Production of a xylose-stimulated beta-glucosidase and a cellulase-free thermostable xylanase by the thermophilic fungus Humicola brevis var. thermoidea under solid state fermentation

Humicola brevis var. thermoidea cultivated under solid state fermentation in wheat bran and water (1:2 w/v) was a good producer of beta-glucosidase and xylanase. After optimization using response surface methodology the level of xylanase reached 5,791.2 +/- A 411.2 U g(-1), while beta-glucosidase production was increased about 2.6-fold, reaching 20.7 +/- A 1.5 U g(-1). Cellulase levels were negligible. Biochemical characterization of H. brevis beta-glucosidase and xylanase activities showed that they were stable in a wide pH range. Optimum pH for beta-glucosidase and xylanase activities were 5.0 and 5.5, respectively, but the xylanase showed 80 % of maximal activity when assayed at pH 8.0. Both enzymes presented high thermal stability. The beta-glucosidase maintained about 95 % of its activity after 26 h in water at 55 A degrees C, with half-lives of 15.7 h at 60 A degrees C and 5.1 h at 65 A degrees C. The presence of xylose during heat treatment at 65 A degrees C protected beta-glucosidase against thermal inactivation. Xylanase maintained about 80 % of its activity after 200 h in water at 60 A degrees C. Xylose stimulated beta-glucosidase activity up to 1.7-fold, at 200 mmol L-1. The notable features of both xylanase and beta-glucosidase suggest that H. brevis crude culture extract may be useful to compose efficient enzymatic cocktails for lignocellulosic materials treatment or paper pulp biobleaching.

Vanillin production from ferulic acid with Pseudomonas fluorescens BF13-1p4

Bioconversion of ferulic acid to vanillin represents an attractive opportunity for replacing synthetic vanillin with a bio-based product, that can be label “natural”, according to current food regulations. Ferulic acid is an abundant phenolic compound in cereals processing by-products, such as wheat bran, where it is linked to the cell wall constituents. In this work, the possibility of producing vanillin from ferulic acid released enzymatically from wheat bran was investigated by using resting cells of Pseudomonas fluorescens strain BF13-1p4 carrying an insertional inactivation of vdh gene and ech and fcs BF13 genes on a low copy number plasmid. Process parameters were optimized both for the biomass production phase and the bioconversion phase using food-grade ferulic acid as substrate and the approach of changing one variable while fixing the others at a certain level followed by the response surface methodology (RSM). Under optimized conditions, vanillin up to 8.46 mM (1.4 g/L) was achieved, whereas highest productivity was 0.53 mmoles vanillin L-1 h-1). Cocktails of a number of commercial enzyme (amylases, xylanases, proteases, feruloyl esterases) combined with bran pre-treatment with steam explosion and instant controlled pressure drop technology were then tested for the release of ferulic acid from wheat bran. The highest ferulic acid release was limited to 15-20 % of the ferulic acid occurring in bran, depending on the treatment conditions. Ferulic acid 1 mM in enzymatic hydrolyzates could be bioconverted into vanillin with molar yield (55.1%) and selectivity (68%) comparable to those obtained with food-grade ferulic acid after purification from reducing sugars with a non polar adsorption resin. Further improvement of ferulic acid recovery from wheat bran is however required to make more attractive the production of natural vanillin from this by-product.

The choice of anesthesia influences oxidative energy metabolism and tissue survival in critically ischemic murine skin

In surgical animal studies anesthesia is used regularly. Several reports in the literature demonstrate respiratory and cardiovascular side effects of anesthesiologic agents. The aim of this study was to compare two frequently used anesthesia cocktails (ketamine/xylazine [KX] versus medetomidine/climazolam/fentanyl [MCF]) in skin flap mouse models. Systemic blood values, local metabolic parameters, and surgical outcome should be analyzed in critical ischemic skin flap models. Systemic hypoxia was found in the animals undergoing KX anesthesia compared with normoxia in the MCF group (sO(2): 89.2% +/- 2.4% versus 98.5% +/- 1.2%, P < 0.01). Analysis of tissue metabolism revealed impaired anaerobic oxygen metabolism and increased cellular damage in critical ischemic flap tissue under KX anesthesia (lactate/pyruvate ratio: KX 349.86 +/- 282.38 versus MCF 64.53 +/- 18.63; P < 0.01 and glycerol: KX 333.50 +/- 83.91 micromol/L versus MCF 195.83 +/- 29.49 micromol/L; P < 0.01). After 6 d, different rates of flap tissue necrosis could be detected (MCF 57% +/- 6% versus KX 68% +/- 6%, P < 0.01). In summary we want to point out that the type of anesthesia, the animal model and the goal of the study have to be well correlated. Comparing the effects of KX and MCF anesthesia in mice on surgical outcome was a novel aspect of our study.

The basel cocktail for simultaneous phenotyping of human cytochrome P450 isoforms in plasma, saliva and dried blood spots.

BACKGROUND AND OBJECTIVE Phenotyping cocktails use a combination of cytochrome P450 (CYP)-specific probe drugs to simultaneously assess the activity of different CYP isoforms. To improve the clinical applicability of CYP phenotyping, the main objectives of this study were to develop a new cocktail based on probe drugs that are widely used in clinical practice and to test whether alternative sampling methods such as collection of dried blood spots (DBS) or saliva could be used to simplify the sampling process. METHODS In a randomized crossover study, a new combination of commercially available probe drugs (the Basel cocktail) was tested for simultaneous phenotyping of CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6 and CYP3A4. Sixteen subjects received low doses of caffeine, efavirenz, losartan, omeprazole, metoprolol and midazolam in different combinations. All subjects were genotyped, and full pharmacokinetic profiles of the probe drugs and their main metabolites were determined in plasma, dried blood spots and saliva samples. RESULTS The Basel cocktail was well tolerated, and bioequivalence tests showed no evidence of mutual interactions between the probe drugs. In plasma, single timepoint metabolic ratios at 2 h (for CYP2C19 and CYP3A4) or at 8 h (for the other isoforms) after dosing showed high correlations with corresponding area under the concentration-time curve (AUC) ratios (AUC0-24h parent/AUC0-24h metabolite) and are proposed as simple phenotyping metrics. Metabolic ratios in dried blood spots (for CYP1A2 and CYP2C19) or in saliva samples (for CYP1A2) were comparable to plasma ratios and offer the option of minimally invasive or non-invasive phenotyping of these isoforms. CONCLUSIONS This new combination of phenotyping probe drugs can be used without mutual interactions. The proposed sampling timepoints have the potential to facilitate clinical application of phenotyping but require further validation in conditions of altered CYP activity. The use of DBS or saliva samples seems feasible for phenotyping of the selected CYP isoforms.

Estudos de proteômica, estruturais e funcionais de proteínas envolvidas na degradação da biomassa lignocelulósica: expansinas microbianas e hidrolases de glicosídeos termofílicas

O Brasil possui uma posição privilegiada quando se refere à produção de etanol. Por questões históricas e geográficas o país é responsável por mais de 30 % da produção mundial de etanol, com uma produção nacional de mais de 28 bilhões de litros em 2014. Para maximizar o rendimento desse processo, está em desenvolvimento a tecnologia associada ao etanol de segunda geração ou etanol lignocelulósico. Os principais desafios desta tecnologia são: melhorar a eficiência de conversão do substrato em produto e a produção em grande escala utilizando substratos de baixo custo. Com o objetivo de melhorar a eficiência do processo de conversão foram estudadas proteínas auxiliares (expansinas) que, em conjunto com celulases, melhoram a despolimerização de biomassa lignocelulósica em açúcares fermentescíveis. Além disso, realizou-se também a caracterização de enzimas ativas de carboidratos (CAZymes) de origem termofílica do organismo Thermogemmatispora sp. T81, devido a capacidade que estas proteínas apresentam de manter a atividade e conformação estrutural em altas temperaturas por um prolongado período de tempo. A partir de análises utilizando bioinformática, os genes que codificam para expansinas de Xanthomonas campestris, Bacillus licheniformis e Trichoderma reesei foram clonados e expressos em E. coli, e seus produtos gênicos (as expansinas) tiveram seus índices de sinergismo (devido atuação conjunta com coquetéis comerciais) e atividade catalítica determinados. Adicionalmente, dispondo de alinhamentos estruturais, foi proposto um mecanismo hidrolítico para elas. Em relação à bactéria Thermogemmatispora sp. T81, foram realizadas análises genômicas e proteômicas, a fim de selecionar enzimas superexpressas em meio celulósico. Seus genes foram clonados heterologamente em E. coli e o produto de expressão caracterizado bioquimicamente (cromatografia, ensaios de atividade e perfil de hidrólise) e estruturalmente (SAXS e dicroísmo circular). Os índices de sinergismo determinados foram de 2,47; 1,96 e 2,44 para as expansinas de Xanthomonas campestris, Bacillus licheniformis e Trichoderma reesei, respectivamente. A partir dos alinhamentos estruturais foi proposto a díade Asp/Glu como sitio catalítico em expansinas. As análises de proteômica possibilitaram a seleção de quatro alvos de clonagem, por apresentarem alto índice de expressão quando a bactéria foi cultivada em meio celulósico. Estas proteínas foram caracterizadas quanto a atividade e apresentaram um perfil comum: temperatura ótima de ação (de 70 a 75 °C), pH ótimo de 5, e hidrolisam preferencialmente substratos hemicelulósicos (xilano). A porcentagem de estruturais secundárias das proteínas em estudo foram confirmadas com predições teóricas ao se utilizar a técnica de dicroísmo circular. Desta maneira, os objetivos iniciais propostos neste projeto foram concluídos com a determinação do grau de sinergismo das proteínas expansinas em estudo e a proposição de um mecanismo de hidrólise para as mesmas, considerando que tais proteínas por mais de 20 anos tiveram sua atividade definida exclusivamente como acessória. Além disso, este estudo contribui com a identificação e seleção de genes para CAZymes termofilícas com aplicação biotecnológica devido às propriedades termoestáveis apresentadas.

Toxin insights into nicotinic acetylcholine receptors

Venomous species have evolved cocktails of bioactive peptides to facilitate prey capture. Given their often exquisite potency and target selectivity, venom peptides provide unique biochemical tools for probing the function of membrane proteins at the molecular level. in the field of the nicotinic acetylcholine receptors (nAChRs), the subtype specific snake alpha-neurotoxins and cone snail alpha-conotoxins have been widely used to probe receptor structure and function in native tissues and recombinant systems. However, only recently has it been possible to generate an accurate molecular view of these nAChR-toxin interactions. Crystal structures of AChBP, a homologue of the nAChR ligand binding domain, have now been solved in complex with alpha-cobratoxin, alpha-conotoxin PnIA and alpha-conotoxin Iml. The orientation of all three toxins in the ACh binding site confirms many of the predictions obtained from mutagenesis and docking simulations on homology models of mammalian nAChR. The precise understanding of the molecular determinants of these complexes is expected to contribute to the development of more selective nAChR modulators. In this commentary, we review the structural data on nAChR-toxin interactions and discuss their implications for the design of novel ligands acting at the nAChR. (c) 2006 Elsevier Inc. All rights reserved.

Adult Alternatives for Social Drinking: A Direction

In their survey/study - Adult Alternatives for Social Drinking: A Direction - by John Dienhart and Sandra Strick, Assistant Professors, Department of Restaurant, Hotel and Institutional Management, Purdue University, Dienhart and Strick begin with: “Changes in consumer habits have brought about a change in the business of selling alcoholic drinks and have impacted upon hotel food and beverage operations. The authors surveyed a sample of hotel corporate food and beverage directors to ascertain how they are handling this challenge.” Dienhart and Strick declare that the alcoholic beverage market, sale and consumption thereof, has taken a bit of a hit in contemporary society. “Even to the casual observer, it's obvious that the bar and beverage industry has undergone a great deal of change in the past few years,” say the authors. “Observations include a change in the types of drinks people are ordering, as well as a decrease in the number of drinks being sold,” they qualify. Dienhart and Strick allude to an increase in the federal excise tax, attacks from alcohol awareness groups, the diminished capacity of bars and restaurants to offer happy hours, increased liability insurance premiums as well as third-party liability issues, and people’s awareness of their own mortality as some of the reasons for the change. To quantify some empirical data on beverage consumption the Restaurant, Hotel, and Institutional Management Department of Purdue University conducted a study “… to determine if observed trends could be documented with hard data.” In regards to the subject, the study asks and answers a lot of interesting questions with the results presented to concerned followers via percentages. Typical of the results are: “When asked whether the corporation experienced a change in alcoholic sales in the past year, 67 percent reported a decrease in the amount of alcohol sold.” “Sixty-two percent of the respondents reported an increase in non-alcoholic sales over the past year. The average size of the increase was 8 percent. What Dienhart and Strick observe is that the decrease in alcoholic beverage consumption has resulted in a net increase for non-alcoholic beverage consumption. What are termed specialty drinks are gaining a foothold in the market, say the authors. “These include traditional cocktails made with alcohol-free products, as well as creative new juice based drinks, cream based drinks, carbonated beverages, and heated drinks,” say Dienhart and Strick by way of citation . Another result of the non-alcoholic consumption trend is the emergence of some novel marketing approaches by beer, wine, and spirits producers, including price increases on their alcohol based beverages as well as the introduction of faux alcoholic drinks like non-alcoholic beer and wine. Who or what is the big winner in all of this? That distinction might go to bottled water!

Modular approach to select bacteriophages targeting Pseudomonas aeruginosa for their application to children suffering with cystic fibrosis

This review discusses the potential application of bacterial viruses (phage therapy) towards the eradication of antibiotic resistant Pseudomonas aeruginosa in children with cystic fibrosis (CF). In this regard, several potential relationships between bacteria and their bacteriophages are considered. The most important aspect that must be addressed with respect to phage therapy of bacterial infections in the lungs of CF patients is in ensuring the continuity of treatment in light of the continual occurrence of resistant bacteria. This depends on the ability to rapidly select phages exhibiting an enhanced spectrum of lytic activity among several well-studied phage groups of proven safety. We propose a modular based approach, utilizing both mono-species and hetero-species phage mixtures. With an approach involving the visual recognition of characteristics exhibited by phages of well studied phage groups on lawns of the standard P. aeruginosa PAO1 strain, the simple and rapid enhancement of the lytic spectrum of cocktails is permitted, allowing the development of tailored preparations for patients capable of circumventing problems associated with phage resistant bacterial mutants.

A Case Against Fancy Decorated Drinks: Multi-Organ Failure After Drinking a Mojito Cocktail

We present the first case of gastro-intestinal perforation caused by a mint twig decorating a cocktail drink. A 76-year-old man was enjoying his Mojito cocktail on a cruise ship near Mexico when he accidently swallowed a mint twig, resulting in ileum perforation. This led to a cascade of events, eventually resulting in life-threatening multi-organ failure. Given this rare but potentially severe complication and the increasing popularity of decorated drinks, a less ‘fancy’ presentation for cocktails and similar drinks may be warranted.

Estructuración de los procesos y la función de recursos humanos alineados con el direccionamiento estratégico en Kathmandu restaurante

El direccionamiento estratégico está destinado para que las organizaciones cumplan con sus metas y tengan perdurabilidad en el tiempo; es una forma metodológica en la cual se establecen La formulación y el planteamiento estratégicos, se identifican las brechas entre el sueño de la empresa y su realidad, para desplegar la plata forma estratégica. Kathmandu es un restaurante conocido por su diversidad cultural gastronómica, ubicado en Usaquén al Norte de Bogotá. Su serviespacio reúne los sabores, aromas, decoración y elixires del oriente, su música incluye una fusión de sonidos que permite materializar la experiencia que quiere brindar para satisfacer las expectativas de sus comensales y brindarles una experiencia diferente. Los fines de semana presentan shows de danza de vientre, para acompañar la comida y el rato que dispongan disfrutar en este lugar. Este restaurante tiene un diferenciador en su servicio, que le ofrece una ventaja competitiva, Kathmandu brinda la posibilidad de que además del deleite de sus platos, los clientes puedan adquirir toda la decoración de este lugar, estas son artesanías originales traídas de Nepal, India, Tailandia, Indonesia y sus alrededores. Otro atrayente de este restaurante son los cocteles alusivos a las regiones del oriente, y la narguile que ofrece a sus clientes. Al pasar de los años, ciertos inconvenientes presentados en el negocio, con los clientes, el personal y las utilidades, han hecho que los gerentes se preocupen por encontrar la raíz del problema y buscar la solución óptima para perdurar en el tiempo con los mejores beneficios posibles, pues las estrategias que estaban implementando no van acorde a su misión y visión, por lo que es necesario realizar una restructuración de ciertos procesos enfocados al direccionamiento estratégico. Se realizó un análisis en una reunión junto a las directivas sobre cada área de la organización donde se evidenciaron problemas que deben ser tratados para hacer de Kathmandu una empresa más eficiente, eficaz y productiva. Y prepararla para afrontar los cambios que el tiempo le presente, en cuanto a preferencias de los clientes, competencia, entre otras, bien sean a corto, mediano o largo plazo. Algunos de los principales problemas encontrados son: el servicio al cliente, la selección adecuada de personal y capacitación de este, la falta de manejo de un conducto jerárquico y repartición de funciones entre los colaboradores, el informe adecuado sobre la inconformidad de los clientes con su solución, el manual de funciones, el manejo de inventarios y su rotación, verificación de calidad de los productos, manejo de proveedores, entre otros. Kathmandu, a pesar de tener problemas evidentes, es una empresa que por los servicios y productos que ofrece sigue generando buenas utilidades, por esto los gerentes no le habían prestado la debida atención a estos inconvenientes que están poniendo en juego la perdurabilidad de la empresa y que afecta la optimización de las utilidades y minimización de costes sin afectar la calidad brindada al cliente. El objetivo de este restaurante es que el área de recursos humanos sea restructurada en cada uno de los procesos para articularla al direccionamiento estratégico de Kathmandu; pues en esta área se generan la mayoría de los problemas presentados anteriormente. El presente proyecto pretende diseñar el direccionamiento estratégico del área recursos humanos del restaurante Kathmandu para superar la problemática identificada y garantizar la efectividad de sus procesos y adicionalmente evidenciar la creencia de éxito sostenido que se manejaba en este lugar. Este proyecto se realizara con una metodología mixta, para tener aspectos de la investigación cuantitativa y cualitativa y poder realizar una investigación más completa que lleve a una óptima solución del problema abarcado. Se utilizara el direccionamiento estratégico como herramienta para replantear el área de recursos humanos de la organización y alcanzar la efectividad en sus procesos.

Soil management systems for sustainable melon cropping in the Submedian of the São Francisco Valley.

Changes in soils management systems, including the application of green manure, are able to increase crop productivity. The aim of this study was to propose a soil management system with the use of green manure to improve the nutritional status and melon productivity in the submedian of the São Francisco Valley. The experiment was installed in Typic Plinthustalf and conducted in split plot. There were two soil tillage systems, tillage (T) and no tillage (NT), and three types of green manure (two vegetal cocktails: VC1- 75% legumes (L) + 25% non-legumes (NL); VC2- 25% L+ 75% NL and spontaneous vegetation (SV)). The experimental design was a randomised block with four replications. Fourteen species of legumes, grasses and oilseeds were used for the composition of the plant cocktails. We evaluated production of the dry shoot and root biomass and carbon and nutrient accumulation by green manures and melon plant. Data were subjected to analysis of variance and the treatment means were compared by Tukey´s test (P<0.05). Shoot biomass production and carbon and nutrient accumulation were higher in plant mixtures compared to spontaneous vegetation. The root system of the plant cocktails added larger quantities of biomass and nutrients to the soil to a depth of 0.60 m when compared to the spontaneous vegetation. The cultivation of plant cocktails with soil tillage, regardless of their composition, is a viable alternative for adding biomass and nutrients to the soil in melon crops in semi-arid conditions, providing productivity increases.