Protein extraction and comparison of stain protocols for analysis of two-dimensional electrophoresis gels

Protein extraction and the proteome of Lactobacillus delbrueckii subsp. bulgaricus were studied using different stains. The reversible silver staining technique was shown to be more sensitive than the irreversible silver stain. Coomassie colloidal was demonstrated to be as sensitive as reversible silver stain; however, the Coomassie colloidal blue solution developed a higher background and for sample preparation was more time-consuming.     

Asymmetric alkene epoxidation by Mn(III)salen catalyst in ionic liquids

The enantioselective epoxidation of 6-cyano-2,2-dimethylchromene (Chrom) catalysed by the Jacobsen catalyst, using sodium hypochlorite (NaOCl) as oxygen source, at room temperature, was performed in a series of 1,3-dialkylimidazolium and tetra-alkyl-dimethylguanidium based ionic liquids. All the room temperature ionic liquids (RTILs) could be used as reaction media for the enantioselective epoxidation of the alkene giving, generally, moderate to good epoxide yields and enantiomeric excesses (ee%).For the series of ionic liquids derived from the 1,3-dialkylimidazolium cation, it was observed some relationship between the RTILs physical properties and the catalytic reaction parameters, exemplified by linear correlations between (i) the ee% and the α Kamlet-Taft parameter (hydrogen bond acidity of the solvent) for CH₂Cl₂ and [C₄m_nim][BF₄] ionic liquids (n = 1 or 2), and (ii) the ee% and the β Kamlet-Taft parameter (hydrogen bond basicity of the solvent) for CH₂Cl₂ and [C₄mim][X] ionic liquids (X = PF₆, NTf₂ or BF₄).All the RTILs could be reused in further catalytic cycles, with the exception of [C₈mim][PF₆]. The reutilisation of the Jacobsen catalyst for four times generally led to a decrease in the epoxide yield and to a slight decrease in the enantioselectivity. The recycling of the catalyst could be improved by imparting an ionic character to the complex through abstraction of the axially coordinated chloride anion (Cat 2). Other oxygen sources, such as iodosylbenzene, hydrogen peroxide and urea-hydrogen peroxide adduct, were also tested coupled with Jacobsen catalyst, but the best results were achieved with NaOCl.     

Evaluation of <Emphasis Type="Italic">Fusarium solani</Emphasis> Hyphae and Conidia Susceptibility to Amphotericin B and Itraconazole: Study of a Clinical Case

Fusarium species are hyaline moulds belonging to the hyalohyphomycosis group that are usually found in the soil and plants. This organism has emerged as a cause of disseminated invasive disease. The correlation between in vitro value and clinical efficacy is low and many patients remain unresponsive to treatment despite in vitro susceptibility. We determined growth control for Fusarium solani using the BioCell-Tracer system that measures the growth rate of a single fungal hypha, and the effect of different concentrations of amphotericin B and itraconazole. The MIC for these two drugs was also determined by a broth microdilution technique, using RPMI 1640. Different MICs for amphotericin B were obtained by the two different methods. This paper describes a case of infection due to Fusarium solani in an allogeneic bone marrow transplanted patient, the microbiological diagnostic, antifungal susceptibility tests for conidia and hypha and clinical correlation.     

Simultaneous Saccharification and Fermentation of Hydrothermal Pretreated Lignocellulosic Biomass: Evaluation of Process Performance Under Multiple Stress Conditions

Industrial lignocellulosic bioethanol processes are exposed to different environmental stresses (such as inhibitor compounds, high temperature, and high solid loadings). In this study, a systematic approach was followed where the liquid and solid fractions were mixed to evaluate the influence of varied solid loadings, and different percentages of liquor were used as liquid fraction to determine inhibitor effect. Ethanol production by simultaneous saccharification and fermentation (SSF) of hydrothermally pretreated Eucalyptus globulus wood (EGW) was studied under combined diverse stress operating conditions (30–38 °C, 60–80 g of liquor from hydrothermal treatment or autohydrolysis (containing inhibitor compounds)/100 g of liquid and liquid to solid ratio between 4 and 6.4 g liquid in SSF/g unwashed pretreated EGW) using an industrial Saccharomyces cerevisiae strain supplemented with low-cost byproducts derived from agro-food industry. Evaluation of these variables revealed that the combination of temperature and higher solid loadings was the most significant variable affecting final ethanol concentration and cellulose to ethanol conversion, whereas solid and autohydrolysis liquor loadings had the most significant impact on ethanol productivity. After optimization, an ethanol concentration of 54 g/L (corresponding to 85 % of conversion and 0.51 g/Lh of productivity at 96 h) was obtained at 37 °C using 60 % of autohydrolysis liquor and 16 % solid loading (liquid to solid ratio of 6.4 g/g). The selection of a suitable strain along with nutritional supplementation enabled to produce noticeable ethanol titers in quite restrictive SSF operating conditions, which can reduce operating cost and boost the economic feasibility of lignocellulose-to-ethanol processes.     

Genotoxicity and Cytotoxicity Evaluation of the Neolignan Analogue 2-(4-Nitrophenoxy)-1Phenylethanone and its Protective Effect Against DNA Damage

Neolignans are secondary metabolites found in various groups of Angiosperms. They belong to a class of natural compounds with great diversity of chemical structures and pharmacological activities. These compounds are formed by linking two phenylpropanoid units. Several compounds that have ability to prevent genetic damage have been isolated from plants, and can be used to prevent or delay the development of tumor cells. Genetic toxicology evaluation is widely used in risk assessment of new drugs in preclinical screening tests. In this study, we evaluated the genotoxicity and cytotoxicity of the neolignan analogue 2-(4-nitrophenoxy)-1-phenylethanone (4NF) and its protective effect against DNA damage using the mouse bone marrow micronucleus test and the comet assay in mouse peripheral blood. Our results showed that this neolignan analogue had no genotoxic activity and was able to reduce induced damage both in mouse bone marrow and peripheral blood. Although the neolignan analogue 4NF was cytotoxic, it reduced cyclophosphamide-induced cytotoxicity. In conclusion, it showed no genotoxic action, but exhibited cytotoxic, antigenotoxic, and anticytotoxic activities.     

Elucidating the role of oxidative stress in the therapeutic effect of rutin on experimental acute pancreatitis

Introduction: Acute pancreatitis (AP) may be severe and cause hospitalization or death, and the available treatment is insufficient to control pancreatic inflammation and pain. Rutin is a natural flavonoid with the potential to treat AP via anti-inflammatory, antinociceptive, and antioxidant activities.

Aim: This study investigated the beneficial effects of rutin on experimental AP induced by l-arginine administration in mice.

Methods: The l-arginine-induced AP model was used in Swiss mice (n?=?6–8). Mice submitted to AP induction were treated with rutin (37.5, 75, or 150?mg kg^?1, p.o.) or vehicle (saline) after 24, 36, 48, and 60?h of AP induction. Abdominal hyperalgesia, serum enzymes, interleukin (IL)-6 levels, pancreatic inflammatory parameters, malondialdehyde (MDA) levels, antioxidant enzyme activities, and 3-nitrotyrosine contents were measured 72?h after induction.

Results: Mice submitted to l-arginine injections developed abdominal hyperalgesia and increased serum amylase, lipase, C-reactive protein and IL-6 concentrations; and increased pancreatic myeloperoxidase activity, edema index, MDA, and 3-nitrotyrosine contents. A marked decrease in catalase activity was observed in the pancreas without alterations of superoxide dismutase (SOD) activity compared with the control group. Rutin treatment significantly impaired all the parameters that were altered by AP induction, but increased catalase and SOD activities in the pancreas compared with the vehicle-treated group.

Conclusion: Rutin treatment exerted a protective effect on l-arginine-induced AP by mechanisms involving the reduction of oxidative stress, which suggests that this flavonoid has a potential for future approaches designed for the management of AP.     

Metabolic shift analysis at high cell densities

Abstract: In high cell density cultures it is virtually inevitable that the environment to which the cells are exposed is heterogeneous. Thus, with suspended cultures, individual cells are subject to temporal changes in their environment whereas with aggregated or immobilized cells, the culture can be considered as being formed by a number of subpopulations, each with its own environmental characteristics. In addition, in a high cell density environment, high concentrations of end products may negatively influence the growth rate. This may result in the selection of organisms with an altered metabolic behaviour or with a decreased sensitivity to the adverse effects of the product. We discuss the consequences of this heterogeneity with regard to carbon source metabolism in view of the ability of many bacterial species to adapt to environmental conditions. Selection of variant organisms was found to occur with Clostridium butyricum when grown for a prolonged time in a medium containing approx. I-50 mM glucose. In contrast to the original strain, these variants could sustain a high maximal growth rate in the presence of butyric acid. In addition, they had acquired the capacity to spontaneously form aggregates and were able to carry out a completely solventogenic fermentation. Heterogeneous metabolic activity in aggregated cells is demonstrated with cultures of Lactobacillus laevolacticus , an aggregateforming lactic acid bacterium that converts glucose completely to o-lactate. By using microelectrodes, we show that the fraction of metabolically active cells decreases with increasing aggregate size: in larger aggregates steep pH gradients occur with the effect that only the outer layer of the aggregate is metabolically active, i.e. contributes to lactic acid formation, whereas with smaller aggregates all cells remain active. As a result, the net specific lactic acid production rate of the population as a whole is not invariably increased with increased aggregate size.