AMP hydrolysis in soluble and microsomal rat cardiac cell fractions: kinetic characterization and molecular identification of 5 `-nucleotidase

The present study describes the enzymatic properties and molecular identification of 5`-nucleotidase in soluble and microsomal fractions from rat cardiac ventricles. Using AMP as a substrate, the results showed that the cation and the concentration required for maximal activity in the two fractions was magnesium at a final concentration of 1 mM. The pH optimum for both fractions was 9.5. The apparent K-m (Michaelis constant) values calculated from the Eadie-Hofstee plot were 59.7 +/- 10.4 mu M and 134.8 +/- 32.1 mu M, with V-max values of 6.7 +/- 0.4 and 143.8 +/- 23.8 nmol P-i/min/mg of protein (means +/- S.D., n = 4) from soluble and microsomal fractions respectively. Western blotting analysis of ecto-5`-nucleotidase revealed a 70 kDa protein in both fractions, with the major proportion present in the microsomal fraction. The presence of these enzymes in the heart probably has a physiological function in adenosine signalling. Furthermore, the presence of ecto-5`-nucleotidase in the microsomal fraction could have a role in the modulation of the excitation-contraction-coupling process through involvement of the Ca2+ influx into the sarcoplasmic reticulum. The measurement of maximal enzyme activities in the two fractions highlights the potential capacity of the different pathways of purine metabolism in the heart.

Relative contribution of pre- and post-synaptic effects to the neostigmine-induced recovery of neuromuscular transmission blocked by vecuronium

The relative contribution of the pre- and post-synaptic effects to the neostigmine-induced recovery of neuromuscular transmission blocked by vecuronium was studied. A conjunction of myographical and electrophysiological techniques was employed. The preparation was the sciatic nerve-extensor digitorum longus muscle of the rat, in vitro. The physiological variables recorded were nerve-evoked twitches (generated at 0.1 Hz), tetanic contractions (generated at 50 Hz) and end-plate potentials (epps), generated in trains of 50 Hz. The epps were analyzed in: amplitude of first epp in the train; mean amplitude of the 30th to the 59th epp in the train (epps-plateau); half-decay time of the epp; early tetanic rundown of epps in the train; plateau tetanic rundown of epps in the train; quantal content of the epps and quantal size. In myographical experiments, a concentration of vecuronium was found (0.8 mu m) that affected both twitches and tetanic contractions and a concentration of neostigmine was found (0.048 mu m) that completely restored the twitch affected by vecuronium. The cellular effects of vecuronium and neostigmine, studied alone or in association, in the above-mentioned concentrations, were scrutinized by means of electrophysiological techniques. These showed that vecuronium alone decreased the peak amplitude, the quantal content of epps and the quantal size and reinforced the tetanic rundown of epps. Neostigmine alone increased the peak amplitude, the quantal content and the half-decay time of the epps. When employed in the presence of vecuronium, neostigmine increased both the quantal content of the epps (via a presynaptic effect) and the half-decay time of the epps (via a postsynaptic effect). Seeing the pre- and the post-synaptic effects of neostigmine were of similar magnitude, they permit to conclude that both these effects contributed significantly to the restoration by neostigmine of the neuromuscular transmission blocked by vecuronium.

Alboserpin, a Factor Xa Inhibitor from the Mosquito Vector of Yellow Fever, Binds Heparin and Membrane Phospholipids and Exhibits Antithrombotic Activity

The molecular mechanism of factor Xa (FXa) inhibition by Alboserpin, the major salivary gland anticoagulant from the mosquito and yellow fever vector Aedes albopictus, has been characterized. cDNA of Alboserpin predicts a 45-kDa protein that belongs to the serpin family of protease inhibitors. Recombinant Alboserpin displays stoichiometric, competitive, reversible and tight binding to FXa (picomolar range). Binding is highly specific and is not detectable for FX, catalytic site-blocked FXa, thrombin, and 12 other enzymes. Alboserpin displays high affinity binding to heparin (K(D) similar to 20 nM), but no change in FXa inhibition was observed in the presence of the cofactor, implying that bridging mechanisms did not take place. Notably, Alboserpin was also found to interact with phosphatidylcholine and phosphatidylethanolamine but not with phosphatidylserine. Further, annexin V (in the absence of Ca(2+)) or heparin outcompetes Alboserpin for binding to phospholipid vesicles, suggesting a common binding site. Consistent with its activity, Alboserpin blocks prothrombinase activity and increases both prothrombin time and activated partial thromboplastin time in vitro or ex vivo. Furthermore, Alboserpin prevents thrombus formation provoked by ferric chloride injury of the carotid artery and increases bleeding in a dose-dependent manner. Alboserpin emerges as an atypical serpin that targets FXa and displays unique phospholipid specificity. It conceivably uses heparin and phosphatidylcholine/phosphatidylethanolamine as anchors to increase protein localization and effective concentration at sites of injury, cell activation, or inflammation.

Thermal Phase Behavior of DMPG Bilayers in Aqueous Dispersions as Revealed by (2)H- and (31)P-NMR

The synthetic lipid 1,2-dimyristoyl-sn-3-phosphoglycerol (DMPG), when dispersed in water/NaCl exhibits a complex phase behavior caused by its almost unlimited swelling in excess water. Using deuterium ((2)H)- and phosphorus ((31)P)-NMR we have studied the molecular properties of DMPG/water/NaCl dispersions as a function of lipid and NaCl concentration. We have measured the order profile of the hydrophobic part of the lipid bilayer with deuterated DMPG while the orientation of the phosphoglycerol headgroup was deduced from the (31)P NMR chemical shielding anisotropy. At temperatures > 30 degrees C we observe well-resolved (2)H- and (31)P NMR spectra not much different from other liquid crystalline bilayers. From the order profiles it is possible to deduce the average length of the flexible fatty acyl chain. Unusual spectra are obtained in the temperature interval of 20-25 degrees C, indicating one or several phase transitions. The most dramatic changes are seen at low lipid concentration and low ionic strength. Under these conditions and at 25 degrees C, the phosphoglycerol headgroup rotates into the hydrocarbon layer and the hydrocarbon chains show larger flexing motions than at higher temperatures. The orientation of the phosphoglycerol headgroup depends on the bilayer surface charge and correlates with the degree of dissociation of DMPG-Na(+). The larger the negative surface charge, the more the headgroup rotates toward the nonpolar region.

Thermoluminescence and optical absorption studies of natural grossular minerals

The optical absorption spectra of two samples of grossular have been measured at room temperature. An intense charge transfer band (UVCT) of iron extends to the visible and near infrared region. Some peaks associated to Fe3+ ions in tetrahedral and octahedral positions have been identified and their energy levels were computed. Mn2+ and Fe2+ ions are responsible with some bands and probably these ions occupy dodecahedral positions. No change in the intensity of optical absorption spectra were found after gamma dose, but only the 505 nm band decreases with irradiation. The OH spectra, consisting of OH overtones at 2750nm and asymmetric OH bands in the near infrared region were observed in the two samples. The heat treatment produces Fe2+ -> Fe3+ and Mn2+ -> Mn3+ by oxidation. This last was observed in sample II only. The thermally stimulated luminescence of both grossular samples has been investigated. Due to differences in iron and manganese concentration, not only a large difference has been observed in their optical absorption behavior, but also a striking difference in their thermoluminescent behavior. Actually, it is not clear whether other impurities such as Ti, Na and K that are present in quite different concentration in grossular I and II are also contributing to the thermoluminescenct properties of both samples. (C) 2008 Elsevier Ltd. All rights reserved.

Structural role of fluoride in the ion-conducting glass system B(2)O(3)-PbO-LiF studied by single- and double-resonance NMR

The local structure of an ion-conducting glass with nominal composition 50B(2)O(3)-10PbO-40LiF has been investigated by complementary (7)Li, (11)B, (19)F, and (207)Pb single- and double-resonance experiments. The results give insight into the structural role of the lithium fluoride additive in borate glasses: (1) LiF is seen to actively participate in the network transformation process contributing to the conversion of three- into four-coordinate boron units, as shown by (11)B single-resonance as well as by (11)B{(19)F} and (19)F{(11)B} double-resonance experiments. (2) (19)F signal quantification experiments suggest substantial fluoride loss, presumably caused by formation of volatile BF(3). A part of the fluoride remains in the dopant role, possibly in the form of small LiF-like cluster domains, which serve as a mobile ion supply. (3) The extent of lithium-fluorine and lead-fluorine interactions has been characterized by (7)Li{(19)F} and (207)Pb{(19)F} REDOR and SEDOR experiments. On the basis of these results, a quantitative structural description of this system has been developed.

A renewable copper electrode as an amperometric flow detector for nitrate determination in mineral water and soft drink samples

A novel approach was developed for nitrate analysis in a FIA configuration with amperometric detection (E=-0.48 V). Sensitive and reproducible current measurements were achieved by using a copper electrode activated with a controlled potential protocol. The response of the FIA amperometric method was linear over the range from 0.1 to 2.5 mmol L(-1) nitrate with a detection limit of 4.2 mu mol L(-1) (S/N = 3). The repeatability of measurements was determined as 4.7% (n=9) at the best conditions (flow rate: 3.0 mL min(-1), sample volume: 150 mu L and nitrate concentration: 0.5 mmol L(-1)) with a sampling rate of 60 samples h(-1). The method was employed for the determination of nitrate in mineral water and soft drink samples and the results were in agreement with those obtained by using a recommended procedure. Studies towards a selective monitoring of nitrite were also performed in samples containing nitrate by carrying out measurements at a less negative potential (-0.20 V). (C) 2009 Elsevier B.V. All rights reserved.

Conformational preferences for some 2-substituted N-methoxy-N-methylacetamides through spectroscopic and theoretical studies

The analysis of the IR carbonyl band of the 2-substituted N-methoxy-N-methylacetamides Y-CH(2)C(O)-N(OMe)Me (Y = F1, OMe 2, OPh 3, Cl 4), supported by B3LYP/6-311++G(3df, 3pd) calculations along with the NBO analysis for 1-4, indicated the existence of cis-gauche conformers i.e. (c) and (g) for 1 and 3, (c(1), c(2)) and (g(1), g(2)) for 2, and (c) and (g(1), g(2)) for 4. In the gas phase, the g conformer population prevails over the c one, for 1 and 3, the (c(1) + c(2)) population prevails over the (g(1) + g(2)) one for 2, and the (g(1) + g(2)) conformer population is more abundant than (c) one for 4. In n-hexane solution, the cis conformer is more abundant for 1-3. The occurrence of Fermi resonance in the nu(CO) region, in n-hexane, precludes the estimative of relative populations of the (c, g(1), g(2)) conformers for 4. The SCI-PCM calculations agree with the solvent effect on the nu(CO) band component relative intensities for 1-3. NBO analysis showed that the n(N) -> pi.(CO), orbital interaction is the main factor which stabilizes the gauche (g, g(1), g(2)) conformers for 1-4 into a larger extent relative to the cis (c, c(1), c(2)) ones. The n(y) -> pi(.)(Co,) sigma(C-Y) -> pi.(CO,) pi(CO) -> sigma(C-Y) and 7co orbital interactions still contribute, but into a minor extent for the stabilization of the gauche conformers relative to the cis ones. The existence of some pyramidalization at the nitrogen atom of the Weinreb amides 1-4 is responsible for the occurrence of Y(delta)-(4)center dot center dot center dot O(delta)-(9) and Y(delta)-(4)center dot center dot center dot N(delta)-(7) short contacts in the gauche (g, g(1), g(2)) conformers, which originates strong repulsive Coulombic interactions, acting in opposition to the large orbital stabilization of the gauche conformer with respect to the cis one. Therefore, a delicate balance of the Coulombic and orbital interactions seems to be responsible for the observed stabilization of the gauche (g, g(1), g(2)) and cis (c, c(1), c(2)) conformers, both in the gas phase and in the solution for 1-4. However, the cis conformer predominance, in non polar solvents, for the 2-substituted N-methoxy-N-methyl acetamides 1-3, bearing in a first raw (fluorine and oxygen) atoms, is in the opposite direction to the gauche conformer preference for the corresponding 2-substituted N,N-dialkyl-acetamides. (C) 2010 Elsevier B.V. All rights reserved.

Efeito anestésico do óleo de cravo em alevinos de lambari

Para avaliar o efeito anestésico do óleo de cravo em lambaris (Astyanax altiparanae), cinco grupos de 30 alevinos (0,6±0,1g) foram expostos às concentrações de 50, 75, 100, 125 e 150mg L-1, medindo-se, respectivamente: tempo de indução à anestesia profunda (caracterizada pela perda de equilíbrio, pela ausência de natação, pela redução dos movimentos operculares e pelas respostas apenas a estímulos táteis mais intensos), tempo de recuperação e taxa de mortalidade após a exposição. Em uma segunda etapa (10 peixes/tratamento), registraram-se, para cada concentração, os tempos de indução e de recuperação após anestesia cirúrgica (movimentos operculares lentos e irregulares e perda de reações a estímulos), anotando-se a mortalidade após seis minutos de exposição. Observou-se redução linear no tempo de indução à anestesia (0,01min mg-1 de anestésico acrescido) e aumento quadrático do tempo de recuperação com a elevação da concentração (resposta máxima estimada em 7,1 minutos). A anestesia profunda foi alcançada em tempo inferior a 1,5 minuto para todas as concentrações, com recuperação mais rápida e sem mortalidade para 50mg L-1. Para indução à anestesia cirúrgica, foram registrados menores tempos nas concentrações 75 e 100mg L-1; porém, com mortalidade de 80% e 100%, respectivamente. A concentração 50mg L-1 promoveu anestesia cirúrgica e recuperação em 3,29±0,71 e 4,97±0,63 minutos, respectivamente, sem mortalidade. Concluiu-se que o óleo de cravo possui efeito anestésico para alevinos de lambari, sendo 50mg L-1 a concentração eficiente e segura para indução à anestesia profunda em até 1,5 minuto e de anestesia cirúrgica em até 3,3 minutos de exposição.

Efeito residual da silicatagem no solo e na produtividade do capim-marandu sob pastejo

A escória siderúrgica é uma alternativa para a correção da acidez dos solos e é constituída de silicato de cálcio. Neste estudo, avaliaram-se os efeitos residuais da aplicação de silicato de cálcio nos atributos químicos do solo e da planta em Latossolo Vermelho distroférrico típico com capim-Marandu (Brachiaria brizantha cv. Marandu), sob intensidades de pastejo em lotação rotacionada. O delineamento experimental foi de blocos ao acaso, com intensidades de pastejo avaliadas pelas ofertas diárias de forragem de 50, 100, 150 e 200 kg t-1 de MS por PV nas parcelas experimentais, enquanto a aplicação superficial de silicato de cálcio combinado com calcário dolomítico, respectivamente, nas doses 0 + 0; 2 + 0; 4 + 0; 6 + 0; 2 + 4; 4 + 2 e 0 + 6 t ha- 1 nas subparcelas com quatro repetições, duas épocas (verão e inverno) e avaliação em três profundidades do solo (0-10, 10-20 e 20-40 cm). Os atributos químicos do solo pH em CaCl2, Ca, Mg, K, H + Al e V, avaliados 720 dias após a aplicação, apresentaram resultados favoráveis do poder residual do silicato de Ca e do calcário. A oferta de forragem 200 kg t-1 e o tratamento somente com calcário dolomítico (0 + 6 t ha-1) elevaram o valor de pH em CaCl2 e o V, principalmente na camada de 0-10 cm. Os teores de Si no solo foram influenciados pelas doses aplicadas de silicato de Ca, apesar de não terem causado alterações significativas nos teores foliares de Si. A composição químico-bromatológica foi afetada somente pelas ofertas e épocas. As ofertas, épocas e a interação época x oferta resultaram em efeitos na produção de matéria seca no pré-pastejo, com maiores produções para a oferta 200 kg t-1 e menores para a de 50 kg t-1 nas duas épocas. O resíduo (pós-pastejo) foi influenciado pelas ofertas e épocas. As ofertas 50 e 100 kg t-1 e o tratamento com 2 t ha-1 de silicato de Ca promoveram as maiores taxas de acúmulo de matéria seca.

Calagem e adubação boratada na produção de feijoeiro

Objetivou-se avaliar os efeitos da aplicação de calcário e boro nos atributos químicos de um Latossolo, estado nutricional e produtividade da cultura do feijoeiro. As doses de calcário empregadas foram crescentes, considerando-se a saturação por bases igual a 70%, correspondendo às doses: zero; 0,6; 1,2 (v = 70%); 1,8 e 2,4 t ha-1. O calcário utilizado apresentava poder relativo de neutralização total (PRNT), poder de neutralização (PN) e reatividade (RE) de 131; 137 e 95%, respectivamente. As doses de boro estudadas foram iguais a 0 (zero); 0,6; 1,2; 1,8; e 2,4 mg dm-3 de B, sendo a referência para definição das mesmas a de 1,2 mg dm-3 de B (ou 1,2 kg ha-1 de B). Utilizou-se como fonte o ácido bórico (H3BO3) p.a. (17% B). Na época do florescimento foi avaliado o estado nutricional das plantas e, aos 90 dias foram avaliadas a fertilidade do solo e a produção de grãos. No solo houve melhoria dos atributos pH, SB, V e na concentração de Ca e Mg, e do B com o emprego da adubação boratada. A interação da calagem com a adubação boratada promoveu maior acúmulo de Ca, Mg e B nas plantas. Os dados apresentados demonstram que o feijoeiro foi responsivo à calagem e à adubação boratada, tendo atingido bons índices de produção com a utilização de 1,8 kg ha-1 de B com doses crescentes de calcário.

Consumo, desempenho e parâmetros econômicos de novilhos Nelore e F1 Brangus x Nelore terminados em pastagens, suplementados com mistura mineral e sal nitrogenado com uréia ou amiréia

Avaliou-se o efeito da suplementação protéica (40% PB) com amiréia ou uréia sobre o consumo de suplemento, desempenho e características econômicas de novilhos terminados em pastagens. Foram utilizados 120 novilhos com 19 meses de idade e 358kg, sendo 60 Nelore e 60 F1 Brangus x Nelore, divididos em três tratamentos com 20 animais, alojados em piquetes de Brachiaria brizantha cv. Marandu de 10 hectares cada, totalizando 120 hectares, sendo dois piquetes por grupo genético e tratamento, pastejados alternadamente a cada pesagem (42 dias). Os tratamentos consistiram em mistura mineral com amiréia-150S (AM), mistura mineral com uréia+milho+enxofre (UR) e mistura mineral (MM). As médias de consumo de suplemento dos animais F1 foram de 206,1; 145,9 e 73,1g/dia, e as dos animais Nelore, 236,0; 205,1 e 94,3g/dia para os tratamentos AM, UR e MM, respectivamente. Para os novilhos Nelore, houve efeito (P<0,05) do suplemento sobre o peso de abate (PA), sendo a média do tratamento UR, 518,85kg, mais alta que a dos demais, 491,89 e 485,20kg, respectivamente, para AM e MM. Para os novilhos F1, foi significativo o efeito da suplementação protéica (P<0,05), com médias de 515,90 e 520,15kg, respectivamente, para os tratamentos UR e AM. A suplementação protéica proporcionou bom desempenho em animais F1 durante períodos de abundância de forragem. O uso de uréia apresentou melhor viabilidade econômica.

Modulation of peritoneal macrophage activity by the saturation state of the fatty acid moiety of phosphatidylcholine

To determine the effects of saturated and unsaturated fatty acids in phosphatidylcholine (PC) on macrophage activity, peritoneal lavage cells were cultured in the presence of phosphatidylcholine rich in saturated or unsaturated fatty acids (sat PC and unsat PC, respectively), both used at concentrations of 32 and 64 µM. The treatment of peritoneal macrophages with 64 µM unsat PC increased the production of hydrogen peroxide by 48.3% compared to control (148.3 ± 16.3 vs 100.0 ± 1.8%, N = 15), and both doses of unsat PC increased adhesion capacity by nearly 50%. Moreover, 64 µM unsat PC decreased neutral red uptake by lysosomes by 32.5% compared to the untreated group (67.5 ± 6.8 vs 100.0 ± 5.5%, N = 15), while both 32 and 64 µM unsat PC decreased the production of lipopolysaccharide-elicited nitric oxide by 30.4% (13.5 ± 2.6 vs 19.4 ± 2.5 µM) and 46.4% (10.4 ± 3.1 vs 19.4 ± 2.5 µM), respectively. Unsat PC did not affect anion production in non-stimulated cells or phagocytosis of unopsonized zymosan particles. A different result pattern was obtained for macrophages treated with sat PC. Phorbol 12-miristate 13-acetate-elicited superoxide production and neutral red uptake were decreased by nearly 25% by 32 and 64 µM sat PC, respectively. Sat PC did not affect nitric oxide or hydrogen peroxide production, adhesion capacity or zymosan phagocytosis. Thus, PC modifies macrophage activity, but this effect depends on cell activation state, fatty acid saturation and esterification to PC molecule and PC concentration. Taken together, these results indicate that the fatty acid moiety of PC modulates macrophage activity and, consequently, is likely to affect immune system regulation in vivo.

Screening process for activity determination of conductive oxide electrodes for organic oxidation

A modified method for the calculation of the normalized faradaic charge (q fN) is proposed. The method involves the simulation of an oxidation process, by cyclic voltammetry, by employing potentials in the oxygen evolution reaction region. The method is applicable to organic species whose oxidation is not manifested by a defined oxidation peak at conductive oxide electrodes. The variation of q fN for electrodes of nominal composition Ti/RuX Sn1-X O2 (x = 0.3, 0.2 and 0.1), Ti/Ir0.3Ti0.7O2 and Ti/Ru0.3Ti0.7O2 in the presence of various concentrations of formaldehyde was analyzed. It was observed that electrodes containing SnO2 are the most active for formaldehyde oxidation. Subsequently, in order to test the validity of the proposed model, galvanostatic electrolyses (40 mA cm-2) of two different formaldehyde concentrations (0.10 and 0.01 mol dm-3) were performed. The results are in agreement with the proposed model and indicate that this new method can be used to determine the relative activity of conductive oxide electrodes. In agreement with previous studies, it can be concluded that not only the nature of the electrode material, but also the organic species in solution and its concentration are important factors to be considered in the oxidation of organic compounds.

Qualidade da água para consumo humano e concentração de fluoreto

O artigo visa analisar a concentração de fluoreto na água para consumo humano, considerando o balanço entre benefícios e riscos à saúde, e produzir subsídios para atualização da legislação brasileira. Estudos de revisão sistemática, documentos oficiais e dados meteorológicos foram examinados. As temperaturas nas capitais brasileiras indicam que o fluoreto deveria variar de 0,6 a 0,9 mg/L para prevenir cárie dentária. Concentração de fluoreto natural de 1,5 mg/L é tolerável para consumo no Brasil se não houver tecnologia de custo-benefício aceitável para ajuste/remoção do seu excesso. A ingestão diária de água com fluoreto em concentração > 0,9 mg/L representa risco à dentição em menores de oito anos de idade e os consumidores deveriam ser expressamente informados desse risco. Considerando a expansão do programa nacional de fluoretação da água para regiões de clima tipicamente tropical, deve-se revisar a Portaria 635/75, relacionada ao fluoreto adicionado às águas de abastecimento público.