RESUMO
Bentazon (Basagran®) belongs to the chemical group of benzothiadiazinones. Thus, this study aimed to estimate the influence of herbicide bentazon (3 µg.L-1, 6 µg.L-1, 12 µg.L-1, 300 µg.L-1) in Danio rerio embryos development. The study tested environmental relevant concentrations of bentazon as well as the limit established for drinking water (300 µg.L-1) in Brazil. We performed behavioral and developmental analyzes during 96 h of exposure. The bentazon measurements after experimental period showed reduction ranging from 5.0 to 18.93% between exposed groups. Our results showed significant differences in the heart rate, which was significantly higher in groups exposed to all bentazon concentrations compared to control groups. The absence of alterations in the behavioral parameters showing that the herbicide bentazon at the concentrations tested had few adverse effects on the development and behavior of the Danio rerio embryos. Considering the toxic point of view, there is a chance that bentazon acts together with other environmental contaminants as an additive or synergistic way.
Assuntos
Herbicidas , Poluentes Químicos da Água , Animais , Herbicidas/toxicidade , Peixe-Zebra , Benzotiadiazinas/toxicidade , Desenvolvimento Embrionário , Poluentes Químicos da Água/toxicidade , Embrião não MamíferoRESUMO
In this study, we prepared Ti4O7 porous electrodes with continuous layered structures characterized by different layer-to-layer distance (from 2 to 10 µm) but the same total void fraction (88-90%), to modulate the electrodes' permeability and the volumetric electrochemical surface area (from 90 to 840 cm2 cm-3). These platforms were evaluated as anodes in the electro-oxidation (EO) of bentazon in a three-electrode cell under galvanostatic conditions, operated both in traditional batch (TB) or batch recycle flow-through (BRFT) modes. The performance was significantly enhanced when the liquid was recirculated through the lamellar structure of the electrodes. In BRFT mode, the electrode interlayer gap was found to be a key factor to control the bentazon and total organic carbon (TOC) conversions. For the best conditions evaluated (BRFT, 10 µm-interlayered Ti4O7 electrodes with a volumetric surface area of 90 cm2 cm-3), the effect of the applied current (1 or 3 mA) and liquid flow rate (10, 12 or 14 mL. min-1) was investigated. Specific energy consumption (SEC) values were estimated to reveal the performance of each of the EO treatments from an energetic point of view. The use of 10 µm-interlayered Ti4O7 electrodes at 1 mA in BRFT mode at a flow rate of 14 mL min-1 showed the best results, yielding 85% bentazon removal, 57% mineralization and SEC values of 0.006 kWh.gTOC-1 after 6 h of treatment. This contribution highlights the use of layered Ti4O7 electrodes as a promising strategy for intensifying EO processes, pointing to a trade-off between the accessibility to the internal electrode structure and the volumetric electrode surface area to enhance the contact between the target molecules and the hydroxyl radicals physisorbed on the electrode surface, while minimizing simultaneously the energy requirements.
Assuntos
Titânio , Poluentes Químicos da Água/análise , Benzotiadiazinas , Eletrodos , OxirreduçãoRESUMO
In this study, silica and functionalized silica materials (3-aminopropyl and 3-mercapto derivatives) were successfully used for the removal of the pesticides bentazone and imazapyc from aqueous solutions. Adsorbent materials were characterized by BET isotherms and FT-IR spectroscopy (confirming the functionalization), and their equilibrium adsorption capacity was evaluated at different ionic strengths. It is observed that the maximum adsorption capacities decrease in the order 3-aminopropyl-derivative > silica >3-mercaptopropyl derivative. An increase in ionic strength produces an enhancement in the removal of pesticides. All isotherms are Ib-type and follow the Langmuir model, suggesting a monolayer physical adsorption process.
Assuntos
Dióxido de Silício , Poluentes Químicos da Água , Adsorção , Benzotiadiazinas , Cinética , Espectroscopia de Infravermelho com Transformada de Fourier , ÁguaRESUMO
The main objective of this work is to demonstrate the viability of solar photoelectro-Fenton (SPEF) process to degrade pesticides in urban wastewater matrix, selecting the herbicide bentazon as a model molecule. In order to provide a correct assessment of the role of the different oxidants and catalysts involved, bentazon was comparatively treated by anodic oxidation with electrogenerated H2O2 (AO-H2O2), electro-Fenton (EF) and UVA-assisted EF (i.e., PEF) processes as well, either in sulfate or chloride media. Trials were made in a stirred tank reactor with an air-diffusion cathode and a boron-doped diamond (BDD), RuO2-based or Pt anode. In chlorinated matrices, the herbicide disappeared more rapidly using a RuO2-based anode because of the generated active chlorine. The best mineralization performance was always obtained using BDD due to its higher oxidation power, which allowed the complete destruction of refractory chloroderivatives. A concentration of 0.50â¯mM Fe2+ was found optimal to catalyze Fenton's reaction, largely enhancing the mineralization process under the action of OH. Among photo-assisted treatments, sunlight was proven superior to a UVA lamp to promote the photolysis of intermediates, owing to its greater UV irradiance and contribution of visible photons, although PEF also allowed achieving a large mineralization. In all cases, bentazon decay obeyed a pseudo-first-order kinetics. SPEF treatment in urban wastewater using BDD at only 16.6â¯mAâ¯cm-2 yielded 63.2% mineralization. A thorough, original reaction pathway for bentazon degradation is proposed, including seven non-chlorinated aromatics, sixteen chloroaromatics and two chloroaliphatics identified by GC-MS, most of them not previously reported in literature. Ion-exclusion HPLC allowed the detection of seven short-chain linear carboxylic acids.
Assuntos
Herbicidas , Poluentes Químicos da Água , Benzotiadiazinas , Técnicas Eletroquímicas , Eletrodos , Eletrólise , Peróxido de Hidrogênio , Oxirredução , Luz SolarRESUMO
A series of nineteen benzothiazin-4-ones from N-(3-aminopropyl) piperidine, 4-(2-aminoethyl)morpholine or 1-(2-aminoethyl)piperidine, aliphatic or aromatic aldehyde and thiosalicylic acid, were synthesized in good yields by multicomponent one-pot reactions. The solvent was toluene and this efficient procedure afforded the desired heterocycles in 5 h. Identification and characterization were achieved by NMR and GC-MS techniques. In vitro AChE activities of all compounds were evaluated in cerebral cortex and hippocampus of rats and in general, the results in cortex were more promising than hippocampus. The benzothiazinone 5Bd showed the best AChE inhibition activity IC50 8.48 µM (cortex) and IC50 39.80 µM (hippocampus). The cytotoxicity of seven compounds in MCR-5 human fibroblast cell by SRB test in 24 h were evaluated and 5Bd suggest preliminary safety, showing no cytotoxicity at 100 µM. Finally, these important findings could be a starting point for the development of new AChE inhibitors agents and will provide the basis for new studies.
Assuntos
Acetilcolinesterase/metabolismo , Benzotiadiazinas/farmacologia , Inibidores da Colinesterase/farmacologia , Animais , Benzotiadiazinas/síntese química , Benzotiadiazinas/química , Células Cultivadas , Inibidores da Colinesterase/síntese química , Inibidores da Colinesterase/química , Relação Dose-Resposta a Droga , Desenho de Fármacos , Fibroblastos/efeitos dos fármacos , Humanos , Masculino , Estrutura Molecular , Ratos , Ratos Wistar , Relação Estrutura-AtividadeRESUMO
In this paper we report the design, synthesis, antinociceptive and anti-inflammatory activities of a series of benzothiazine N-acylhydrazones 14ah, planned by structural modification of piroxicam (1), a non steroidal anti-inflammatory drug. Among the synthesized analogues, compounds 14f (LASSBio-1637) and 14g (LASSBio-1639) were identified as novel antinociceptive and anti-inflammatory prototypes, active by oral administration, acting by a mechanism of action that seems to be different from that of piroxicam, since they were inactive as an inhibitor of cyclooxygenase (COX-1 and COX-2) at concentrations of 10 mM.
Assuntos
Analgésicos , Anti-Inflamatórios , Inibidores de Ciclo-Oxigenase/farmacologia , Piroxicam , Administração Oral , Analgésicos/síntese química , Analgésicos/farmacologia , Animais , Anti-Inflamatórios/síntese química , Anti-Inflamatórios/farmacologia , Benzotiadiazinas/síntese química , Benzotiadiazinas/farmacologia , Ciclo-Oxigenase 1/química , Ciclo-Oxigenase 2/química , Proteínas de Membrana/química , Camundongos , Estrutura Molecular , Piroxicam/análogos & derivados , Piroxicam/síntese química , Piroxicam/química , Piroxicam/farmacologiaRESUMO
The knowledge of the host-guest complexes using cyclodextrins (CDs) has prompted an increase in the development of new formulations. The capacity of these organic host structures of including guest within their hydrophobic cavities, improves physicochemical properties of the guest. In the case of pesticides, several inclusion complexes with cyclodextrins have been reported. However, in order to explore rationally new pesticide formulations, it is essential to know the effect of cyclodextrins on the properties of guest molecules. In this study, the inclusion complexes of bentazon (Btz) with native ßCD and two derivatives, 2-hydroxypropyl-ß-cyclodextrin (HPCD) and sulfobutylether-ß-cyclodextrin (SBECD), were prepared by two methods: kneading and freeze-drying, and their characterization was investigated with different analytical techniques including Fourier transform infrared spectroscopy (FT-IR), differential thermal analysis (DTA), X-ray diffractometry (XRD) and differential pulse voltammetry (DPV). All these approaches indicate that Btz forms inclusion complexes with CDs in solution and in solid state, with a stoichiometry of 1:1, although some of them are obtained in mixtures with free Btz. The calculated association constant of the Btz/HPCD complex by DPV was 244±19 M(-1) being an intermediate value compared with those obtained with ßCD and SBECD. The use of CDs significantly increases Btz photostability, and depending on the CDs, decreases the surface tension. The results indicated that bentazon forms inclusion complexes with CDs showing improved physicochemical properties compared to free bentazon indicating that CDs may serve as excipient in herbicide formulations.
Assuntos
Benzotiadiazinas/farmacologia , Ciclodextrinas/química , Herbicidas/farmacologia , Química Farmacêutica/métodos , Físico-Química/métodos , Desenho de Fármacos , Eletroquímica/métodos , Cinética , Modelos Químicos , Oligossacarídeos/química , Praguicidas/química , Fotoquímica/métodos , Espectroscopia de Infravermelho com Transformada de Fourier/métodos , Propriedades de Superfície , Difração de Raios XRESUMO
The fluorimetric signal produced by bentazone retained in selected solid surfaces was investigated. Among the different tested supports, only a microporous nylon membrane produced the desired signal. The quantitative study was carried out by second-order calibration using parallel factor analysis, allowing the determination in a highly interfering medium. A detection limit of 0.4 ng mL(-1), a prediction relative error of 8%, and a sample frequency of ten samples per hour were obtained in spiked natural waters using green analytical chemistry principles.
Assuntos
Benzotiadiazinas/isolamento & purificação , Fluorometria/métodos , Herbicidas/isolamento & purificação , Membranas Artificiais , Nylons/química , Poluentes Químicos da Água/isolamento & purificação , Limite de Detecção , Água/análiseRESUMO
Bentazon removal by Ganoderma lucidum cultured in liquid and solid state conditions was compared in this work. In solid state cultures, the fungus produced both ligninolytic enzymes, namely laccase and Mn peroxidase. In liquid cultures, the main ligninolytic enzyme produced was laccase. In both types of cultures bentazon improved the production of laccase without significant alteration in the production of Mn peroxidase. In solid state cultures, where high levels of both laccase and Mn peroxidase activities were found, the fungus was more resistant to the action of the herbicide (50 mM in solid state cultures against 20 mM in liquid cultures) and more efficient in removing bentazon (90% removal against 55% in liquid cultures after 10 days of cultivation). Furthermore, the solid state culture filtrates were more efficient in the in vitro degradation of bentazon than the liquid culture filtrates. These observations suggest that both enzymes, laccase and Mn peroxidase, are involved in bentazon degradation. The results further suggest that solid state cultures of Ganoderma lucidum could be useful in strategies designed to reduce environmental contamination by bentazon.
Assuntos
Benzotiadiazinas/metabolismo , Meios de Cultura/química , Herbicidas/metabolismo , Reishi/metabolismo , Biotransformação , Proteínas Fúngicas/metabolismo , Lacase/metabolismo , Peroxidases/metabolismoRESUMO
Triasulfuron (TS) is a widely used sulfonylurea herbicide which inhibits the acetolactate synthase in broad-leaf weeds and in some wheat crop grasses (Triticum aestivum L.). Residues can be found in soil and superficial water with high toxicity to primary producers. In cereals, TS metabolism depends on cytochromes P450 (CYPs), the age of seedlings and the interaction with compounds. The genotoxicity of TS was demonstrated in the wing spot test of Drosophila melanogaster, an in vivo assay based on the loss of heterozygosity of the mwh and flr markers in the wing imaginal disk cells of larvae fed with chemical agents. Chronic treatments with analytical grade TS, commercial formulation TS (Amber) 75WG) (0.5mg/mL) and commercial formulation bentazon (Basagran) 480) (0.24mg/mL) were performed with three-day-old larvae of the standard (ST) and the high bioactivation (HB) crosses with regulated and high constitutive levels of CYPs, respectively. To demonstrate the effect of winter wheat metabolism on TS genotoxicity, T. aestivum L. seedlings were immersed for 4h in these herbicides, and aqueous extracts (AEs) of the roots were prepared to expose the larvae. TS and Amber 75WG produced similar genotoxic effects in both crosses. Wheat metabolism modulated the genotoxicity because the AEs yielded statistically significant lower spot frequencies in the HB cross than in the ST cross. Differences between the two crosses of the wing spot test in D. melanogaster must be related to CYPs levels. Basagran 480 was genotoxic only in the HB cross, and wheat metabolism did not modulate its genotoxicity.
Assuntos
Perda de Heterozigosidade/efeitos dos fármacos , Testes de Mutagenicidade , Mutagênicos/análise , Plântula , Compostos de Sulfonilureia/análise , Triticum , Asas de Animais , Animais , Benzotiadiazinas/análise , Benzotiadiazinas/metabolismo , Cruzamentos Genéticos , Sistema Enzimático do Citocromo P-450/metabolismo , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Marcadores Genéticos , Herbicidas/análise , Herbicidas/metabolismo , Larva/genética , Larva/metabolismo , Testes de Mutagenicidade/métodos , Mutagênicos/metabolismo , Proteínas de Plantas/metabolismo , Plântula/enzimologia , Compostos de Sulfonilureia/metabolismo , Fatores de Tempo , Triticum/enzimologiaRESUMO
Chlorophyll fluorescence techniques are used for the detection of toxic substances in samples of photosynthetic cells by measuring chlorophyll a fluorimetric parameters, which are a response of the PSII physiological status. This work was conducted to determine the effects of the herbicide bentazon (CASRN 25057-89-0) on growth and maximum quantum yield of photosystem II (Fv/Fm) in cells of the marine diatom Skeletonema costatum. Unialgal cultures were exposed to several bentazon concentrations and its effects on algal growth and Fv/Fm were determined. The traditional algal growth inhibition test (algal biomass measurements) and DCMU-induced chlorophyll a variable fluorescence measurements were determined. Our results showed that even low concentrations of bentazon rapidly lead to Fv/Fm decrease, while the effects on algal growth were detected after 24 h of exposure. The LOEC (2.81 mg L(-1)) and EC50 (13.0 mg L(-1)) determined through Fv/Fm experiments were lower than the LOEC (22.5 mg L(-1)) and EC50 (24.0 mg L(-1)) determined through algal growth inhibition experiments. This confirms that the Fv/Fm is a more sensitive parameter than algal growth for monitoring the effects of bentazon. The present results have demonstrated the applicability of Fv/Fm parameter to access the early toxicity of bentazon, as well as other PSII-inhibition compounds, before significant changes occurred in the original concentration and bioavailability of these toxicants during longer exposure times.
Assuntos
Benzotiadiazinas/toxicidade , Diatomáceas/crescimento & desenvolvimento , Diatomáceas/ultraestrutura , Herbicidas/toxicidade , Complexo de Proteína do Fotossistema II/metabolismo , Biomassa , Clorofila/metabolismo , Interpretação Estatística de Dados , Diatomáceas/efeitos dos fármacos , Diurona/toxicidade , Relação Dose-Resposta a Droga , FluorescênciaRESUMO
Testing of pesticide leaching models against comprehensive field-scale measurements is necessary to increase confidence in their predictive ability when used as regulatory tools. Version 5.1 of the MACRO model was tested against measurements of water flow and the behaviour of bromide, bentazone [3-isopropyl-1H-2,1,3-benzothiadiazin-4(3H)-one-2,2-dioxide] and imidacloprid [1-(6-chloro-3-pyridylmethyl)-N-nitroimidazolidin-2-ylideneamine] in a cracked clay soil. In keeping with EU (FOCUS) procedures, the model was first calibrated against the measured moisture profiles and bromide concentrations in soil and in drain water. Uncalibrated pesticide simulations based on laboratory measurements of sorption and degradation were then compared with field data on the leaching of bentazone and imidacloprid. Calibrated parameter values indicated that a high degree of physical non-equilibrium (i.e. strong macropore flow) was necessary to describe solute transport in this soil. Comparison of measured and simulated bentazone concentration profiles revealed that the bulk of the bentazone movement in this soil was underestimated by MACRO. Nevertheless, the model simulated the dynamics of the bentazone breakthrough in drain water rather well and, in particular, accurately simulated the timing and the concentration level of the early bentazone breakthrough in drain water. The imidacloprid concentration profiles and its persistence in soil were simulated well. Moreover, the timing of the early imidacloprid breakthrough in the drain water was simulated well, although the simulated concentrations were about 2-3 times larger than measured. Deep groundwater concentrations for all substances were underestimated by MACRO, although it simulated concentrations in the shallow groundwater reasonably well. It is concluded that, in the context of ecotoxicological risk assessments for surface water, MACRO can give reasonably good simulations of pesticide concentrations in water draining from cracking clay soils, but that prior calibration against hydrologic and tracer data is desirable to reduce uncertainty and improve accuracy.
Assuntos
Brometos/análise , Modelos Químicos , Praguicidas/análise , Solo/análise , Água/análise , Silicatos de Alumínio/análise , Benzotiadiazinas/análise , Calibragem , Argila , Temperatura Alta , Imidazóis/análise , Neonicotinoides , Nitrocompostos/análiseRESUMO
We have previously reported that topical application of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) to the rat neocortex prevents the effects of a subsequent application of N-methyl-d-aspartic acid (NMDA). Activation of NMDA receptors is involved in the pathogenesis of epileptic activity. Therefore, we examined if topically applied AMPA could affect changes in the somatosensory evoked potentials (SEPs) and electrocorticogram (ECoG) epileptic spikes caused by bicuculline. AMPA (50 microM) prevented the epileptiform activity to a level that was comparable to that caused by diazepam (3 mg/kg i.p.) or clomethiazole (100 mg/kg i.p.). Also, the epileptiform activity was suppressed by the AMPAR antagonist, CNQX, or the blocker of AMPAR desensitization, cyclothiazide. In the hippocampal slice, bicuculline-induced changes in the population spike potentials recorded from the CA1 cells were not affected by AMPA. We conclude that in the complex neuronal network of the rat neocortex, epileptiform activity can be suppressed in a variety of strategies that target the AMPA receptors: (1) blocking AMPA receptors, (2) promoting an apparent desensitization of AMPA receptors (possibly on the pyramidal neurons) or (3) reducing an apparent desensitization of AMPA receptors (possibly on the inhibitory GABA-ergic interneurons).
Assuntos
Epilepsia/fisiopatologia , Neocórtex/fisiopatologia , Receptores de AMPA/fisiologia , 6-Ciano-7-nitroquinoxalina-2,3-diona/farmacologia , Animais , Anticonvulsivantes/farmacologia , Anticonvulsivantes/uso terapêutico , Benzotiadiazinas/farmacologia , Bicuculina , Clormetiazol/farmacologia , Clormetiazol/uso terapêutico , Diazepam/farmacologia , Diazepam/uso terapêutico , Eletroencefalografia/métodos , Epilepsia/induzido quimicamente , Epilepsia/prevenção & controle , Potenciais Somatossensoriais Evocados/efeitos dos fármacos , Potenciais Somatossensoriais Evocados/fisiologia , Agonistas de Aminoácidos Excitatórios/farmacologia , Agonistas de Aminoácidos Excitatórios/uso terapêutico , Antagonistas de Aminoácidos Excitatórios/farmacologia , Hipocampo/citologia , Técnicas In Vitro , Masculino , Neocórtex/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/farmacologia , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/uso terapêuticoRESUMO
BACKGROUND: Liposuction, one of the most common operations performed by plastic surgeons, is not free of complications. One of the most common factors is patient hypothermia, a factor little studied but one capable of producing severe arrhythmias and cardiac arrest. A comparative clinical study was conducted to determine what effect using tumescent infiltration solutions at room temperature and at body temperature has on vital signs. METHODS: Two similar groups of 15 healthy female subjects were studied. In the first group (group A), subcutaneous solutions were infiltrated at room temperature (24 degrees C), and in the second group (group B), solutions were infiltrated at body temperature (37 degrees C). Vital signs (i.e., heart rate, respiratory rate, temperature, and blood pressure) were monitored every 15 minutes until the basal vital signs were attained. Variables such as age, body mass index, infiltrated and aspirated liquids, and surgery time were very similar for both groups. RESULTS: Although there were differences in heart rate, respiratory rate, and arterial pressure, they were not statistically significant. Nevertheless, the differences between groups A and B for body temperature (34.9 +/- 1.1 degrees C versus 35.7 +/- 1.3 degrees C, respectively) and for the time necessary to attain basal vital signs (120 +/- 8 minutes versus 69 +/- 4 minutes, respectively) were statistically significant (p < 0.05). CONCLUSIONS: Despite the existence of a significant change in the body temperature in healthy female subjects during manipulation of the temperature of the infiltration solution, this change had no important effect on the intraoperative hemodynamic values. Nevertheless, it could have a more significant effect on patients with greater surgical risk.
Assuntos
Temperatura Alta , Lipectomia/métodos , Adolescente , Adulto , Benzotiadiazinas , Óxidos S-Cíclicos , Feminino , Hemodinâmica , Humanos , Período IntraoperatórioRESUMO
Testing of pesticide leaching models is important to increase confidence in their use in pesticide registration procedures world-wide. The chromatographic PEARL model was tested against the results of a field leaching study on a cracking clay soil with a tracer (bromide), a mobile pesticide (bentazone) and a moderately sorbing, persistent pesticide (imidacloprid). Input parameters for water flow and solute transport were obtained from site-specific measurements and from literature. The model was tested using a stepwise approach in which each sub-model was sequentially and separately tested. Uncalibrated simulations for the water flow resulted in moisture profiles that were too wet. Calibration of the hydraulic relationships resulted in a good description of the moisture profiles. Calibration of the dispersion length was necessary to obtain a good description of bromide leaching. The calibrated dispersion length was 61 cm, which is very long and indicates a large non-uniformity of solute transport. The half-life of bentazone had to be calibrated to obtain a good description of its field persistence. The calibrated half-life was 2.5 times shorter than the half-life derived from the laboratory studies. Concentrations of bentazone in drain water and groundwater were described reasonably well by PEARL. Although measured and simulated persistence of imidacloprid in soil corresponded well, the bulk of the imidacloprid movement was overestimated by PEARL. However, imidacloprid concentrations in drain water were underestimated. In spite of the extensive calibration of water flow and tracer movement, the behaviour of the moderately sorbing pesticide imidacloprid could not be simulated. This indicates that the convection-dispersion equation cannot be used for accurate simulation of pesticide transport in cracking clay soils (even if extremely long dispersion length is used). Comparison of the model results from a poorly sorbed chemical (bentazone) and a moderately sorbed chemical (imidacloprid) were useful in defining the limitations of using a chromatographic flow model to simulate the effects of preferential flow.
Assuntos
Modelos Químicos , Praguicidas/química , Poluentes do Solo/análise , Poluentes Químicos da Água/análise , Benzotiadiazinas/química , Brometos/química , Simulação por Computador , Herbicidas/química , Imidazóis/química , Inseticidas/química , Neonicotinoides , Nitrocompostos , TemperaturaRESUMO
The mammalian kidney bumetanide-sensitive Na(+)-K(+)-2Cl(-) and thiazide-sensitive Na(+)-Cl(-) cotransporters are the major pathways for salt reabsorption in the thick ascending limb of Henle's loop and distal convoluted tubule, respectively. These cotransporters serve as receptors for the loop- and thiazide-type diuretics, and inactivating mutations of corresponding genes are associated with development of Bartter's syndrome type I and Gitleman's disease, respectively. Structural requirements for ion translocation and diuretic binding specificity are unknown. As an initial approach for analyzing structural determinants conferring ion or diuretic preferences in these cotransporters, we exploited functional differences and structural similarities between Na(+)-K(+)-2Cl(-) and Na(+)-Cl(-) cotransporters to design and study chimeric proteins in which the NH(2)-terminal and/or COOH-terminal domains were switched between each other. Thus six chimeric proteins were produced. Using the heterologous expression system of Xenopus laevis oocytes, we observed that four chimeras exhibited functional activity. Our results revealed that, in the Na(+)-K(+)-2Cl(-) cotransporter, ion translocation and diuretic binding specificity are determined by the central hydrophobic domain. Thus NH(2)-terminal and COOH-terminal domains do not play a role in defining these properties. A similar conclusion can be suggested for the Na(+)-Cl(-) cotransporter.
Assuntos
Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Alça do Néfron/metabolismo , Receptores de Droga/genética , Receptores de Droga/metabolismo , Simportadores de Cloreto de Sódio-Potássio/genética , Simportadores de Cloreto de Sódio-Potássio/metabolismo , Simportadores , Animais , Benzotiadiazinas , Bumetanida/farmacologia , Proteínas de Transporte/química , Polaridade Celular/fisiologia , Clonagem Molecular , Diuréticos/farmacologia , Feminino , Mutagênese , Oócitos , Estrutura Terciária de Proteína , Ratos , Receptores de Droga/química , Proteínas Recombinantes de Fusão/metabolismo , Inibidores de Simportadores de Cloreto de Sódio/farmacologia , Simportadores de Cloreto de Sódio , Simportadores de Cloreto de Sódio-Potássio/química , Membro 1 da Família 12 de Carreador de Soluto , Membro 3 da Família 12 de Carreador de Soluto , Xenopus laevisRESUMO
The thiazide-sensitive Na+:Cl- cotransporter is the major salt transport pathway in the distal convoluted tubule of the kidney, and a role of this cotransporter in blood pressure homeostasis has been defined by physiological studies on pressure natriuresis and by its involvement in monogenic diseases that feature arterial hypotension or hypertension. Data base analysis revealed that 135 single nucleotide polymorphisms along the human SLC12A3 gene that encodes the Na+:Cl- cotransporter have been reported. Eight are located within the coding region, and one results in a single amino acid change; the residue glycine at the position 264 is changed to alanine (G264A). This residue is located within the fourth transmembrane domain of the predicted structure. Because Gly-264 is a highly conserved residue, we studied the functional properties of this polymorphism by using in vitro mutagenesis and the heterologous expression system in Xenopus laevis oocytes. G264A resulted in a significant and reproducible reduction ( approximately 50%) in (22)Na+ uptake when compared with the wild type cotransporter. The affinity for extracellular Cl- and for thiazide diuretics was increased in G264A. Western blot analysis showed similar immunoreactive bands between the wild type and the G264A cotransporters, and confocal images of oocytes injected with enhanced green fluorescent protein-tagged wild type and G264A cotransporter showed no differences in the protein surface expression level. These observations suggest that the G264A polymorphism is associated with reduction in the substrate translocation rate of the cotransporter, due to a decrease in the intrinsic activity. Our study also reveals a role of the transmembrane segment 4 in defining the affinity for extracellular Cl- and thiazide diuretics.
Assuntos
Proteínas de Transporte/metabolismo , Rim/metabolismo , Receptores de Droga/metabolismo , Simportadores , Adulto , Sequência de Aminoácidos , Animais , Benzotiadiazinas , Sítios de Ligação/genética , Proteínas de Transporte/química , Proteínas de Transporte/genética , Cloretos/metabolismo , Diuréticos , Feminino , Humanos , Transporte de Íons , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Polimorfismo de Nucleotídeo Único , Ratos , Receptores de Droga/química , Receptores de Droga/genética , Inibidores de Simportadores de Cloreto de Sódio/metabolismo , Simportadores de Cloreto de Sódio , Membro 3 da Família 12 de Carreador de Soluto , Relação Estrutura-AtividadeRESUMO
The purpose of the present study was to determine the major functional, pharmacological, and regulatory properties of the flounder thiazide-sensitive Na-Cl cotransporter (flTSC) to make a direct comparison with our recent characterization of the rat TSC (rTSC; Monroy A, Plata C, Hebert SC, and Gamba G. Am J Physiol Renal Physiol 279: F161-F169, 2000). When expressed in Xenopus laevis oocytes, flTSC exhibits lower affinity for Na(+) than for Cl(-), with apparent Michaelis-Menten constant (K(m)) values of 58.2 +/- 7.1 and 22.1 +/- 4.2 mM, respectively. These K(m) values are significantly higher than those observed in rTSC. The Na(+) and Cl(-) affinities decreased when the concentration of the counterion was lowered, suggesting that the binding of one ion increases the affinity of the transporter for the other. The effect of several thiazides on flTSC function was biphasic. Low concentrations of thiazides (10(-9) to 10(-7) M) resulted in activation of the cotransporter, whereas higher concentrations (10(-6) to 10(-4) M) were inhibitory. In rTSC, this biphasic effect was observed only with chlorthalidone. The affinity for thiazides in flTSC was lower than in rTSC, but the affinity in flTSC was not affected by the Na(+) or the Cl(-) concentration in the uptake medium. In addition to thiazides, flTSC and rTSC were inhibited by Hg(2+), with an apparent higher affinity for rTSC. Finally, flTSC function was decreased by activation of protein kinase C with phorbol esters and by hypertonicity. In summary, we have found significant regulatory, kinetic, and pharmacological differences between flTSC and rTSC orthologues.
Assuntos
Benzotiadiazinas , Linguado/metabolismo , Inibidores de Simportadores de Cloreto de Sódio/farmacologia , Simportadores de Cloreto de Sódio-Potássio/metabolismo , Animais , Clortalidona/farmacologia , Diuréticos , Ativação Enzimática/efeitos dos fármacos , Feminino , Soluções Hipertônicas , Técnicas In Vitro , Cinética , Mercúrio/toxicidade , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Ligação Proteica , Biossíntese de Proteínas , Proteína Quinase C/metabolismo , Ratos , Simportadores de Cloreto de Sódio-Potássio/efeitos dos fármacos , Especificidade da Espécie , Relação Estrutura-Atividade , Xenopus laevisRESUMO
OBJECTIVE: To evaluate the effects of diet and medication, either isolated or associated, on serum levels of uric acid in patients with hyperuricemia. METHODS: We studied patients from the Hypertension Unit of the University of Goias who had hyperuricemia (men > or =8.5 mg/dL and women > or =7.5 mg/dL). We divided the patients into three groups: G1 (low purine diet), G2 (low purine diet + medication), and G3 (medication only). Patients received allopurinol, 150 mg/day titrated up to 300 mg/dL when necessary. Patients were evaluated with regards to their lifestyles (diet, smoking, physical, activity, alcohol consumption), uric acid, blood pressure, use of medication, body mass index, cholesterol, and triglyceride. Follow-up took place in weeks 0 (M1), 6 (M2), 12 (M3) during the intervention and in week 36 (M4) after the study was completed. RESULTS: Fifty-five patients participated in the study, 31 women, mean age 54.4+/-10.6 years, body mass index 28.6+/-3.9 kg/m2. A similar reduction (p<0.001) in uric acid levels occurred in the three intervention groups. In week 36 (M4), after 24 weeks without intervention, a tendency toward elevation of uricemia was noted in G2 and G3, and a continuous drop in uricemia was noted in G1. No significant modifications were observed in the other variables analyzed. CONCLUSION: Considering the cost x benefit relationship, a diet low in purine should be the 1st therapeutic option for controlling hyperuricemia in patients with similar characteristic to the ones presented in this study.
Assuntos
Alopurinol/administração & dosagem , Dieta , Hipertensão/sangue , Ácido Úrico/sangue , Benzotiadiazinas , Doenças Cardiovasculares/etiologia , Diuréticos , Ingestão de Energia , Etanol/efeitos adversos , Exercício Físico , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Risco , Fumar/efeitos adversos , Inibidores de Simportadores de Cloreto de Sódio/uso terapêutico , Resultado do TratamentoRESUMO
We report on a 29-year-old patient who received high doses of prednisone and cyclosporine for the treatment of Still disease. She consulted about dorsolumbar pain leading to physical disability. She presented multiple vertebral fractures, decreased lumbar bone mineral density in the rank of osteoporosis, high bone turnover, and associated hypercalciuria. Cyclosporine and corticoids induced severe changes in bone and mineral metabolism. All patients in treatment with these drugs should undergo radiology, bone densitometry and biochemical determinations of mineral metabolism at the beginning of therapy. Treatment with high doses of intravenous pamidronate (225 mg in 3 months), calcitonin (200/400 IU daily), tiazide (25 mg/daily), and kinesiotherapy mitigated the pain quickly and she recovered motility. We discuss this approach of treating osteoporosis with corticoids and immunosuppressors according to the present knowledge of bone biology.