A systematic approach to identify STRE-binding proteins of the gsn glycogen synthase gene promoter in Neurospora crassa

The gene encoding glycogen synthase in Neurospora crassa (gsn) is transcriptionally down-regulated when mycelium is exposed to a heat shock from 30 to 45 degrees C. The gsn promoter has one stress response element (STRE) motif that is specifically bound by heat shock activated nuclear proteins. In this work, we used biochemical approaches together with mass spectrometric analysis to identify the proteins that bind to the STRE motif and could participate in the gsn transcription regulation during heat shock. Crude nuclear extract of heat-shocked mycelium was prepared and fractionated by affinity chromatography. The fractions exhibiting DNA-binding activity were identified by electrophoretic mobility shift assay (EMSA) using as probe a DNA fragment containing the STRE motif DNA-protein binding activity was confirmed by Southwestern analysis. The molecular mass (MM) of proteins was estimated by fractionating the crude nuclear extract by SDS-PAGE followed by EMSA analysis of the proteins corresponding to different MM intervals. Binding activity was detected at the 30-50 MM kDa interval. Fractionation of the crude nuclear proteins by IEF followed by EMSA analysis led to the identification of two active fractions belonging to the pIs intervals 3.54-4.08 and 6.77-7.31. The proteins comprising the MM and pI intervals previously identified were excised from a 2-DE gel, and subjected to mass spectrometric analysis (MALDI-TOF/TOF) after tryptic digestion. The proteins were identified by search against the MIPS and MIT N. crassa databases and five promising candidates were identified. Their structural characteristics and putative roles in the gsn transcription regulation are discussed.

Multiple minima hypersurfaces procedures for biomimetic ligands screening

In this work the interaction of the pesticide carbaryl with two groups of biomimetic ligands, peptides and MIPs was screened by multiple minima hypersurfaces (MMH) procedures, through the AM1 semiempirical method. Data related to the properties of the molecular association of the complex biomimetic ligand-pesticide were obtained and compared with another molecular modeling algorithm named Leapfrog, as included in the Sybyl software package, and experimental results from the literature, remarking good correlation between them. All important MMH program parameters (cells number, box size, conformers) were studied and optimized with the aim of getting the minimum computation time without losing the correlation with experimental data. The data demonstrated that MMH approach can be used as a fast biomimetic ligand screening tool for MIPs. In the case of peptides the computation time was not comparable with the molecular dynamics methods conventionally used for this approach. © 2011 Springer Science+Business Media B.V.

Polímeros impressos com íons: fundamentos, estratégias de preparo e aplicações em química analítica

Chemical imprinting technology has been widely used as a valuable tool in selective recognition of a given target analyte (molecule or metal ion), yielding a notable advance in the development of new analytical protocols. Since their discovery, molecularly imprinted polymers (MIPs) have been extensively studied with excellent reviews published. However, studies involving ion imprinted polymers (IIPs), in which metal ions are recognized in the presence of closely related inorganic ions, remain scarce. Thus, this review involved a survey of different synthetic approaches for preparing ion imprinted adsorbents and their application for the development of solid phase extraction methods, metal ion sensors (electrodes and optodes) and selective membranes.

Desenvolvimento de biossensor baseado em extrato de açaí e sensor biomimético para detecção de hexazinona

Pós-graduação em Química - IQ

Synthesis and characterization of cross-linked molecularly imprinted polyacrylamide for the extraction/preconcentration of glyphosate and aminomethylphosphonic acid from water samples

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

A new biomimetic sensor based on molecularly imprinted polymers for highly sensitive and selective determination of hexazinone herbicide

A new selective sensor based on molecularly imprinted polymers (MIPs) was developed for the determination of hexazinone (HXZ) in environmental samples. MIPs were synthesized using a non-covalent approach, and selection of the monomers employed in the polymerization reaction was carried out by molecular modeling. Three functional monomers with high (2-vinylpyridine (MP17)) and intermediate (methacrylic acid (MP12) and acrylamide (MP5)) energies of binding to the template (HXZ) were selected for preparation of the MIPs, in order to conduct comparative studies and validate the theoretical data. For sensor construction, carbon pastes were modified with each MIP or NIP (non-imprinted polymer), and HXZ determination was performed using differential pulse adsorptive cathodic stripping voltammetry (DPAdCSV). All parameters affecting the sensor response were optimized. In HCl at pH 2.5, the sensor prepared with MP17 (5% w/w in the paste) showed a dynamic linear range between 1.9 × 10−11 and 1.1 × 10−10 mol L−1, and a detection limit of 2.6 × 10−12 mol L−1, under the following conditions: accumulation time of 200 s at a potential of −0.5V, scan rate of 50 mVs−1, pulse amplitude of 60 mV, and pulse width of 50 ms. The sensor was selective in the presence of other similar compounds, and was successfully applied to the analysis of HXZ in river water samples.

Análise do comportamento de compra de medicamentos isentos de prescrição e da automedicação

Os Medicamentos Isentos de Prescrição (MIPs) são de venda livre que têm como propósito tratar sintomas e distúrbios menores, sendo responsáveis pela movimentação de bilhões de dólares a cada ano mundialmente. É permitida a veiculação de propagandas direta ao consumidor, não sendo obrigatória a apresentação da prescrição. A disponibilidade e o consumo de MIPs podem impactar a saúde, economia e a promoção do uso racional de medicamentos, portanto se faz necessário compreender o comportamento e as motivações dos consumidores pelos MIPs e a prática da automedicação. Para tanto, conduziu-se um estudo entre março e junho de 2014 por meio de entrevistas com perguntas abertas referentes a classes terapêuticas adquiridas para automedicação, frequência com que os consumidores compraram MIPs e se automedicaram, os problemas de saúde que os motivaram a se automedicarem e a comprarem MIPs e à prática da automedicação. Foram entrevistados 47 consumidores entre 18 e 70 anos residentes em Araraquara/SP. A indicação familiar é o fator de influência mais recorrente (14), seguida pela indicação do balconista da drogaria (11) e amigos (6). A classe de MIPs mais consumida pelos entrevistados é a de analgésicos, que está relacionada com a ocorrência de dores de cabeça e musculares, sendo que a compra de MIPs ocorre, no mínimo, uma vez por mês. As motivações para a compra são principalmente o preço (33) seguido da marca (30). A automedicação ocorre também para o alívio de dores e a maioria alegou saber o conceito da automedicação porém, ainda assim, afirmou administrar por conta própria medicamentos MIPs e/ou sujeitos a prescrições, sem orientação de um profissional de saúde.

Desenvolvimento de sistema de análise para valerato de estradiol baseado em polímeros biomiméticos

Abstracts : The development of analytic methods more selective and sensitive is of great importance for a better quality in the determination of chemical species, therefore increasing the reliability of the results. In this way, the optimization of separation/concentration is still necessary. The use of Molecularly Imprinted Polymers - MIPs have demonstrated to be an efficient tool of analysis with a great potential in minimizing limitations of separation/concentration techniques traditionally employed. In general, the MIPs are obtained by polymerization in the presence of a template to be imprinted so that a polymeric skeleton is formed around the future analyte. In the present work, the template used is Estradiol Valerate (EV), compound used in the hormone replacement therapy (HRT) during climacteric. After the polymerization in bulk and in an anaerobic environment using MAA, EGDMA, AIBN, acetonitrile and VE, the obtained MIP was powdered, sifted (<120 μm) and placed in a soxhlet system containing ethanol at 60 °C, in order to remove the imprinted molecule through six successive washes in periods of 24 hours. The water used in the washings was analyzed using HPLC and spectrophotometry UV/Vis. Then, the obtained MIP was dried at room temperature and 150 mg was inset in SPE cartridges in order to evaluate the polymer's efficiency in the analyte pre-concentration and extraction. To do so, 100,0 mL of VE standard solution (2mg L-1) were pre-concentrated at 4,0 mL min-1 and eluted with 10,0 mL ethanol at 1,0 mL min-1, obtaining recoveries of 53%. Additionally, a NIP (non-imprinting polymer) was prepared to compare the obtained results, in which the recovery was 80%. In the same way, studies were conducted using commercial Strata™-X cartridges, obtaining 53% recovery. Since, the results did not reflect that than was expected, in relation with the MIP efficiency in the recovery, a computational ...