The structural origin and biological function of pH-sensitivity in firefly luciferases

Firefly luciferases are called pH-sensitive because their bioluminescence spectra display a typical red-shift at acidic pH, higher temperatures, and in the presence of heavy metal cations, whereas other beetle luciferases (click beetles and railroadworms) do not, and for this reason they are called pH-insensitive. Despite many studies on firefly luciferases, the origin of pH-sensitivity is far from being understood. This subject is revised in view of recent results. Some substitutions of amino-acid residues influencing pH-sensitivity in firefly luciferases have been identified. Sequence comparison, site-directed mutagenesis and modeling studies have shown a set of residues differing between pH-sensitive and pH-insensitive luciferases which affect bioluminescence colors. Some substitutions dramatically affecting bioluminescence colors in both groups of luciferases are clustered in the loop between residues 223-235 (Photinus pyralis sequence). A network of hydrogen bonds and salt bridges involving the residues N229-S284-E311-R337 was found to be important for affecting bioluminescence colors. It is suggested that these structural elements may affect the benzothiazolyl side of the luciferin-binding site affecting bioluminescence colors. Experimental evidence suggest that the residual red light emission in pH-sensitive luciferases could be a vestige that may have biological importance in some firefly species. Furthermore, the potential utility of pH-sensitivity for intracellular biosensing applications is considered. © The Royal Society of Chemistry and Owner Societies.

Anaerobic treatment of wastewater from coffee pulping in upflow anaerobic sludge blanket (UASB) in two stages

In this work, the efficiency of two-stage upflow anaerobic sludge blanket (UASB) reactors was evaluated in bench scale, for treating a liquid effluent from coffee pulping. Hydraulic detention times (HDT) were 4.0; 5.2 and 6.2 days, resulting in organic loading rates (OLR) of 5.8; 3.6 and 3.0g total COD per (L-d) in the first reactor (Rl) and HDT of 2.0; 2.6 and 3.1 days with OLR of 5.8; 0.5 and 0.4 g total COD per (L-d) in the second reactor (R2). The medium values of total COD affluent varied from 15.440 to 23.040 mg O 2/L, and in the effluent to the reactors 1 and 2 were from l.lOO to 11.500 mg 0 2/L and 420 to 9.000 mg O 2/L, respectively. The medium values of removal efficiencies of total COD and TSS varied from 66 to 98% and 93 to 97%, respectively, in the system of treatment with the UASB reactors, in two stages. The content of methane in the biogas varied from 69 to 89% in the Rl and from 52 to 73% in the R2. The maximum volumetric methane production of 0.483 m 3 CH 4per (m 3 reactor d) was obtained with OLR of 3.6 g total COD per (L reactor d) and HDT of 6.2 days in the Rl. The volatile fatty acids concentration was kept below 100mg/L with HDT of 5.2 and 6.2 days in the Rl and HDT of 2.6 and 3.1 days in the R2.

Pyrearinus termitilluminans larval click beetle luciferase: Active site properties, structure and function relationships and comparison with other beetle luciferases

Several beetle luciferases have been cloned and sequenced. However, most studies on structure and function relationships and bioanalytical applications were done with firefly luciferases, which are pH sensitive. Several years ago we cloned Pyrearinus termitilluminans larval click beetle luciferase, which displays the most blue-shifted bioluminescence among beetle luciferases and is pH insensitive. This enzyme was expressed in E. coli, purified, and its properties investigated. This luciferase shows slower luminescence kinetics, KM values comparable to other beetle luciferases and high catalytic constant. Fluorescence studies with 8-anilino-1-naphtalene-sulfonic acid (1,8-ANS) and modeling studies suggest that the luciferin binding site of this luciferase is very hydrophobic, supporting the solvent and orientation polarizability effects as determining mechanisms for bioluminescence colors. Although pH insensitive in the range between pH 6-8, at pH 10 this luciferase displays a remarkable red-shift and broadening of the bioluminescence spectrum. Modeling studies suggest that the residue C312 may play an important role in bioluminescence color modulation. Compared to other beetle luciferases, Pyrearinus termitilluminans luciferase also displays higher thermostability and sustained luminescence in a bacterial cell environment, which makes this luciferase particularly suitable for in vivo cell analysis and bioimaging. © The Royal Society of Chemistry and Owner Societies 2009.

Effect of light energy density on conversion degree and hardness of dual-cured resin cement

This study evaluated the effect of different light energy densities on conversion degree (CD) and Knoop hardness number (KHN) of RelyX ARC (RLX) resin cement. After manipulation according to the manufacturer's instructions, RLX was inserted into a rubber mold (0.8 mm X 5 mm) and covered with a Mylar strip. The tip of the lightcuring unit (LCU) was positioned in contact with the Mylar surface. Quartz-tungsten-halogen (QTH) and light-emitting diode (LED) LCUs with light densities of 10, 20 and 30 J/cm2 were used to light-cure the specimens. After light curing, the specimens were stored dry in lightproof containers at 37°C. After 24 hours, the CD was analyzed by FT-Raman and, after an additional 24-hours, samples were submitted to Knoop hardness testing. The data of the CD (%) and KHN were submitted to two-way ANOVA and the Tukey's test (α=0.05). QTH and LED were effective light curing units. For QTH, there were no differences among the light energy densities for CD or KHN. For LED, there was a significant reduction in CD with the light energy density set at 10 J/cm2. KHN was not influenced by the lightcuring unit and by its light energy density. © Operative Dentistry.

Reactores aeróbios de lecho fluidizado trifásico con circulación interna: Caracterización hidrodinámica y del soporte

Aerobic internal-loop reactors use active biomass attached in a supporting media (biofilm) with the advantage of retaining a big biomass concentration in a small physical space, removing carbonaceous matter and nitrogen in only one reactor. Liquid circulation occurs due to hydrostatic pressure difference produced by air injection in the riser. In biphasic conditions liquid circulation velocities, gas holdup and oxygen transfer coefficient in four different reactor configurations were studied. For the three-phase conditions, the same parameters in just one of those configurations were evaluated. Also, there were three granular supporting media characterized. On the other hand, the relationship between internal and external tube areas and supporting media concentrations influence the liquid velocity, gas holdup and oxygen mass transfer values and some important supporting media characteristics were observed and compared.

Plasma confinement in tokamaks with robust torus

We present a non-linear symplectic map that describes the alterations of the magnetic field lines inside the tokamak plasma due to the presence of a robust torus (RT) at the plasma edge. This RT prevents the magnetic field lines from reaching the tokamak wall and reduces, in its vicinity, the islands and invariant curve destruction due to resonant perturbations. The map describes the equilibrium magnetic field lines perturbed by resonances created by ergodic magnetic limiters (EMLs). We present the results obtained for twist and non-twist mappings derived for monotonic and non-monotonic plasma current density radial profiles, respectively. Our results indicate that the RT implementation would decrease the field line transport at the tokamak plasma edge. © 2010 Elsevier B.V. All rights reserved.

Validação de método de análise quantitativa para a triancinolona a partir de complexo ternário por espectrofotometria de UV-Vis

Triamcinolone (TRI), a drug widely used in the treatment of ocular inflammatory diseases, is practically insoluble in water, which limits its use in eye drops. Cyclodextrins (CDs) have been used to increase the solubility or dissolution rate of drugs. The purpose of the present study was to validate a UV-Vis spectrophotometric method for quantitative analysis of TRI in inclusion complexes with beta-cyclodextrin (B-CD) associated with triethanolamine (TEA) (ternary complex). The proposed analytical method was validated with respect to the parameters established by the Brazilian regulatory National Agency of Sanitary Monitoring (ANVISA). The analytical measurements of absorbance were made at 242nm, at room temperature, in a 1-cm path-length cuvette. The precision and accuracy studies were performed at five concentration levels (4, 8, 12, 18 and 20μg.mL -1). The B-CD associated with TEA did not provoke any alteration in the photochemical behavior of TRI. The results for the measured analytical parameters showed the success of the method. The standard curve was linear (r2 > 0.999) in the concentration range from 2 to 24 μg.mL -1. The method achieved good precision levels in the inter-day (relative standard deviation-RSD <3.4%) and reproducibility (RSD <3.8%) tests. The accuracy was about 80% and the pH changes introduced in the robustness study did not reveal any relevant interference at any of the studied concentrations. The experimental results demonstrate a simple, rapid and affordable UV-Vis spectrophotometric method that could be applied to the quantitation of TRI in this ternary complex.

Use of Physiological Parameters in Screening Drought Tolerance in Sugarcane Genotypes

The physiological response of four commercial sugarcane genotypes to water stress was evaluated by measuring the photochemical efficiency of the photosystem II (chlorophyll a fluorescence ratio, F v/F m), estimated chlorophyll content (SPAD unit), leaf temperature (LT) and leaf relative water content (RWC). A field trial was established in the subtropical area with well-watered and water-stressed genotypes, in completely randomized blocks with four replicates in a 4 × 2 × 3 factorial design (genotype × irrigation × evaluation date). Physiological measurements were done during a 90 day-period of formative stage of plants. The analysis of variance showed that the interaction of genotype × irrigation × evaluation date had a significant effect for three physiological markers tested, F v/F m, SPAD unit and RWC. Under non-stressed conditions, all genotypes showed similar responses for the four markers. Under water deficiency stress, two drought-tolerant genotypes, HOCP01-523 and TCP89-3505 displayed higher values for F v/F m, SPAD unit and RWC, and lower values for LT, and could be classified as tolerant. It is therefore possible to use these physiological water stress associated traits as scorable marker traits for selecting drought-tolerant sugarcane genotypes in future breeding programs. © 2011 Society for Sugar Research & Promotion.

Electrical Power Distribution System modeling with VHDL-AMS for the construction of a Real-Time Digital Simulator using FPGAS devices

This paper presents a distribution feeder simulation using VHDL-AMS, considering the standard IEEE 13 node test feeder admitted as an example. In an electronic spreadsheet all calculations are performed in order to develop the modeling in VHDL-AMS. The simulation results are compared in relation to the results from the well knowing MatLab/Simulink environment, in order to verify the feasibility of the VHDL-AMS modeling for a standard electrical distribution feeder, using the software SystemVision™. This paper aims to present the first major developments for a future Real-Time Digital Simulator applied to Electrical Power Distribution Systems. © 2012 IEEE.

An efficient implementation of tabu search in feeder reconfiguration of distribution systems

This paper presents an efficient tabu search algorithm (TSA) to solve the problem of feeder reconfiguration of distribution systems. The main characteristics that make the proposed TSA particularly efficient are a) the way in which the neighborhood of the current solution was defined; b) the way in which the objective function value was estimated; and c) the reduction of the neighborhood using heuristic criteria. Four electrical systems, described in detail in the specialized literature, were used to test the proposed TSA. The result demonstrate that it is computationally very fast and finds the best solutions known in the specialized literature. © 2012 IEEE.

Effect of experimental photopolymerized coatings on the hydrophobicity of a denture base acrylic resin and on Candida albicans adhesion

Objective: This study investigated the effect of experimental photopolymerized coatings, containing zwitterionic or hydrophilic monomers, on the hydrophobicity of a denture base acrylic resin and on Candida albicans adhesion. Methods: Acrylic specimens were prepared with rough and smooth surfaces and were either left untreated (control) or coated with one of the following experimental coatings: 2-hydroxyethyl methacrylate (HE); 3-hydroxypropyl methacrylate (HP); and 2-trimethylammonium ethyl methacrylate chloride (T); and sulfobetaine methacrylate (S). The concentrations of these constituent monomers were 25%, 30% or 35%. Half of the specimens in each group (control and experimentals) were coated with saliva and the other half remained uncoated. The surface free energy of all specimens was measured, regardless of the experimental condition. C. albicans adhesion was evaluated for all specimens, both saliva conditioned and unconditioned. The adhesion test was performed by incubating specimens in C. albicans suspensions (1 × 10 7 cell/mL) at 37 °C for 90 min. The number of adhered yeasts were evaluated by XTT (2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-5-[{phenylamino} carbonyl]-2H-tetrazolium-hydroxide) method. Results: For rough surfaces, coatings S (30 or 35%) and HP (30%) resulted in lower absorbance values compared to control. These coatings exhibited more hydrophilic surfaces than the control group. Roughness increased the adhesion only in the control group, and saliva did not influence the adhesion. The photoelectron spectroscopy analysis (XPS) confirmed the chemical changes of the experimental specimens, particularly for HP and S coatings. Conclusions: S and HP coatings reduced significantly the adhesion of C. albicans to the acrylic resin and could be considered as a potential preventive treatment for denture stomatitis. © 2012 Elsevier Ltd.

On the application of nanostructured electrodes prepared by Ti/TiO2/WO3 template : A case study of removing toxicity of indigo using visible irradiation

New assays with HepG2 cells indicate that Indigo Carmine (IC), a dye that is widely used as additive in many food and pharmaceutical industries exhibited cytotoxic effects. This work describes the development of a bicomponent nanostructured Ti/TiO2/WO3 electrode prepared by template method and investigates its efficiency in a photoelectrocatalytic method by using visible light irradiation and applied potential of 1V. After 2h of treatment there are reduction of 97% discoloration, 62% of mineralization and formation of three byproducts assigned as: 2-amine-5-sulfo-benzoic acid, 2,3-dioxo-14-indole-5-sulfonic acid, and 2-amino-α-oxo-5-sulfo-benzeneacetic acid were identified by HPLC-MS/MS. But, cytotoxicity was completely removed after 120min of treatment. © 2013 Elsevier Ltd.

Sugarcane bagasse ozonolysis pretreatment: Effect on enzymatic digestibility and inhibitory compound formation

Sugarcane bagasse was pretreated with ozone to increase lignocellulosic material digestibility. Bagasse was ozonated in a fixed bed reactor at room temperature, and the effect of the two major parameters, ozone concentration and sample moisture, was studied. Acid insoluble and total lignin decreased whereas acid soluble lignin increased in all experiments. Pretreatment barely attacked carbohydrates, with cellulose and xylan recovery rates being >92%. Ozonolysis increased fermentable carbohydrate release considerably during enzymatic hydrolysis. Glucose and xylose yields increased from 6.64% and 2.05%, for raw bagasse, to 41.79% and 52.44% under the best experimental conditions. Only xylitol, lactic, formic and acetic acid degradation compounds were found, with neither furfural nor HMF (5-hydroxymethylfurfural) being detected. Washing detoxification provided inhibitor removal percentages above 85%, increasing glucose hydrolysis, but decreasing xylose yield by xylan solubilization. SEM analysis showed structural changes after ozonization and washing. © 2013 Elsevier Ltd.

Electrochemical oxidation of wastewater containing aromatic amines using a flow electrolytic reactor

Aromatic amines are environmental pollutants and represent one of the most important classes of industrial and natural chemicals. Some types of complex effluents containing these chemical species, mainly those originated from chemicals plants are not fully efficiently treated by conventional processes. In this work, the use of electrochemical technology through an electrolytic pilot scale flow reactor is considered for treatment of wastewater of a chemical industry manufacturer of antioxidant and anti-ozonant substances used in rubber. Experimental results showed that was possible to remove between 65% and 95% of apparent colour and chemical oxygen demand removal between 30 and 90% in 60 min of treatment, with energy consumption rate from 26 kWh m-3 to 31 kWh m-3. Absorbance, total organic carbon and toxicity analyses resulted in no formation of toxic by-products. The results suggest that the presented electrochemical process is a suitable method for treating this type of wastewater, mainly when pre-treated by aeration. Copyright © 2013 Inderscience Enterprises Ltd.