Trimethoprim-selective electrodes with molecularly imprinted polymers acting as ionophores and potentiometric transduction on graphite solid-contact

This work proposes a new biomimetic sensor material for trimethoprim. It is prepared by means of radical polymerization, having trimethylolpropane trimethacrylate as cross-linker, benzoyl peroxide as radicalar iniciator, chloroform as porogenic solvent, and methacrylic acid and 2-vinyl pyridine as monomers. Different percentages of sensor in a range between 1 and 6% were studied. Their behavior was compared to that obtained with ion-exchanger quaternary ammonium salt (additive tetrakis(p-chlorophenyl)borate or tetraphenylborate). The effect of an anionic additive in the sensing membrane was also tested. Trimethoprim sensors with 1% of imprinted particles from methacrylic acid monomers showed the best response in terms of slope (59.7 mV/decade) and detection limit (4.01 × 10− 7 mol/L). These electrodes displayed also a good selectivity towards nickel, manganese aluminium, ammonium, lead, potassium, sodium, iron, chromium, sulfadiazine, alanine, cysteine, tryptophan, valine and glycine. The sensors were not affected by pH changes from 2 to 6. They were successfully applied to the analysis of water from aquaculture.

Selective recognition in potentiometric transduction of amoxicillin by molecularly imprinted materials

The indiscriminate use of antibiotics in food-producing animals has received increasing attention as a contributory factor in the international emergence of antibiotic-resistant bacteria (Woodward in Pesticide, veterinary and other residues in food, CRC Press, Boca Raton, 2004). Numerous analytical methods for quantifying antibacterial residues in edible animal products have been developed over years (Woodward in Pesticide, veterinary and other residues in food, CRC Press, Boca Raton, 2004; Botsoglou and Fletouris in Handbook of food analysis, residues and other food component analysis, Marcel Dekker, Ghent, 2004). Being Amoxicillin (AMOX) one of those critical veterinary drugs, efforts have been made to develop simple and expeditious methods for its control in food samples. In literature, only one AMOX-selective electrode has been reported so far. In that work, phosphotungstate:amoxycillinium ion exchanger was used as electroactive material (Shoukry et al. in Electroanalysis 6:914–917, 1994). Designing new materials based on molecularly imprinted polymers (MIPs) which are complementary to the size and charge of AMOX could lead to very selective interactions, thus enhancing the selectivity of the sensing unit. AMOX-selective electrodes used imprinted polymers as electroactive materials having AMOX as target molecule to design a biomimetic imprinted cavity. Poly(vinyl chloride), sensors of methacrylic acid displayed Nernstian slopes (60.7 mV/decade) and low detection limits (2.9 × 10−5 mol/L). The potentiometric responses were not affected by pH within 4–5 and showed good selectivity. The electrodes were applied successfully to the analysis of real samples.

Host-Tailored Sensors for Carnitine Potentiometric Measurements based on Surface Molecular Imprinting

Graduate Student Symposium on Molecular Imprinting 2013, na Queen’s University, Belfast, United Kingdom, 15 a 17 de Agosto de 2013

Inoculation with Metal-Mobilizing Plant-GrowthPromoting Rhizobacterium Bacillus sp. SC2b and Its Role in Rhizoremediation

A plant growth-promoting bacterial (PGPB) strain SC2b was isolated from the rhizosphere of Sedum plumbizincicola grown in lead (Pb)/zinc (Zn) mine soils and characterized as Bacillus sp. based on (1) morphological and biochemical characteristics and (2) partial 16S ribosomal DNA sequencing analysis. Strain SC2b exhibited high levels of resistance to cadmium (Cd) (300 mg/L), Zn (730 mg/L), and Pb (1400 mg/L). This strain also showed various plant growth-promoting (PGP) features such as utilization of 1-aminocyclopropane-1-carboxylate, solubilization of phosphate, and production of indole-3-acetic acid and siderophore. The strain mobilized high concentration of heavy metals from soils and exhibited different biosorption capacity toward the tested metal ions. Strain SC2b was further assessed for PGP activity by phytagar assay with a model plant Brassica napus. Inoculation of SC2b increased the biomass and vigor index of B. napus. Considering such potential, a pot experiment was conducted to assess the effects of inoculating the metal-resistant PGPB SC2b on growth and uptake of Cd, Zn and Pb by S. plumbizincicola in metal-contaminated agricultural soils. Inoculation with SC2b elevated the shoot and root biomass and leaf chlorophyll content of S. plumbizincicola. Similarly, plants inoculated with SC2b demonstrated markedly higher Cd and Zn accumulation in the root and shoot system, indicating that SC2b enhanced Cd and Zn uptake by S. plumbizincicola through metal mobilization or plant-microbial mediated changes in chemical or biological soil properties. Data demonstrated that the PGPB Bacillus sp. SC2b might serve as a future biofertilizer and an effective metal mobilizing bioinoculant for rhizoremediation of metal polluted soils.

Effect of sialic acid loss on dendritic cell maturation

Sialic acids are key structural determinants and contribute to the functionality of a number of immune cell receptors. Previously, we demonstrated that differentiation of human dendritic cells (DCs) is accompanied by an increased expression of sialylated cell surface structures, putatively through the activity of the ST3Gal.I and ST6Gal.I sialyltransferases. Furthermore, DC endocytosis was reduced upon removal of the cell surface sialic acid residues by neuraminidase. In the present work, we evaluate the contribution of the sialic acid modifications in DC maturation. We demonstrate that neuraminidase-treated human DCs have increased expression of major histocompatibility complex (MHC) and costimulatory molecules, increased gene expression of specific cytokines and induce a higher proliferative response of T lymphocytes. Together, the data suggest that clearance of cell surface sialic acids contributes to the development of a T helper type 1 proinflammatory response. This postulate is supported by mouse models, where elevated MHC class II and increased maturation of specific DC subsets were observed in DCs harvested from ST3Gal.I(-/-) and ST6Gal.I(-/-) mice. Moreover, important qualitative differences, particularly in the extent of reduced endocytosis and in the peripheral distribution of DC subsets, existed between the ST3Gal.I(-/-) and ST6Gal.I(-/-) strains. Together, the data strongly suggest not only a role of cell surface sialic acid modifications in maturation and functionality of DCs, but also that the sialic acid linkages created by different sialyltransferases are functionally distinct. Consequently, with particular relevance to DC-based therapies, cell surface sialylation, mediated by individual sialyltransferases, can influence the immunogenicity of DCs upon antigen loading.

Pulmonary thromboembolism in AIDS patient with chronic venous insufficiency, pulmonary tuberculosis and breast cancer: a case report and pathophysiology review

Recent literature reports thrombotic episodes occurring in patients with HIV infection associated with other abnormalities including neoplasms and infections predisposing to a hypercoagulable state. We report a 47-year-old woman who developed pulmonary thromboembolism in association with HIV infection, pulmonary tuberculosis and breast cancer. She was treated with rifampin, isoniazid, pyrazinamide; heparin, phenprocoumon, zidovudine, lamivudine and efavirenz. Acid fast bacilli were visualized in a sputum smear and three months after, Mycobacterium tuberculosis was isolated from lymph node biopsy during a episode of immune reconstitution. The isolated mycobacteria showed sensitivity to all first-line drugs. HIV infection, breast cancer and pulmonary tuberculosis have several mechanisms that induce hypercoagulable state and can lead to thromboembolic complications. Pulmonary thromboembolism in this patient was a diagnostic challenge because of all the other severe diseases that she experienced at the same time.

The Spectrum of Mutations and Molecular Pathogenesis of Hemophilia A in 181 Portuguese Patients

Disease-causing alterations within the F8 gene were identified in 177 hemophilia A families of Portuguese origin. The spectrum of non-inversion F8 mutations in 101 families included 67 different alterations, namely: 36 missense, 8 nonsense and 4 splice site mutations, as well as 19 insertions/deletions. Thirty-four of these mutations are novel. Molecular modeling allowed prediction of the conformational changes introduced by selected amino acid substitutions and their correlation with the patients' phenotypes. The relatively frequent, population-specific, missense mutations together with de novo alterations can lead to significant differences in the spectrum of F8 mutations among different populations.

BIiossensor Enzimático para avaliação da Capacidade Antioxidente

Os radicais livres formam-se naturalmente nos organismos vivos, pois a sua produção/geração está interligada com o processo de produção de energia (respiração), processos inflamatórios (fagocitose), regulação do crescimento celular, sinalização intercelular e síntese de substâncias biológicas relevantes. Estes também podem ser introduzidos por vias exógenas (poluição, radiação, tabaco, alimentação, etc). Os radicais livres têm capacidade de reagir com o material nucleico (ADN e ARN), proteínas e substâncias oxidáveis, causando danos oxidativos responsáveis pelo envelhecimento e originar doenças degenerativas, tais como, o cancro, arteriosclerose, artrite reumatoide, entre outras. De forma a combater os efeitos pejorativos provocados pelos radicais, os organismos vivos desenvolveram complexos sistemas de defesa antioxidante. Estes sistemas são constituídos por antioxidantes endógenos, produzidos pelos seres vivos, tais como enzimas ou por antioxidantes exógenos obtidos por via da alimentação (por exemplo o ácido ascórbico). Neste sentido, um antioxidante tem capacidade de eliminar ou reduzir a propagação da cadeia de geração de radicais livres. Neste trabalho foi desenvolvido um biossensor enzimático para a quantificação da capacidade antioxidante total de matrizes alimentares. A construção deste biossensor consistiu na eletroimobilização da adenina no elétrodo de pasta de carbono (EPC) ou na adsorção física da dA20 na superfície do EPC. O dano oxidativo foi induzido pelo radical hidroxilo gerado pela reação de Fenton. Nesta dissertação, foi estudada a capacidade de alguns antioxidantes em eliminar o efeito pejorativo dos radicais livres e combater a integridade das bases de adenina ou do dA20.Os antioxidantes estudados foram o ácido ascórbico e alguns ácidos fenólicos como o ácido hidroxibenzoico (ácido gálico) e ácidos hidroxicinâmicos (ácido cafeico e ácido cumárico). Estes antioxidantes têm a capacidade de neutralizar o radical hidroxilo e proteger a adenina/dA20 imobilizado na superfície do EPC. O comportamento da Lacase foi estudado na presença do ácido gálico e do ácido ascórbico. Os estudos eletroquímicos foram realizados através da voltametria de onda quadrada (VOQ), sendo que a interação entre a adenina/ou o dA20 imobilizada na superfície do EPC e os radicais livres na ausência e presença de antioxidantes foi avaliada por meio de mudanças no pico anódico produzido pela oxidação da adenina /dA20. Os resultados demonstraram que estes biossensores permitem a avaliação da capacidade antioxidante total em águas aromatizadas.

Impacto Ambiental do Processo de Tingimento de uma Indústria de Curtumes e Avaliação do Desempenho da ETAR

Um dos objetivos desta dissertação de mestrado foi avaliar o impacto ambiental do processo de tingimento usado na indústria de curtumes, em termos de volume de efluente produzido e sua carga poluente. Pretendeu-se também encontrar alternativas ao nível do processo de tingimento que levem a uma melhoria na qualidade do efluente produzido. Outro objetivo deste trabalho foi avaliar o funcionamento da ETAR existente na empresa JR Fontes, no que diz respeito aos parâmetros em estudo (carência química de oxigénio - CQO, pH, crómio e teor de sólidos suspensos totais - SST), propondo alterações que permitam respeitar os valores de emissão exigidos na licença de descarga da empresa. Verificou-se que o processo de tingimento aplicado pela empresa JR Fontes é muito poluente, em termos de carga orgânica e matéria em suspensão. Este facto é comprovado pelos resultados obtidos para os três processos estudados: Montana, Galáctico e Navak. Todos os processos apresentam um efluente com pH ácido (aproximado a 3) e valores de CQO superiores a 3550 mg O2/L, sendo o processo Navak aquele que apresenta o valor mais alto para o banho composto, 8362 mg O2/L. Relativamente ao teor de crómio, o banho de recurtume com concentração mais elevada de crómio total é o banho 1 do processo Navak, com 2297 mg/L, sendo que a concentração destes banhos é sempre elevada, igual ou superior a 746 mg/L. No que diz respeito à matéria em suspensão, o processo Navak é novamente o mais poluente, com um valor de 3842 mg SST/L, não sendo obtidos nos outros processos, valores inferiores a 1205 mg SST/L. Na realização de um processo alternativo de tingimento verificou-se que é possível diminuir a carga orgânica do efluente originado. A aplicação de recurtumes sintéticos deu origem a efluentes menos poluentes. O melhor valor obtido para a CQO foi de 1113 mg O2/L, sendo obtidos valores não superiores a 7185 mg O2/L para processos de características semelhantes aos aplicados nesta indústria. Relativamente à ETAR, apesar das restrições ao funcionamento que esta apresenta, no geral pode ser considerada eficiente, embora não consiga atingir o objetivo pretendido de remoção para a CQO, ou seja o valor de 1100 mg O2/L. Nos restantes parâmetros é cumprido o limite de emissão (350 mg SST/L, pH entre 6 e 9 e 2 mg Cr/L para o crómio total): os SST apresentam o valor de 98 mg SST/L, o crómio total de 1,2 mg Cr/L e o pH encontra-se entre 8 e 9. São aqui feitas duas abordagens para solucionar os problemas existentes na ETAR. A primeira considera um ajuste no tratamento e equipamentos existentes, através da reconstrução do tanque de equalização, da substituição do coagulante por sulfato ferroso e da reconstrução do sedimentador, assim como a substituição das tubagens por umas de maior diâmetro, solucionando assim problemas de manutenção de toda a instalação e do incumprimento da legislação. A outra abordagem implica a substituição do sistema de afinação existente, os filtros de areia e carvão ativados, por um sistema de membranas de ultrafiltração ou por dois filtros de carvão ativado. Para tratar um efluente com um valor de CQO de 3000 mg O2/L com o carvão estudado, seriam necessários 132 kg de carvão por coluna, aproveitando-se os equipamentos existentes. No caso das membranas filtrantes, estas são eficazes, reduzindo a CQO em cerca de 70%. Ao contrário do carvão, a aplicação deste sistema implicaria a aquisição de novos equipamentos. Futuramente propõe-se a avaliação dos vários tipos de reagentes usados no tingimento de couro de forma a aplicar no tratamento aqueles que produzam as características desejadas no produto final e a que apresentem um menor resultado de CQO. Propõe-se também o estudo de viabilidade da remoção de crómio dos banhos de recurtume e consequente avaliação da aplicabilidade de um tratamento biológico em substituição ou como complemento do tratamento existente.

MnFe2O4@CNT-N as novel electrochemical nanosensor for determination of caffeine, acetaminophen and ascorbic acid

For the first time, a glassy carbon electrode (GCE) modified with novel N-doped carbon nanotubes (CNT-N) functionalized with MnFe2O4 nanoparticles (MnFe2O4@CNT-N) has been prepared and applied for the electrochemical determination of caffeine (CF), acetaminophen (AC) and ascorbic acid (AA). The electrochemical behaviour of CF, AC and AA on the bare GCE, CNT-N/GCE and MnFe2O4@CNT-N/GCE were carefully investigated using cyclic voltammetry (CV) and square-wave voltammetry (SWV). Compared to bare GCE and CNT-N modified electrode, the MnFe2O4@CNT-N modified electrode can remarkably improve the electrocatalytic activity towards the oxidation of CF, AC and AA with an increase in the anodic peak currents of 52%, 50% and 55%, respectively. Also, the SWV anodic peaks of these molecules could be distinguished from each other at the MnFe2O4@CNT-N modified electrode with enhanced oxidation currents. The linear response ranges for the square wave voltammetric determination of CF, AC and AA were 1.0 × 10−6 to 1.1 × 10−3 mol dm−3, 1.0 × 10−6 to 1.0 × 10−3 mol dm−3 and 2.0 × 10−6 to 1.0 × 10−4 mol dm−3 with detection limit (S/N = 3) of 0.83 × 10−6, 0.83 × 10−6 and 1.8 × 10−6 mol dm−3, respectively. The sensitivity values at the MnFe2O4@CNT-N/GCE for the individual determination of AC, AA and CF and in the presence of the other molecules showed that the quantification of AA and CF show no interferences from the other molecules; however, AA and CF interfered in the determination of AC, with the latter molecule showing the strongest interference. Nevertheless, the obtained results show that MnFe2O4@CNT-N composite material acted as an efficient electrochemical sensor towards the selected biomolecules.

ABCC Subfamily Vacuolar Transporters are Involved in Pb (Lead) Detoxification in Saccharomyces cerevisiae

The present work has as objective to contribute for the elucidation of the mechanism associated with Pb detoxification, using the yeast Saccharomyces cerevisiae as a model organism. The deletion of GTT1 or GTT2 genes, coding for functional glutathione transferases (GST) enzymes in S. cerevisiae, caused an increased susceptibility to high Pb concentrations (500-1000 μmol L(-1)). These results suggest that the formation of glutathione-Pb conjugate (GS-Pb), dependent of GSTs, is important in Pb detoxification. The involvement of ATP-binding cassette (ABC) vacuolar transporters, belonging to class C subfamily (ABCC) in vacuolar compartmentalization of Pb, was evaluated. For this purpose, mutant strains disrupted in YCF1, VMR1, YBT1 or BPT 1 genes were used. All mutants tested, without vacuolar ABCC transporters, presented an increased sensitivity to 500-1000 μmol L(-1) Pb comparative to wild-type strain. Taken together, the obtained results suggest that Pb detoxification, by vacuolar compartmentalization, can occur as a result of the concerted action of GSTs and vacuolar ABCC transporters. Pb is conjugated with glutathione, catalysed by glutathione transferases and followed to the transport of GS-Pb conjugate to the vacuole by ABCC transporters.

Optimization of the Tartrate-Resistant Acid Phosphatase Detection by Histochemical Method

According to the new KDIGO (Kidney Disease Improving Global Outcomes) guidelines, the term of renal osteodystrophy, should be used exclusively in reference to the invasive diagnosis of bone abnormalities. Due to the low sensitivity and specificity of biochemical serum markers of bone remodelling,the performance of bone biopsies is highly stimulated in dialysis patients and after kidney transplantation. The tartrate-resistant acid phosphatase (TRACP) is an iso-enzyme of the group of acid phosphatases, which is highly expressed by activated osteoclasts and macrophages. TRACP in osteoclasts is in intracytoplasmic vesicles that transport the products of bone matrix degradation. Being present in activated osteoclasts, the identification of this enzyme by histochemistry in undecalcified bone biopsies is an excellent method to quantify the resorption of bone. Since it is an enzymatic histochemical method for a thermolabile enzyme, the temperature at which it is performed is particularly relevant. This study aimed to determine the optimal temperature for identification of TRACP in activated osteoclasts in undecalcified bone biopsies embedded in methylmethacrylate. We selected 10 cases of undecalcified bone biopsies from hemodialysis patients with the diagnosis of secondary hyperparathyroidism. Sections of 5 μm were stained to identify TRACP at different incubation temperatures (37º, 45º, 60º, 70º and 80ºC) for 30 minutes. Activated osteoclasts stained red and trabecular bone (mineralized bone) was contrasted with toluidine blue. This approach also increased the visibility of the trabecular bone resorption areas (Howship lacunae). Unlike what is suggested in the literature and in several international protocols, we found that the best results were obtained with temperatures between 60ºC and 70ºC. For technical reasons and according to the results of the present study, we recommended that, for an incubation time of 30 minutes, the reaction should be carried out at 60ºC. As active osteoclasts are usually scarce in a bone section, the standardization of the histochemistry method is of great relevance, to optimize the identification of these cells and increase the accuracy of the histomosphometric results. Our results, allowing an increase in osteoclasts contrast, also support the use of semi-automatic histomorphometric measurements.

Exploring cinnamic acid scaffold: development of promising neuroprotective lipophilic antioxidants

New lipophilic hydroxycinnamic acid based derivatives were designed and synthesized and their antioxidant and neuroprotective activities evaluated. The chemical modification introduced in the cinnamic acid scaffold leads to compounds with amplified lipophilicity and in general with increased antioxidant activity when compared to natural models (caffeic and ferulic acids). The compounds did not display cytotoxicity and present a significant neuroprotective effect against 6-OH-DA induced damage to SH-SY5Y cells. Compound 6 stands out as an efficient radical scavenger and iron(II) chelator that ensures drug-like properties. Moreover, neuroprotection against oxidative damage was observed even at low concentration (1 μM). Therefore, compound 6 developed by a biology-oriented approach displays a combination of important features for a further optimization process that will generate a new effective antioxidant with therapeutic application for oxidative-stress-related events, namely neurodegenerative diseases.

Tranexamic Acid for Major Spinal Surgery in Children. A Retrospective Study

Introduction: Paediatric patients who undergo posterior spinal fusion surgery to correct scoliosis often require multiple blood transfusions. Tranexamic acid is a synthetic antifibrinolytic drug that reduces transfusion requirements in scoliosis surgery (1),(2),(3). Methods: To evaluate the efficacy of prophylactic tranexamic acid (TA) (initial dose of 10mg/kg and infusion of 1mg.kg(-1).h(-1)) in reducing perioperative blood transfusion requirements, we reviewed patients files and compared the amount of blood lost and blood transfused in the perioperative period of 12 patients (54.5%) that received TA and 10 patients (45.5%) who did not received TA. T-Student test was applied. Results: The average difference of blood losses (2,67 +/- 6,06ml) and blood transfused (212,9 +/- 101,1ml) between the two groups was not statistically significant (p>0.05). No thrombotic complications were detected in either group. Discussion: Results of the current study showed that prophylactic low dose of TA did not have a significant effect in the management of intraoperative blood loss and transfusion requirements in children undergoing scoliosis surgery. It is important to emphasize that our study is retrospective and that the size of the sample is small. Further studies are needed to evaluate the efficacy and safety of TA on paediatric scoliosis surgery.