition of s-cis-1,3-butadiene on all the different bonds of the Ih-C80 and D5h-C80 cages and their endohedral derivatives. First, we discuss the thermodynamic and kinetic aspects of the cycloaddition reaction on the hollow fullerenes and the two isomers of Sc3N@C80. Afterwards, the effect of the nature of the metal nitride is analyzed in detail. In general, our BP86/TZP//BP86/DZP calculations indicate that [5,6] bonds are more reactive than [6,6] bonds for the two isomers. The [5,6] bond D5h-b, which is the most similar to the unique [5,6] bond type in the icosahedral cage, Ih-a, is the most reactive bond in M3N@D5h-C80 regardless of M. Sc3N@C80 and Lu3N@C80 give similar results; the regioselectivity is, however, significantly reduced for the larger and more electropositive M=Gd, as previously found in similar metallofullerenes. Calculations also show that the D5h isomer is more reactive from the kinetic point of view than the Ih one in all cases which is in good agreement with experiments