Phenol biodegradation by a microbial consortium: application of artificial neural network (ANN) modelling

In this study, an effective microbial consortium for the biodegradation of phenol was grown under different operational conditions, and the effects of phosphate concentration (1.4 g L-1, 2.8 g L-1, 4.2 g L-1), temperature (25 degrees C, 30 degrees C, 35 degrees C), agitation (150 rpm, 200 rpm, 250 rpm) and pH (6, 7, 8) on phenol degradation were investigated, whereupon an artificial neural network (ANN) model was developed in order to predict degradation. The learning, recall and generalization characteristics of neural networks were studied using data from the phenol degradation system. The efficiency of the model generated by the ANN was then tested and compared with the experimental results obtained. In both cases, the results corroborate the idea that aeration and temperature are crucial to increasing the efficiency of biodegradation.

Bilayer graphene on h-BN substrate: investigating the breakdown voltage and tuning the bandgap by electric field

By performing density functional theory calculations we show that it is possible to make the electronic bandgap in bilayer graphene supported on hexagonal boron nitride (h-BN) substrates tunable. We also show that, under applied electric fields, it is possible to insert states from h-BN into the bandgap, which generate a conduction channel through the substrate making the system metallic. In addition, we verify that the breakdown voltage strongly depends on the number of h-BN layers. We also show that both the breakdown voltage and the bandgap tuning are independent of the h-BN stacking order.

Angiotensin-(3-4) counteracts the Angiotensin II inhibitory action on renal Ca2+-ATPase through a cAMP/PKA pathway

We recently demonstrated that Angiotensin-(3-4) [Ang-(3-4)], an Ang II-derived dipeptide, overcomes inhibition of plasma membrane Ca2+-ATPase promoted by nanomolar concentrations of Ang II in basolateral membranes of renal proximal tubule cells, with involvement of a so far unknown AT(2)R-dependent and NO-independent mechanism. The present study investigates the signaling pathway triggered by Ang-(3-4) that is responsible for counteracting the inhibitory effect of Ang II, and attempts to elucidate the functional interaction of the dipeptide with Ang II at the level of AT(2)R. Stimulation by cholera toxin of G(s)alpha protein structurally linked to AT(2)R as revealed by their co-immunoprecipitation mimicked the effect of Ang-(3-4) on Ca2+-ATPase activity. Furthermore, addition of dibutyril-cAMP (db-cAMP) mimicked Ang-(3-4), whereas the specific PKA inhibitor, PKAi((5-24)) peptide, suppressed the counter-regulatory effect of Ang-(3-4) and the AT(2)R agonist, CGP42112A. Membrane-associated PKA activity was stimulated by Ang-(3-4) or CGP42112A to comparable levels as db-cAMP, and the Ang-(3-4) effect was abrogated by the AT(2)R antagonist PD123319, whereas the AT(1)R antagonist Losartan had no effect. Ang-(3-4) stimulated PKA-mediated phosphorylation of Ca2+-ATPase and activated PKA to comparable levels. Binding assays demonstrated that Ang-(3-4) could not displace H-3-Ang II from HEK 293T cells expressing AT(2)R, but 10(-10) mol/L Ang-(3-4) resulted in the appearance of a probable higher-affinity site (picomolar range) for Ang II. The results presented herein demonstrate that Ang-(3-4), acting as an allosteric enhancer, suppresses Ang II-mediated inhibition of Ca2+-ATPase through an AT(2)R/cAMP/PKA pathway, after inducing conformational changes in AT(2)R that results in generation of higher-affinity sites for Ang II. (C) 2012 Elsevier B.V. All rights reserved.

Inhibitory activity of liposomal flavonoids during oxidative metabolism of human neutrophils upon stimulation with immune complexes and phorbol ester

Context and objective: The massive production of reactive oxygen species by neutrophils during inflammation may cause damage to tissues. Flavonoids act as antioxidants and have anti-inflammatory effects. In this study, liposomes loaded with these compounds were evaluated as potential antioxidant carriers, in attempt to overcome their poor solubility and stability. Materials and methods: Liposomes containing quercetin, myricetin, kaempferol or galangin were prepared by the ethanol injection method and analyzed as inhibitors of immune complex (IC) and phorbol ester-stimulated neutrophil oxidative metabolism by luminol (CLlum) and lucigenin-enhanced (CLluc) chemiluminescence (CL) assays. The mechanisms involved this activity of liposomal flavonoids, such as cytotoxicity and superoxide anion scavenging capacity, and their effect on phagocytosis of ICs were also investigated. Results and discussion: The results showed that the inhibitory effect of liposomal flavonoids on CLlum and CLluc is inversely related to the number of hydroxyl groups in the flavonoid B ring. Moreover, phagocytosis of liposomes by neutrophils does not seem to necessarily promote such activity, as the liposomal flavonoids are also able to reduce CL when the cells are pretreated with cytochalasin B. Under assessed conditions, the antioxidant liposomes are not toxic to the human neutrophils and do not interfere with IC-induced phagocytosis. Conclusion: The studied liposomes can be suitable carriers of flavonoids and be an alternative for the treatment of diseases in which a massive oxidative metabolism of neutrophils is involved.

Sensitivity of Eisenia andrei (Annelida, Oligochaeta) to a commercial formulation of abamectin in avoidance tests with artificial substrate and natural soil under tropical conditions

Obtaining ecotoxicological data on pesticides in tropical regions is imperative for performing more realistic risk analysis, and avoidance tests have been proposed as a useful, fast and cost-effective tool. Therefore, the present study aimed to evaluate the avoidance behavior of Eisenia andrei to a formulated product, Vertimec(A (R)) 18 EC (a.i abamectin), in tests performed on a reference tropical artificial soil (TAS), to derive ecotoxicological data on tropical conditions, and a natural soil (NS), simulating crop field conditions. In TAS tests an adaptation of the substrate recommended by OECD and ISO protocols was used, with residues of coconut fiber as a source of organic matter. Concentrations of the pesticide on TAS test ranged from 0 to 7 mg abamectin/kg (dry weight-d.w.). In NS tests, earthworms were exposed to samples of soils sprayed in situ with: 0.9 L of Vertimec(A (R)) 18 EC/ha (RD); twice as much this dosage (2RD); and distilled water (Control), respectively, and to 2RD: control dilutions (12.5, 25, 50, 75%). All tests were performed under 25 +/- A 2A degrees C, to simulate tropical conditions, and a 12hL:12hD photoperiod. The organisms avoided contaminated TAS for an EC50,48h = 3.918 mg/kg soil d.w., LOEC = 1.75 mg/kg soil d.w. and NOEC = 0.85 mg/kg soil d.w. No significant avoidance response occurred for any NS test. Abamectin concentrations in NS were rather lower than EC50, 48h and LOEC determined in TAS tests. The results obtained contribute to overcome a lack of ecotoxicological data on pesticides under tropical conditions, but more tests with different soil invertebrates are needed to improve pesticides risk analysis.

Shear bond strength between Ni-Cr alloy bonded to a ceramic substrate

Shear bond strength between Ni-Cr alloy bonded to a ceramic substrate Introduction: The aim of this study was to evaluate the shear bond strength between a Ni-Cr alloy and a ceramic system submitted or not to thermocycling. Materials and methods: Forty-eight cylinder blocks of Ni-Cr with 3.0 mm diameter by 4.0 mm hight and 48 disc-shaped specimens (7.0 mm in diameter by 2.0 mm thick) composed of ceramic were prepared. The Ni-Cr cylinder blocks were randomised in two groups of 24 specimens each. One group was submitted to air-particle abrasion (sandblasting) with 50 mu m Al2O3 (0.4-0.7 MPa) during 20 s, and the other group was submitted to mechanical retentions with carbide burrs. Each group was subdivided into other two groups (n = 12), submitted or not to thermocycling (500 cycles, 5-55 degrees C). The cylinder blocks were bonded to the disc-shaped ceramic specimens under 10 N of load. The shear bond strengths (MPa) were measured using a universal testing machine at a cross head speed of 0.5 mm/min and 200 kgf of load. The data were submitted to statistical analysis (ANOVA and Tukey's test). Results: The air-particle abrasion group exhibited significantly higher shear bond strength when compared to drilled group (p < 0.05). Conclusions: Thermocycling decreased significantly the bond strengths for all groups tested.

Identification of Regions Involved in Substrate Binding and Dimer Stabilization within the Central Domains of Yeast Hsp40 Sis1

Protein folding, refolding and degradation are essential for cellular life and are regulated by protein homeostatic processes such those that involve the molecular chaperone DnaK/Hsp70 and its co-chaperone DnaJ. Hsp70 action is initiated when proteins from the DnaJ family bind an unfolded protein for delivery purposes. In eukaryotes, the DnaJ family can be divided into two main groups, Type I and Type II, represented by yeast cytosolic Ydj1 and Sis1, respectively. Although sharing some unique features both members of the DnaJ family, Ydj1 and Sis1 are structurally and functionally distinct as deemed by previous studies, including the observation that their central domains carry the structural and functional information even in switched chimeras. In this study, we combined several biophysical tools for evaluating the stability of Sis1 and mutants that had the central domains (named Gly/Met rich domain and C-terminal Domain I) deleted or switched to those of Ydj1 to gain insight into the role of these regions in the structure and function of Sis1. The mutants retained some functions similar to full length wild-type Sis1, however they were defective in others. We found that: 1) Sis1 unfolds in at least two steps as follows: folded dimer to partially folded monomer and then to an unfolded monomer. 2) The Gly/Met rich domain had intrinsically disordered characteristics and its deletion had no effect on the conformational stability of the protein. 3) The deletion of the C-terminal Domain I perturbed the stability of the dimer. 4) Exchanging the central domains perturbed the conformational stability of the protein. Altogether, our results suggest the existence of two similar subdomains in the C-terminal domain of DnaJ that could be important for stabilizing each other in order to maintain a folded substrate-binding site as well as the dimeric state of the protein.

Effect of Substrate Concentration on Dark Fermentation Hydrogen Production Using an Anaerobic Fluidized Bed Reactor

The effect of substrate (glucose) concentration on the stability and yield of a continuous fermentative process that produces hydrogen was studied. Four anaerobic fluidized bed reactors (AFBRs) were operated with a hydraulic retention time (HRT) from 1 to 8 h and an influent glucose concentration from 2 to 25 gL(-1). The reactors were inoculated with thermally pre-treated anaerobic sludge and operated at a temperature of 30 degrees C with an influent pH around 5.5 and an effluent pH of about 3.5. The AFBRs with a HRT of 2 h and a feed strength of 2, 4, and 10 gL(-1) showed satisfactory H-2 production performance, but the reactor fed with 25 gL(-1) of glucose did not. The highest hydrogen yield value was obtained in the reactor with a glucose concentration of 2 gL(-1) when it was operated at a HRT of 2 h. The maximum hydrogen production rate value was achieved in the reactor with a HRT of 1 h and a feed strength of 10 gL(-1). The AFBRs operated with glucose concentrations of 2 and 4 gL(-1) produced greater amounts of acetic and butyric acids, while AFBRs with higher glucose concentrations produced a greater amount of solvents.

Insights into Phosphate Cooperativity and Influence of Substrate Modifications on Binding and Catalysis of Hexameric Purine Nucleoside Phosphorylases

The hexameric purine nucleoside phosphorylase from Bacillus subtilis (BsPNP233) displays great potential to produce nucleoside analogues in industry and can be exploited in the development of new anti-tumor gene therapies. In order to provide structural basis for enzyme and substrates rational optimization, aiming at those applications, the present work shows a thorough and detailed structural description of the binding mode of substrates and nucleoside analogues to the active site of the hexameric BsPNP233. Here we report the crystal structure of BsPNP233 in the apo form and in complex with 11 ligands, including clinically relevant compounds. The crystal structure of six ligands (adenine, 2'deoxyguanosine, aciclovir, ganciclovir, 8-bromoguanosine, 6-chloroguanosine) in complex with a hexameric PNP are presented for the first time. Our data showed that free bases adopt alternative conformations in the BsPNP233 active site and indicated that binding of the co-substrate (2'deoxy) ribose 1-phosphate might contribute for stabilizing the bases in a favorable orientation for catalysis. The BsPNP233-adenosine complex revealed that a hydrogen bond between the 5' hydroxyl group of adenosine and Arg(43*) side chain contributes for the ribosyl radical to adopt an unusual C3'-endo conformation. The structures with 6-chloroguanosine and 8-bromoguanosine pointed out that the Cl-6 and Br-8 substrate modifications seem to be detrimental for catalysis and can be explored in the design of inhibitors for hexameric PNPs from pathogens. Our data also corroborated the competitive inhibition mechanism of hexameric PNPs by tubercidin and suggested that the acyclic nucleoside ganciclovir is a better inhibitor for hexameric PNPs than aciclovir. Furthermore, comparative structural analyses indicated that the replacement of Ser(90) by a threonine in the B. cereus hexameric adenosine phosphorylase (Thr(91)) is responsible for the lack of negative cooperativity of phosphate binding in this enzyme.

Two structurally discrete GH7-cellobiohydrolases compete for the same cellulosic substrate fiber

Background: Cellulose consisting of arrays of linear beta-1,4 linked glucans, is the most abundant carbon-containing polymer present in biomass. Recalcitrance of crystalline cellulose towards enzymatic degradation is widely reported and is the result of intra-and inter-molecular hydrogen bonds within and among the linear glucans. Cellobiohydrolases are enzymes that attack crystalline cellulose. Here we report on two forms of glycosyl hydrolase family 7 cellobiohydrolases common to all Aspergillii that attack Avicel, cotton cellulose and other forms of crystalline cellulose. Results: Cellobiohydrolases Cbh1 and CelD have similar catalytic domains but only Cbh1 contains a carbohydrate-binding domain (CBD) that binds to cellulose. Structural superpositioning of Cbh1 and CelD on the Talaromyces emersonii Cel7A 3-dimensional structure, identifies the typical tunnel-like catalytic active site while Cbh1 shows an additional loop that partially obstructs the substrate-fitting channel. CelD does not have a CBD and shows a four amino acid residue deletion on the tunnel-obstructing loop providing a continuous opening in the absence of a CBD. Cbh1 and CelD are catalytically functional and while specific activity against Avicel is 7.7 and 0.5 U. mg prot-1, respectively specific activity on pNPC is virtually identical. Cbh1 is slightly more stable to thermal inactivation compared to CelD and is much less sensitive to glucose inhibition suggesting that an open tunnel configuration, or absence of a CBD, alters the way the catalytic domain interacts with the substrate. Cbh1 and CelD enzyme mixtures on crystalline cellulosic substrates show a strong combinatorial effort response for mixtures where Cbh1 is present in 2: 1 or 4: 1 molar excess. When CelD was overrepresented the combinatorial effort could only be partially overcome. CelD appears to bind and hydrolyze only loose cellulosic chains while Cbh1 is capable of opening new cellulosic substrate molecules away from the cellulosic fiber. Conclusion: Cellobiohydrolases both with and without a CBD occur in most fungal genomes where both enzymes are secreted, and likely participate in cellulose degradation. The fact that only Cbh1 binds to the substrate and in combination with CelD exhibits strong synergy only when Cbh1 is present in excess, suggests that Cbh1 unties enough chains from cellulose fibers, thus enabling processive access of CelD.

From otoacoustic emission to late auditory potentials P300: the inhibitory effect

This study verifies the effects of contralateral noise on otoacoustic emissions and auditory evoked potentials. Short, middle and late auditory evoked potentials as well as otoacoustic emissions with and without white noise were assessed. Twenty-five subjects, normal-hearing, both genders, aged 18 to 30 years, were tested. In general, latencies of the various auditory potentials were increased at noise conditions, whereas amplitudes were diminished at noise conditions for short, middle and late latency responses combined in the same subject. The amplitude of otoacoustic emission decreased significantly in the condition with contralateral noise in comparison to the condition without noise. Our results indicate that most subjects presented different responses between conditions (with and without noise) in all tests, thereby suggesting that the efferent system was acting at both caudal and rostral portions of the auditory system.

Influence of the Substrate and Precursor on the Magnetic and Magneto-transport Properties in Magnetite Films

We have investigated the magnetic and transport properties of nanoscaled Fe3O4 films obtained from Chemical Vapor Deposition (CVD) technique using [(FeFe2III)-Fe-II(OBut)(8)] and [Fe-2(III)(OBut)(6)] precursors. Samples were deposited on different substrates (i.e., MgO (001), MgAl2O4 (001) and Al2O3 (0001)) with thicknesses varying from 50 to 350 nm. Atomic Force Microscopy analysis indicated a granular nature of the samples, irrespective of the synthesis conditions (precursor and deposition temperature, T-pre) and substrate. Despite the similar morphology of the films, magnetic and transport properties were found to depend on the precursor used for deposition. Using [(FeFe2III)-Fe-II(OBut)(8)] as precursor resulted in lower resistivity, higher M-S and a sharper magnetization decrease at the Verwey transition (T-V). The temperature dependence of resistivity was found to depend on the precursor and T-pre. We found that the transport is dominated by the density of antiferromagnetic antiphase boundaries (AF-APB's) when [(FeFe2III)-Fe-II(OBut)(8)] precursor and T-pre = 363 K are used. On the other hand, grain boundary-scattering seems to be the main mechanism when [Fe-2(III)(OBut)(6)] is used. The Magnetoresistance (MR(H)) displayed an approximate linear behavior in the high field regime (H > 796 kA/m), with a maximum value at room-temperature of similar to 2-3 % for H = 1592 kA/m, irrespective from the transport mechanism.

Growth of seedlings Schizolobium amazonicum (Huber ex Ducke) on substrate fertilized with nitrogen, phosphorus and potassium

The nutritional management of seedlings in the nursery is one of the most important practices that influence seedling quality. The aim of this work was to evaluate the effect of nitrogen, phosphorus and potassium on the development of Schizolobium amazonicum seedlings grown in 250 cm(3) containers with a commercial substrate in the North of Mato Grosso State, Brazil. The experimental design was completely randomized design with five treatments and five replications, each replication being represented by 24 seedlings. The treatments were: control (only commercial substrate); nitrogen fertilization (150 g m(-3) N using ammonium sulfate + 1.0 kg of ammonium sulfate dissolved in 100 L of water and applied in coverage); phosphorus fertilization (300 g P2O5 m(-3) using simple superphosphate); potassium fertilization (100 g m(-3) K2O using potassium chloride + 0.3 kg of potassium chloride dissolved in 100 L of water and applied in coverage) and; complete (a mixture of the three nutrients, 150, 300 and 100 g m(-3) N, P2O5 and K2O, respectively + 1.0 kg of ammonium sulfate + 0.3 kg of potassium chloride). The commercial substrate was composted milled pine bark plus vermiculite. Evaluations of the seedlings were performed at 90 days after sowing. The complete treatment (NPK) gave the highest values for biometric and best plant indices, which express the quality. When analyzing nutrients in isolation; potassium had the lowest effect. Based on these results it can be recommended to fertilize Schizolobium amazonicum seedlings in nurseries with 150, 300 and 100 g m(-3) of N, P2O5 and K2O, respectively, plus 1.0 kg of sulfate ammonium and 0.3 kg of potassium chloride applied in coverage.

Cyclooxygenase inhibitory properties of nor-neolignans from Styrax pohlii

Chemical investigation of the n-hexane and EtOAc fractions of the ethanolic extract from Styrax pohlii (Styracaceae) aerial parts resulted in the isolation of the benzofuran nor-neolignan derivatives egonol (1), homoegonol (2), homoegonol gentiobioside (3), homoegonol glucoside (4) and egonol gentiobioside (5). This is the first report of compounds 1-5 in S. pohlii. Compounds 1-5, the acetyl derivatives 1a and 2a, the ethanolic extract (EE), the n-hexane fraction (HF) and EtOAc fraction (EF) were tested for their inhibitory activities against COX-1 and COX-2. The results showed that EE, HF, EF and compounds 1-5 and 1 a-2 a shown weak to moderate inhibition of COX-1 and COX-2. Among the assayed nor-neolignans, 4 gave a COX-1 inhibition of 35.7% at 30 mu M. Compound 5 displayed a COX-2 inhibition of 19.7% at 30 mu M.

Mechanisms underlying the inhibitory effects of uroguanylin on NHE3 transport activity in renal proximal tubule

Lessa LM, Carraro-Lacroix LR, Crajoinas RO, Bezerra CN, Dariolli R, Girardi AC, Fonteles MC, Malnic G. Mechanisms underlying the inhibitory effects of uroguanylin on NHE3 transport activity in renal proximal tubule. Am J Physiol Renal Physiol 303: F1399-F1408, 2012. First published September 5, 2012; doi: 10.1152/ajprenal.00385.2011.-We previously demonstrated that uroguanylin (UGN) significantly inhibits Na+/H+ exchanger (NHE)3-mediated bicarbonate reabsorption. In the present study, we aimed to elucidate the molecular mechanisms underlying the action of UGN on NHE3 in rat renal proximal tubules and in a proximal tubule cell line (LLC-PK1). The in vivo studies were performed by the stationary microperfusion technique, in which we measured H+ secretion in rat renal proximal segments, through a H+-sensitive microelectrode. UGN (1 mu M) significantly inhibited the net of proximal bicarbonate reabsorption. The inhibitory effect of UGN was completely abolished by either the protein kinase G (PKG) inhibitor KT5823 or by the protein kinase A (PKA) inhibitor H-89. The effects of UGN in vitro were found to be similar to those obtained by microperfusion. Indeed, we observed that incubation of LLC-PK1 cells with UGN induced an increase in the intracellular levels of cAMP and cGMP, as well as activation of both PKA and PKG. Furthermore, we found that UGN can increase the levels of NHE3 phosphorylation at the PKA consensus sites 552 and 605 in LLC-PK1 cells. Finally, treatment of LLC-PK1 cells with UGN reduced the amount of NHE3 at the cell surface. Overall, our data suggest that the inhibitory effect of UGN on NHE3 transport activity in proximal tubule is mediated by activation of both cGMP/PKG and cAMP/PKA signaling pathways which in turn leads to NHE3 phosphorylation and reduced NHE3 surface expression. Moreover, this study sheds light on mechanisms by which guanylin peptides