Adapted colorimetric method for measurement of feline urinary glycosaminoglycans

Several methods have been employed to quantify urinary glycosaminoglycans (GAGs), such as chromatography associated with electrophoresis and colorimetric methods, cheaper and faster ones, which employ mainly azure A and B, alcian blue, and dimethylmethylene blue (DMB). The purpose of this study was to standardize a reproducible and cheap method to measure total urinary GAGs in feline urine. Two colorimetric methods based on DMB were tested with chondroitin sulfate C as standard. Urine samples were obtained from 12 healthy cats and some modifications were made for the chosen method to be adequate. The modified technique using DMB acetate buffer carried out in this study can be used to measure feline urinary GAGs. © 2012 Springer-Verlag London

Efeito da idade dos frangos de corte sobre a atividade enzimática e digestibilidade dos nutrientes do farelo de soja e da soja integral

Esse trabalho visou investigar o efeito da idade de aves sobre a digestibilidade dos nutrientes da soja integral extrusada (SIE), soja integral tostada à vapor (SITV) e farelo de soja com óleo (FSO) na produção de enzimas digestivas do pâncreas. Cinco ensaios de digestibilidade foram conduzidos com frangos de corte de uma, duas, três, quatro e seis semanas de idade. Foi utilizada a metodologia de coleta total de excretas. A atividade das enzimas amilase e tripsina pancreática aumentou linearmente com a idade das aves, assim como o crescimento alométrico do pâncreas. A maior taxa de crescimento ocorreu na segunda semana, coincidindo com a fase de maior aumento da atividade das enzimas digestivas. Entretanto, para a atividade da lipase, o efeito da idade foi diferente para cada alimento. Para aves alimentadas com SITV, a atividade dessa enzima cresceu linearmente com a idade, enquanto nas alimentadas com SIE, FSO e ração, o efeito foi quadrático. Os coeficientes de digestibilidade da matéria seca e extrato etéreo e os valores de energia metabolizável dos tipos de sojas variaram em proporções diferentes em função da idade. Também, observou-se correlação positiva entre a digestibilidade do extrato etéreo e a atividade de lipase. Os valores de energia metabolizável aparente corrigida (EMAn) e verdadeira corrigida (EMVn) determinados para a SIE apresentaram comportamento quadrático em função da idade, ocorrendo aumento da energia metabolizável (EM) até a 3ª semana de idade e diminuindo a partir da 4ª semana. Entretanto, a em da SITV, FSO e da ração não foi afetada pela idade da ave. O aproveitamento da energia dos alimentos varia com a idade das aves, em função de sua dependência da atividade enzimática.This work aimed to investigate the effect of broilers age in the digestibility of nutrients in extruded and toasted full fat soybean (EFFS and TFFS) and soybean meal plus oil (SMO), and in the production of digestive pancreatic enzymes. Five digestibility assays were carried out with broilers of one, two, three, four , and six weeks of age. Total collected excreta methodology was applied. The activities of amylase and pancreatic trypsin enzymes increased linearly with the broilers age, as well as the allometric growth of the pancreas. The highest allometric growth ratio occurred in the second week, coincidentally with the period of increasing activity of the digestive enzymes. However, for lipase activity, distinct age affect was found for each feeds. For birds fed on TFFS, the enzyme activity increased linearly with age, while those fed on EFFS, SMO, and concentrate a quadratic effect was observed. The digestibility coefficients for dry matter and ether extract and the soybeans metabolizable energy (ME) varied at different proportions by age. A positive correlation between ether extract digestibility and lipase activity was also observed. The results of corrected apparent and actual metabolizable energy for EFFS presented a quadratic profile in function of age, increasing ME up to the third week and decreasing after this period. However, ME of TFFS, SMO and concentrate was not affected by chicks age. The nutrient digestibility of feedstuffs varies with chicks age, due to the different enzymatic activities

Carbohydrate metabolism of Xylella fastidiosa: Detection of glycolytic and pentose phosphate pathway enzymes and cloning and expression of the enolase gene

The objective of this work was to assess the functionality of the glycolytic pathways in the bacterium Xylella fastidiosa. To this effect, the enzymes phosphoglucose isomerase, aldolase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase of the glycolytic pathway, and glucose 6-phosphate dehydrogenase of the Entner-Doudoroff pathway were studied, followed by cloning and expression studies of the enolase gene and determination of its activity. These studies showed that X. fastidiosa does not use the glycolytic pathway to metabolize carbohydrates, which explains the increased duplication time of this phytopatogen. Recombinant enolase was expressed as inclusion bodies and solubilized with urea (most efficient extractor), Triton X-100, and TCA. Enolase extracted from X. fastidiosa and from chicken muscle and liver is irreversibly inactivated by urea. The purification of enolase was partial and resulted in a low yield. No enzymatic activity was detected for either recombinant and native enolases, aldolase, and glyceraldehyde-3-phosphate dehydrogenase, suggesting that X. fastidiosa uses the Entner-Doudoroff pathway to produce pyruvate. Evidence is presented supporting the idea that the regulation of genes and the presence of isoforms with regulation patterns might make it difficult to understand the metabolism of carbohydrates in X. fastidiosa

Carbohydrate metabolism of Xylella fastidiosa: Detection of glycolytic and pentose phosphate pathway enzymes and cloning and expression of the enolase gene

The objective of this work was to assess the functionality of the glycolytic pathways in the bacterium Xylella fastidiosa. To this effect, the enzymes phosphoglucose isomerase, aldolase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase of the glycolytic pathway, and glucose 6-phosphate dehydrogenase of the Entner-Doudoroff pathway were studied, followed by cloning and expression studies of the enolase gene and determination of its activity. These studies showed that X. fastidiosa does not use the glycolytic pathway to metabolize carbohydrates, which explains the increased duplication time of this phytopatogen. Recombinant enolase was expressed as inclusion bodies and solubilized with urea (most efficient extractor), Triton X-100, and TCA. Enolase extracted from X. fastidiosa and from chicken muscle and liver is irreversibly inactivated by urea. The purification of enolase was partial and resulted in a low yield. No enzymatic activity was detected for either recombinant and native enolases, aldolase, and glyceraldehyde-3-phosphate dehydrogenase, suggesting that X. fastidiosa uses the Entner-Doudoroff pathway to produce pyruvate. Evidence is presented supporting the idea that the regulation of genes and the presence of isoforms with regulation patterns might make it difficult to understand the metabolism of carbohydrates in X. fastidiosa