Toxic effect and inability of L-homoserine to be a nitrogen source for growth of Escherichia coli resolved by a combination of in vivo evolution engineering and omics analyses.

L-homoserine is a pivotal intermediate in the carbon and nitrogen metabolism of E. coli. However, this non-canonical amino acid cannot be used as a nitrogen source for growth. Furthermore, growth of this bacterium in a synthetic media is potently inhibited by L-homoserine. To understand this dual effect, an adapted laboratory evolution (ALE) was applied, which allowed the isolation of a strain able to grow with L-homoserine as the nitrogen source and was, at the same time, desensitized to growth inhibition by this amino acid. Sequencing of this evolved strain identified only four genomic modifications, including a 49 bp truncation starting from the stop codon of thrL. This mutation resulted in a modified thrL locus carrying a thrL* allele encoding a polypeptide 9 amino acids longer than the thrL encoded leader peptide. Remarkably, the replacement of thrL with thrL* in the original strain MG1655 alleviated L-homoserine inhibition to the same extent as strain 4E, but did not allow growth with this amino acid as a nitrogen source. The loss of L-homoserine toxic effect could be explained by the rapid conversion of L-homoserine into threonine via the thrL*-dependent transcriptional activation of the threonine operon thrABC. On the other hand, the growth of E. coli on a mineral medium with L-homoserine required an activation of the threonine degradation pathway II and glycine cleavage system, resulting in the release of ammonium ions that were likely recaptured by NAD(P)-dependent glutamate dehydrogenase. To infer about the direct molecular targets of L-homoserine toxicity, a transcriptomic analysis of wild-type MG1655 in the presence of 10 mM L-homoserine was performed, which notably identified a potent repression of locomotion-motility-chemotaxis process and of branched-chain amino acids synthesis. Since the magnitude of these effects was lower in a ΔthrL mutant, concomitant with a twofold lower sensitivity of this mutant to L-homoserine, it could be argued that growth inhibition by L-homoserine is due to the repression of these biological processes. In addition, L-homoserine induced a strong upregulation of genes in the sulfate reductive assimilation pathway, including those encoding its transport. How this non-canonical amino acid triggers these transcriptomic changes is discussed.

Kinetic modeling of ethanol production by Scheffersomyces stipitis from xylose.

This work focuses on the kinetics of ethanol production by Scheffersomyces stipitis on xylose with the development of a mathematical model considering the effect of substrate and product concentrations on growth rate. Experiments were carried out in batch and continuous modes, with substrate concentration varying from 7.2 to 145 g L(-1). Inhibitory effects on cell growth, substrate uptake, and ethanol production rates were found to be considerable. Kinetic parameters were obtained through linear and non-linear regression methods. Experiments in continuous mode were performed at different dilution rates to evaluate the inhibitory effect of ethanol. A mixed mathematical model which combined Andrews and Levenspiel's models, combining substrate and product inhibition, was used. A quasi-Newton routine was applied to obtain a more accurate fitting of kinetic parameters. The parameters such as cell to product factor (YP/X) and limiting cell yield (YX) were shown to be dependent on substrate concentration. The kinetic model fitted satisfactorily the experimental data.

Discriminação com três diferentes contingências em SA: extinção, reforçamento e punição, extinção e punição / Discrimination using three different contingences in SA: extinction, reinforcement and punishment, extinction and punishment

Estudos relataram maior eficácia no treino discriminativo quando choque elétrico (usado como punidor ou como reforçador negativo) foi combinado à extinção na presença de SΔ. O presente estudo explorou a generalidade desses dados com jato de ar quente (JAQ). No Experimento 1, três grupos de dois ratos aprenderam discriminação escuro (SD) versus claro (SΔ) em três diferentes contingências para SΔ: extinção, reforçamento+punição e extinção+ punição. Observou-se uma maior eficiência do treino combinando extinção+punição na produção da discriminação. No Experimento 2, as funções claro versus escuro foram invertidas em relação ao Experimento 1. Observou-se equivalência entre reforçamento+punição e extinção+ punição, ambas mais eficazes que a extinção sozinha. De forma geral, os dados confirmam a eficiência do estabelecimento de contingências de punição positiva combinada com extinção na presença de SΔ em contexto de treino discriminativo utilizando o JAQ como aversivo, ampliando a generalidade do fenômeno.

Previous studies reported better efficacy in discrimination training when electric shock (used as punisher or as negative reinforcer stimulus) was combined with extinction in presence of SΔ. The present study explored the generality of such data with Hot Air Blast (HAB). In Experiment 1, three groups of two rats learned discrimination between light (SΔ) versusdark (SD) under three different contingencies for SΔ: extinction, reinforcement+ punishment, and extinction+punishment. The data showed more training efficacy when combining punishment and extinction. In Experiment 2, the functions of light and dark stimuli were reversed. The data showed equivalence between punishment and extinction+ punishment, with both more efficient than extinction alone. Generally, the data confirm the efficacy of arranging punishment contingencies combined with extinction in presence ofin context of discrimination training with HBA as aversive stimulus, increasing the generality of the phenomenon.

Efeito antioxidante da ficocianina em pescado salgado-seco / Antioxidant effect of phycocianin on dried-salted fish

A conservação e o armazenamento de alimentos com elevados teores lipídicos podem conduzir ao desencadeamento do processo oxidativo, causando modificações nas características sensoriais e redução da vida útil dos mesmos. A ficocianina é um pigmento extraído da microalga Spirulina platensis que apresenta propriedades antioxidantes, por isso, objetivou-se verificar o efeito antioxidante da ficocianina durante o armazenamento do pescado salgado. O pescado salgado foi produzido com o peixe Pacu (Piaractus mesopotamicus, Holmberg, 1887), o qual foi submetido a 5 tratamentos (T1: sem adição de antioxidante, T2: com adição de antioxidante sintético BHT na concentração de 0,01 por cento, T3, T4 e T5: com a adição de ficocianina nas concentrações de 0,5 por cento, 1 por cento e 1,5 por cento, respectivamente). As amostras foram analisadas nos tempos: zero, 30 d e 60 d de armazenamento através do índice de TBA. Pelos resultados observou-se que a presença do antioxidante natural ficocianina ocasionou uma inibição no processo de oxidação lipídica, com índices de TBA, estatisticamente iguais (p>0,05) ao tratamento com uso de antioxidante sintético.

Conservation and storage of food with a lipidic content may unchain oxidative processes, causing modifications of sensorial characteristics and reduction of useful life. Phycocianin is a pigment of Spirulina platensis, which presents antioxidant proprieties. The objective of this work was to verify the antioxidant effect of phycocyanin during the storage of salted fish. The salted fish was produced with the pacu species (Piaractus mesopotamicus Holmberg, 1887), which was submitted to 5 treatments (T1: without addition of antioxidant; T2: with addition of synthetic antioxidant BHT in the concentration of 0.01 percent, T3, T4 and T5: with addition of phycocianin at concentrations of 0.5 percent, 1 percent and 1.5 percent respectively). Samples were analyzed at zero (0), 30 and 60 days of storage determining the physics-chemical quality through the index of TBA. The results showed that the addition of the natural antioxidant phycocianin caused the inhibition of the process of oxidation, with TBA index statistically equal (p>0.05) of that obtained with the use of synthetic antioxidant.

Antioxidant effect of phycocyanin on oxidative stress induced with monosodium glutamate in rats

The objective of this work was to study the antioxidant effect of phycocyanin on the oxidative stress induced by monosodium glutamate in the rats. The tests were performed with 32 rats of Wistar breed, divided into four groups, which were administered saline solution of phycocyanin, monosodium glutamate and monosodium glutamate plus phycocyanin. Sulfhydryl groups and the secondary substances derived from lipid oxidation were determined through the level of TBA. The evaluation of these values and the level of sulfhydryl showed that the administration of phycocyanin presented significant antioxidant effect (p < 0.05) reducing the oxidative stress induced by the monosodium glutamate in vivo.

Potencial amilolítico do grão de milho maltado no processo de sacarificação do mesmo cereal / Amylolytic potential of maize malt in the process of sacarification of the same cereal

Objetivou-se avaliar o potencial de sacarificação do malte de milho para produção de álcool deste cereal. Para tal, foi realizada germinação do grão de milho a 20 ºC por 5 dias. O potencial amilolítico do malte de milho foi otimizado por meio de um PlanejamentoFatorial Completo 22 com 3 pontos centrais, sendo as variáveis de estudo a temperatura de incubação e concentração de substrato [S]. A sacarificação foi realizada por meio de um Planejamento de Mistura com três pontos centrais, sendo as variáveis de estudo a concentração de malte e concentração de substrato, durante 6 horas. A fermentação foi conduzida com a melhor condição de sacarificação, em reator de 5 L e volume útil de 2 L. O pH foi fixado em 4,5-5. O Brix foi acertado para o valor de 14º. O mosto hidrolisado foi esterilizado e inoculado com 5,0 g.L-1 de Saccharomyces cerevisiae proveniente de fermento comercial liofilizado. O sistema foi submetido a 2 horas de aeração inicial (170 LO2.h -1) e mantido na temperatura de 30 °C. Durante o processo de fermentação, foram realizadas determinações da concentração de células (g.L-1), açúcares redutores (mg.mL-1) e álcool (g.L-1). As variáveis temperatura de incubação e concentração de substrato foram significativos (p< 0,05) na condição de estudo de 20 % de substrato e incubação a 70 °C. Os experimentos centrais com 50 % de malte apresentaram os melhores parâmetros cinéticos com taxa volumétrica de formação de produto em etanol de 2,81 getanol.L-1.h-1 que corresponde a 8,0 ºGL e conversão de substrato em células 0,127 gcélula.gglicose-1.

The aim of this research was the production of maize malt and the evaluation of its use for alcohol production. The maize grain was allowed to germinate at 20°C for five days. The amylolitic potential of malt was optimized through a Complete Factorial Planning 22 with three central points, where the study variables were the temperature of incubation and the substrate concentration [S]. The sacarification was performed through a Planning of Mixture with three central points, where the study variables were the malt concentration and [S] during six 6 hours. The fermentation was carried out using the best sacarification condition, in a five-liter reactor and a two-liter working volume. The pH was maintained at 4.5 – 5.0. The Brix was corrected to 14º. The sterile hydrolyzedproduct was inoculated with 5.0 g.L-1 Saccharomyces cerevisiae. The system was submitted to two hours of initial aeration (170 LO2.h-1) and kept at 30°C. During the fermentation process, analyses were made to determine the cell concentrations (g.L-1), reducingsugars (mg.mL-1), alcohol (g.L-1) and °Brix. The temperature of incubation and substrate concentration were significant factors (p<0.05) in the determination of the amylolytic potential under the conditions of study utilizing 20% of substrate and incubation at 70°C. The central experiments with 50% malt presented the best kinetic parameters with a volumetric rate of ethanol formation of 2.81 gethanol.L-1.h-1, which correspond to 8.0 ºGL, and a substrate conversion in cells of 0.127gcells.gglucose -1.

Análise da topografia axial dos terceiros molares inclusos através da radiografia panorâmica dos maxilares em relação à classificação de Winter / Molar analysis of the axial topography of third enclosed through the panoramic x-ray of the maxilares in relation to the classification of Winter

A radiografia panorâmica é um dos mais freqüentes exames complementares solicitados na clínica odontológica sendo de suma importância para avaliação das inclusões dentais e nos planejamentos cirúrgicos. O objetivo deste trabalho é analisar os terceiros molares inclusos através da radiografia panorâmica em relação à classificação de Winter (1926). Foram analisadas 200 radiografias selecionadas aleatoriamente de uma clínica particular de radiologia odontológica em Caruaru-PE, de pacientes numa faixa etária entre 18 e 30 anos de ambos os gêneros, no período de janeiro a junho de 2006. Das radiografias analisadas 58% (n = 116) eram do gênero feminino e 42% (n = 84) eram do gênero masculino. A média de idade foi de 22,4 anos e a maior freqüência foi o elemento 28, 26,5% (n = 164) do qual 26,5% (n = 69) eram do gênero masculino e 27,2% (n = 95) seguido do elemento 18, 24,1% (n = 163) ambos com a posição vertical mais freqüente.