Deteção eletroquímica da norfloxacina e trimetoprim em amostras de urina utilizando polímero molecularmente impresso modificado e grafeno

A norfloxacina e o trimetoprim são dois antibióticos antibacterianos usados para o tratamento de infeções urinárias, intestinais e respiratórias. A maioria dos fármacos exige uma dosagem que garanta os níveis de segurança e eficácia de atuação. A necessidade de dosear os medicamentos e os seus metabolitos é assim um controlo imperioso e em muitos casos regular no tratamento de um paciente. Neste trabalho desenvolveram-se dois sensores eletroquímicos para a deteção da norfloxacina (NFX) e do trimetoprim (TMP), usando como superfície de suporte o carbono vítreo. A busca de novos materiais que conferiram maior seletividade e sensibilidade aos sistemas de deteção e por outro lado apresentem menores riscos para o paciente quando usados em dispositivos que permitam uma análise point-of-care, é especialmente importante e pode ser uma parte crucial do processo de decisão clínica. Assim, os polímeros molecularmente impresos enquadram-se nesse perfil e o seu uso tem vindo a ser cada vez mais avaliado. A impressão molecular é uma tecnologia capaz de produzir polímeros que incorporam as moléculas do analito e que após remoção por solventes específicos, permitem dotá-los de locais específicos de reconhecimento estereoquímico. A seleção do pirrol como polímero molecularmente impresso (MIP) permitiu construir com sucesso os sensores para doseamento dos antibióticos. A fim de aumentar a sensibilidade do método incorporou-se grafeno na superfície do elétrodo. Este material tem vindo a ser largamente utilizado devido às suas propriedades: estrutura molecular, condutividade elétrica e aumento da superfície são algumas das características que mais despertam o interesse para a sua aplicação neste projeto. Os sensores desenvolvidos foram incorporados em sistemas eletroquímicos. Os métodos voltamétricos aplicados foram a voltametria cíclica, a voltametria de onda quadrada e ainda a impedância. As condições de análise foram otimizadas no que respeita à polimerização do pirrol (concentração do polímero, número de ciclos de eletropolimerização e respetivos potenciais aplicados, tempo de incubação, solvente de remoção do analito), ao pH da solução do fármaco, à gama de concentrações dos antibióticos e aos parâmetros voltamétricos dos métodos de análise. Para cada um dos antibióticos um elétrodo não-impresso foi também preparado, usando o procedimento de polimerização mas sem a presença da molécula do analito, e foi usado como controlo. O sensor desenvolvido para o trimetoprim foi usado no doseamento do fármaco em amostras de urina. As amostras foram analisadas sem qualquer diluição, apenas foram centrifugadas para remoção de proteínas e algum interferente. Os sensores construídos apresentaram comportamento linear na gama de concentrações entre 102 e 107 mol/L. Os resultados mostram boa precisão (desvios padrão inferiores a 11%) e os limites de deteção foram de 8,317 e 1,307 mol/L para a norfloxacina e o trimetoprim, respetivamente. Para validação do método foram ainda efetuados ensaios de recuperação tendo obtido valores superiores a 94%.

Analysis of the combined exposure to noise and ototoxic substances—pilot study

Abstract: Ototoxic substances have been associated to damage of the auditory system, and its effects are potentiated by noise exposure. The present study aims at analyzing auditory changes from combined exposure to noise and organic solvents, through a pilot study in the furniture industry sector. Audiological tests were performed on 44 workers, their levels of exposure to toluene, xylene and ethylbenzene were determined and the levels of noise exposure were evaluated. The results showed that workers are generally exposed to high noise levels and cabin priming filler and varnish sector workers have high levels of exposure to toluene. However, no hearing loss was registered among the workers. Workers exposed simultaneously to noise and ototoxic substances do not have a higher degree of hearing loss than those workers exposed only to noise. Thus, the results of this study did not show that the combined exposure to noise and the organic solvent is associated with hearing disorders.

Green production of zero-valent iron nanoparticles using tree leaf extracts

The interest in zero-valent iron nanoparticles has been increasing significantly since the development of a green production method in which extracts from natural products or wastes are used. However, this field of application is yet poorly studied and lacks knowledge that allows the full understanding of the production and application processes. The aim of the present work was to evaluate the viability of the utilization of several tree leaves to produce extracts which are capable of reducing iron(III) in aqueous solution to form nZVIs. The quality of the extracts was evaluated concerning their antioxidant capacity. The results show that: i) dried leaves produce extracts with higher antioxidant capacities than non-dried leaves, ii) the most favorable extraction conditions (temperature, contact time, and volume:mass ratio) were identified for each leaf, iii) with the aim of developing a green, but also low-cost,method waterwas chosen as solvent, iv) the extracts can be classified in three categories according to their antioxidant capacity (expressed as Fe(II) concentration): >40 mmol L−1; 20–40 mmol L−1; and 2–10 mmol L−1; with oak, pomegranate and green tea leaves producing the richest extracts, and v) TEManalysis proves that nZVIs (d=10–20 nm) can be produced using the tree leaf extracts.

Validation of a single-extraction procedure for sequential analysis of vitamin E, cholesterol, fatty acids, and total fat in seafood

Food lipid major components are usually analyzed by individual methodologies using diverse extractive procedures for each class. A simple and fast extractive procedure was devised for the sequential analysis of vitamin E, cholesterol, fatty acids, and total fat estimation in seafood, reducing analyses time and organic solvent consumption. Several liquid/liquid-based extractive methodologies using chlorinated and non-chlorinated organic solvents were tested. The extract obtained is used for vitamin E quantification (normal-phase HPLC with fluorescence detection), total cholesterol (normal-phase HPLC with UV detection), fatty acid profile, and total fat estimation (GC-FID), all accomplished in <40 min. The final methodology presents an adequate linearity range and sensitivity for tocopherol and cholesterol, with intra- and inter-day precisions (RSD) from 3 to 11 % for all the components. The developed methodology was applied to diverse seafood samples with positive outcomes, making it a very attractive technique for routine analyses in standard equipped laboratories in the food quality control field.

Determination of ochratoxin A in bread: evaluation of microwave-assisted extraction using an orthogonal composite design coupled with response surface methodology

An analytical method using microwave-assisted extraction (MAE) and liquid chromatography (LC) with fluorescence detection (FD) for the determination of ochratoxin A (OTA) in bread samples is described. A 24 orthogonal composite design coupled with response surface methodology was used to study the influence of MAE parameters (extraction time, temperature, solvent volume, and stirring speed) in order to maximize OTA recovery. The optimized MAE conditions were the following: 25 mL of acetonitrile, 10 min of extraction, at 80 °C, and maximum stirring speed. Validation of the overall methodology was performed by spiking assays at five levels (0.1–3.00 ng/g). The quantification limit was 0.005 ng/g. The established method was then applied to 64 bread samples (wheat, maize, and wheat/maize bread) collected in Oporto region (Northern Portugal). OTAwas detected in 84 % of the samples with a maximum value of 2.87 ng/g below the European maximum limit established for OTA in cereal products of 3 ng/g.

Extração e caracterização de extratos de Jatropha gossypiifolia L.: avaliação da sua atividade antimicrobiana e antioxidante

Trabalho Final de Mestrado para obtenção do grau de mestre em Engenharia Química e Biológica

Simulação de processos de adsorção recorrendo ao Aspen Chromatography

As operações de separação por adsorção têm vindo a ganhar importância nos últimos anos, especialmente com o desenvolvimento de técnicas de simulação de leitos móveis em colunas, tal como a cromatografia de Leito Móvel Simulado (Simulated Moving Bed, SMB). Esta tecnologia foi desenvolvida no início dos anos 60 como método alternativo ao processo de Leito Móvel Verdadeiro (True Moving Bed, TMB), de modo a resolver vários dos problemas associados ao movimento da fase sólida, usuais nestes métodos de separação cromatográficos de contracorrente. A tecnologia de SMB tem sido amplamente utilizada em escala industrial principalmente nas indústrias petroquímica e de transformação de açúcares e, mais recentemente, na indústria farmacêutica e de química fina. Nas últimas décadas, o crescente interesse na tecnologia de SMB, fruto do alto rendimento e eficiente consumo de solvente, levou à formulação de diferentes modos de operação, ditos não convencionais, que conseguem unidades mais flexíveis, capazes de aumentar o desempenho de separação e alargar ainda mais a gama de aplicação da tecnologia. Um dos exemplos mais estudados e implementados é o caso do processo Varicol, no qual se procede a um movimento assíncrono de portas. Neste âmbito, o presente trabalho foca-se na simulação, análise e avaliação da tecnologia de SMB para dois casos de separação distintos: a separação de uma mistura de frutose-glucose e a separação de uma mistura racémica de pindolol. Para ambos os casos foram considerados e comparados dois modos de operação da unidade de SMB: o modo convencional e o modo Varicol. Desta forma, foi realizada a implementação e simulação de ambos os casos de separação no simulador de processos Aspen Chromatography, mediante a utilização de duas unidades de SMB distintas (SMB convencional e SMB Varicol). Para a separação da mistura frutose-glucose, no quediz respeito à modelização da unidade de SMB convencional, foram utilizadas duas abordagens: a de um leito móvel verdadeiro (modelo TMB) e a de um leito móvel simulado real (modelo SMB). Para a separação da mistura racémica de pindolol foi considerada apenas a modelização pelo modelo SMB. No caso da separação da mistura frutose-glucose, procedeu-se ainda à otimização de ambas as unidades de SMB convencional e Varicol, com o intuito do aumento das suas produtividades. A otimização foi realizada mediante a aplicação de um procedimento de planeamento experimental, onde as experiências foram planeadas, conduzidas e posteriormente analisadas através da análise de variância (ANOVA). A análise estatística permitiu selecionar os níveis dos fatores de controlo de modo a obter melhores resultados para ambas as unidades de SMB.

Construção de curvas binodais para sistemas ternários, contendo biodiesel, álcool e água

Trabalho Final de Mestrado para obtenção do grau de Mestre em Engenharia Química

Self-reported hearing performance in workers exposed to solvents

OBJECTIVE: To compare hearing performance relating to the peripheral and central auditory system between solvent-exposed and non-exposed workers. METHODS: Forty-eight workers exposed to a mixture of solvents and 48 non-exposed control subjects of matched age, gender and educational level were selected to participate in the study. The evaluation procedures included: pure-tone audiometry (500 - 8,000 Hz), to investigate the peripheral auditory system; the Random Gap Detection test, to assess the central auditory system; and the Amsterdam Inventory for Auditory Disability and Handicap, to investigate subjects' self-reported hearing performance in daily-life activities. A Student t test and analyses of covariance (ANCOVA) were computed to determine possible significant differences between solvent-exposed and non-exposed subjects for the hearing level, Random Gap Detection test and Amsterdam Inventory for Auditory Disability and Handicap. Pearson correlations among the three measures were also calculated. RESULTS: Solvent-exposed subjects exhibited significantly poorer hearing thresholds for the right ear than non-exposed subjects. Also, solvent-exposed subjects exhibited poorer results for the Random Gap Detection test and self-reported poorer listening performance than non-exposed subjects. Results of the Amsterdam Inventory for Auditory Disability and Handicap were significantly correlated with the binaural average of subject pure-tone thresholds and Random Gap Detection test performance. CONCLUSIONS: Solvent exposure is associated with poorer hearing performance in daily life activities that relate to the function of the peripheral and central auditory system.

Aplicação de filmes de suberina na produção de materiais bio-funcionais

Trabalho Final de Mestrado para obtenção do grau de Mestre em Engenharia Química e Biológica

Mining protein structure data

The principal topic of this work is the application of data mining techniques, in particular of machine learning, to the discovery of knowledge in a protein database. In the first chapter a general background is presented. Namely, in section 1.1 we overview the methodology of a Data Mining project and its main algorithms. In section 1.2 an introduction to the proteins and its supporting file formats is outlined. This chapter is concluded with section 1.3 which defines that main problem we pretend to address with this work: determine if an amino acid is exposed or buried in a protein, in a discrete way (i.e.: not continuous), for five exposition levels: 2%, 10%, 20%, 25% and 30%. In the second chapter, following closely the CRISP-DM methodology, whole the process of construction the database that supported this work is presented. Namely, it is described the process of loading data from the Protein Data Bank, DSSP and SCOP. Then an initial data exploration is performed and a simple prediction model (baseline) of the relative solvent accessibility of an amino acid is introduced. It is also introduced the Data Mining Table Creator, a program developed to produce the data mining tables required for this problem. In the third chapter the results obtained are analyzed with statistical significance tests. Initially the several used classifiers (Neural Networks, C5.0, CART and Chaid) are compared and it is concluded that C5.0 is the most suitable for the problem at stake. It is also compared the influence of parameters like the amino acid information level, the amino acid window size and the SCOP class type in the accuracy of the predictive models. The fourth chapter starts with a brief revision of the literature about amino acid relative solvent accessibility. Then, we overview the main results achieved and finally discuss about possible future work. The fifth and last chapter consists of appendices. Appendix A has the schema of the database that supported this thesis. Appendix B has a set of tables with additional information. Appendix C describes the software provided in the DVD accompanying this thesis that allows the reconstruction of the present work.

Ibuprofen-loaded poly(trimethylene carbonate-co-ϵ- caprolactone) electrospun fibres for nerve regeneration

The development of scaffolds that combine the delivery of drugs with the physical support provided by electrospun fibres holds great potential in the field of nerve regeneration. Here it is proposed the incorporation of ibuprofen, a well-known non-steroidal anti-inflammatory drug, in electrospun fibres of the statistical copolymer poly(trimethylene carbonate-co-ε-caprolactone) [P(TMC-CL)] to serve as a drug delivery system to enhance axonal regeneration in the context of a spinal cord lesion, by limiting the inflammatory response. P(TMC-CL) fibres were electrospun from mixtures of dichloromethane (DCM) and dimethylformamide (DMF). The solvent mixture applied influenced fibre morphology, as well as mean fibre diameter, which decreased as the DMF content in solution increased. Ibuprofen-loaded fibres were prepared from P(TMC-CL) solutions containing 5% ibuprofen (w/w of polymer). Increasing drug content to 10% led to jet instability, resulting in the formation of a less homogeneous fibrous mesh. Under the optimized conditions, drug-loading efficiency was above 80%. Confocal Raman mapping showed no preferential distribution of ibuprofen in P(TMC-CL) fibres. Under physiological conditions ibuprofen was released in 24h. The release process being diffusion-dependent for fibres prepared from DCM solutions, in contrast to fibres prepared from DCM-DMF mixtures where burst release occurred. The biological activity of the drug released was demonstrated using human-derived macrophages. The release of prostaglandin E2 to the cell culture medium was reduced when cells were incubated with ibuprofen-loaded P(TMC-CL) fibres, confirming the biological significance of the drug delivery strategy presented. Overall, this study constitutes an important contribution to the design of a P(TMC-CL)-based nerve conduit with anti-inflammatory properties.

Cytochrome c nitrite reductase from desulfovibrio desulfuricans ATCC 27774

The Journal of Biological Chemistry Vol. 278, No. 19, Issue of May 9, pp. 17455–17465, 2003

A further investigation of the cytochrome b5–cytochrome c complex

J Biol Inorg Chem (2003) 8: 777–786

Broad-temperature range spectroscopy of the two-centre modular redox metalloprotein Desulfovibrio desulfuricans desulfoferrodoxin

Dalton Trans., 2003, 3328-3338