RESUMO
Analyses of natural waters frequently show elevated levels of total aluminum (Al) attributable to acid extraction of Al from the total suspended solids (TSS) minerals. Hence, there is a need for an analytical method that measures only bioavailable Al. Natural waters high in TSS were collected to study the chronic effects of Al on Ceriodaphnia dubia. In the collected waters TSS ranged from 30 to 411 mg/L; total Al concentrations ranged from 2.0 to 44.8 mg/L. The TSS in natural waters inhibited reproduction of C. dubia up to 40% in comparison to the same filtered waters. This inhibition did not correlate with the concentration of TSS or total Al; it was attributed to nutritional deficiency and was prevented by increasing the food supply. To demonstrate that toxicity can be measured in natural waters, samples with elevated TSS were spiked with soluble Al, and survival and reproduction were measured in chronic studies performed at pH 6.3 and 8.0. To properly characterize the Al concentrations in the toxicity studies, a method was needed that could discriminate bioavailable Al from mineral forms of Al. An extraction method at pH 4 for bioavailable Al was developed and evaluated using C. dubia chronic toxicity studies in the presence of TSS. It is concluded that the proposed method is better able to discriminate chronic toxicity effects attributable to bioavailable Al from mineralized nontoxic forms of Al compared with existing methods using total or total recoverable Al (i.e., extraction at pH ≤ 1.5). We propose that this new method be used when assessing the potential for Al in natural surface waters to cause toxicity. Environ Toxicol Chem 2019;38:1668-1681. © 2019 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals, Inc. on behalf of SETAC.
Assuntos
Alumínio/toxicidade , Cladocera/efeitos dos fármacos , Água Doce/química , Minerais/química , Poluentes Químicos da Água/toxicidade , Alumínio/metabolismo , Animais , Disponibilidade Biológica , Cladocera/metabolismo , Poluentes Químicos da Água/metabolismoRESUMO
The complex chemistry of iron (Fe) at circumneutral pH in oxygenated waters and the poor correlation between ecotoxicity results in laboratory and natural waters have led to regulatory approaches for iron based on field studies (US Environmental Protection Agency Water Quality Criteria and European Union Water Framework Directive proposal for Fe). The results of the present study account for the observed differences between laboratory and field observations for Fe toxicity to algae (Pseudokirchneriella subcapitata). Results from standard 72-h assays with Fe at pH 6.3 and pH 8 resulted in similar toxicity values measured as algal biomass, with 50% effect concentrations (EC50) of 3.28 mg/L and 4.95 mg/L total Fe(III), respectively. At the end of the 72-h exposure, however, dissolved Fe concentrations were lower than 30 µg/L for all test concentrations, making a direct toxic effect of dissolved iron on algae unlikely. Analysis of nutrient concentrations in the artificial test media detected phosphorus depletion in a dose-dependent manner that correlated well with algal toxicity. Subsequent experiments adding excess phosphorus after Fe precipitation eliminated the toxicity. These results strongly suggest that observed Fe(III) toxicity on algae in laboratory conditions is a secondary effect of phosphorous depletion. Environ Toxicol Chem 2017;36:952-958. © 2016 SETAC.
Assuntos
Clorófitas/efeitos dos fármacos , Compostos Férricos/toxicidade , Modelos Teóricos , Fósforo/análise , Poluentes Químicos da Água/toxicidade , Biomassa , Clorófitas/crescimento & desenvolvimento , Ecotoxicologia , Compostos Férricos/química , Água Doce/química , Concentração de Íons de Hidrogênio , Solubilidade , Testes de Toxicidade , Poluentes Químicos da Água/química , Qualidade da Água/normasRESUMO
The objective of the present study was to assess the predictive capacity of the acute Cu biotic ligand model (BLM) as applied to chronic Cu toxicity to Daphnia magna in freshwaters from Chile and synthetic laboratory-prepared waters. Samples from 20 freshwater bodies were taken, chemically characterized, and used in the acute Cu BLM to predict the 21-d chronic Cu toxicity for D. magna. The half-maximal effective concentration (EC50) values, determined using the Organisation for Economic Co-operation and Development (OECD) 21-d reproduction test (OECD Method 211), were compared with the BLM simulated EC50 values. The same EC50 comparison was performed with the results of 19 chronic tests in synthetic media, with a wide range of hardness and alkalinity and a fixed 2 mg/L dissolved organic carbon (DOC) concentration. The acute BLM was modified only by adjustment of the accumulation associated with 50% of an effect value (EA50). The modified BLM model was able to predict, within a factor of two, 95% of the 21-d EC50 and 89% of the 21-d half-maximal lethal concentrations (LC50) in natural waters, and 100% of the 21-d EC50 and 21-d LC50 in synthetic waters. The regulatory implications of using a slightly modified version of an acute BLM to predict chronic effects are discussed.
Assuntos
Cobre/toxicidade , Daphnia/efeitos dos fármacos , Água Doce/química , Modelos Biológicos , Animais , Chile , Concentração de Íons de Hidrogênio , Dose Letal Mediana , Ligantes , Poluentes Químicos da Água/toxicidadeRESUMO
Pollution of soil with mine wastes results in both Cu enrichment and soil acidification. This confounding effect may be very important in terms of phytotoxicity, because pH is a key parameter influencing Cu solubility in soil solution. Laboratory toxicity tests were used to assess the effect of acidification by acidic mine wastes on Cu solubility and on root elongation of barley (Hordeum vulgare L.). Three contrasting substrates (two soils and a commercial sand) and two acidic, Cu-rich mine wastes (oxidized tailings [OxT] and smelter dust [SmD]) were selected as experimental materials. Substrates were spiked with a fixed amount of either SmD or OxT, and the pH of experimental mixtures was then modified in the range of 4.0 to 6.0 and 7.0 using PIPES (piperazine-1,4-bis(2-ethanesulfonic acid)), MES (2-(N-morpholino)ethanesulfonic acid), and MOPS (3-(N-Morpholino)-propanesulfonic acid) buffers. Chemical (pore-water Cu and pH) and toxicological (root length of barley plants) parameters were determined for experimental mixtures. Addition of SmD and OxT to substrates resulted in acidification (0.11-1.16 pH units) and high levels of soluble Cu and Zn. Neutralization of experimental mixtures with MES (pH 6.0) and MOPS (pH 7.0) buffers resulted in a marked decrease in soluble Cu and Zn, but the intensity of the effect was substrate-dependent. Adjustment of soil pH above the range normally considered to be toxic to plants (pH in water extract, > 5.5) significantly reduced metal toxicity in barley, but phytotoxicity was not completely eliminated. The present results stress the importance of considering confounding effects on derivation of toxicity thresholds to plants when using laboratory phytotoxicity tests.
Assuntos
Cobre/análise , Cobre/toxicidade , Resíduos Industriais/análise , Mineração , Raízes de Plantas/efeitos dos fármacos , Poluentes do Solo/análise , Solo/análise , Fatores de Confusão Epidemiológicos , Hordeum , Concentração de Íons de Hidrogênio , Resíduos Industriais/efeitos adversos , Raízes de Plantas/crescimento & desenvolvimento , Poluentes do Solo/toxicidade , Solubilidade , Testes de ToxicidadeRESUMO
A better understanding of exposure to and effects of copper-rich pollutants in soils is required for accurate environmental risk assessment of copper. A greenhouse experiment was conducted to study copper bioavailability and bioaccumulation in agricultural soils spiked with different types of copper-rich mine solid wastes (copper ore, tailing sand, smelter dust, and smelter slag) and copper concentrate. A copper salt (copper sulfate, CuSO4) that frequently is used to assess soil copper bioavailability and phytotoxicity also was included for comparison. Results showed that smelter dust, tailing sand, and CuSO4 are more likely to be bioavailable and, thus, toxic to plants compared with smelter slag, concentrate, and ore at equivalent total copper concentrations. Differences may be explained by intrinsic differences in copper solubilization from the source materials, but also by their capability to decrease soil pH (confounding effect). The copper toxicity and bioaccumulation in plants also varied according to soil physicochemical characteristics (e.g., pH and total organic carbon) and the available levels of plant nutrients, such as nitrogen, phosphorus, and potassium. Chemistry/mineralogy of mine materials, soil/pore-water chemistry, and plant physiological status thus should be integrated for building adequate models to predict phytotoxicity and environmental risk of copper.
Assuntos
Agricultura , Cobre/análise , Solo/análise , Disponibilidade Biológica , Monitoramento Ambiental/métodos , Concentração de Íons de Hidrogênio , Resíduos Industriais , Metais Pesados/análise , Proteínas de Plantas/química , Medição de Risco , Poluentes do SoloRESUMO
The objective of this study was to assess the predictive capacity of the biotic ligand model (BLM) for acute copper toxicity to daphnids as applied to a number of freshwaters from Chile and to synthetic laboratory-prepared waters. Thirty-seven freshwater bodies were sampled, chemically characterized, and used to determine the copper concentration associated with the 50% of mortality (LC50) for Daphnia magna, Daphnia pulex, and Daphnia obtusa (native to Chile). The data were then used to run three versions of the acute copper BLM, and the predicted LC50s were compared to the observed ones. The same was done with synthetic assay media at various hardness and dissolved organic carbon (DOC) levels. The BLM versions differed in the affinity constants for some biotic ligand-ion pairs, stability constants for inorganic Cu complexes, and assumptions regarding Cu binding to DOC. All three versions showed a high degree of predictive performance, mostly within a twofold range of observed toxicity values. The D. obtusa data set was used to compare water quality criteria (WQC) derived from the observed toxicity values with those derived from either the BLM or the U.S. Environmental Protection Agency (U.S. EPA) procedure. For most low DOC waters, the three procedures generated similar WQCs. For the high-DOC waters, the EPA-derived criteria were significantly lower, that is, greatly overprotective. The results are also discussed in terms of the validation of the BLM for regulatory use.
Assuntos
Cobre/toxicidade , Daphnia/efeitos dos fármacos , Ligantes , Poluentes da Água/toxicidade , Água/química , Animais , Carbono/análise , Carbono/química , Chile , Cobre/metabolismo , Daphnia/metabolismo , Modelos Biológicos , Compostos Orgânicos/química , Valor Preditivo dos Testes , Testes de Toxicidade Aguda/métodos , Poluentes da Água/metabolismoRESUMO
Modification of Escherichia coli phosphofructokinase-2 (Pfk-2) with pyrene maleimide (PM) results in a rapid inactivation of the enzyme. The loss of enzyme activity correlates with the incorporation of 2 mol of PM/mol of subunit and the concomitant dissociation of the dimeric enzyme. The two modified residues were identified as Cys-238 and Cys-295. In the presence of the negative allosteric effector, MgATP, Cys-238 was the only modified cysteine residue. Kinetic characterization of the Cys-238-labelled Pfk-2 indicates that the enzyme is fully active, with the kinetic constants ( K(m), kcat) being almost identical to the ones obtained for the native enzyme. The modified enzyme is a monomer in the absence of ligands and, like the native enzyme, behaves as a tetramer in the presence of the nucleotide. However, in the presence of fructose-6-phosphate (fru-6-P) and ATP(-4), the enzyme behaves as a dimer, suggesting that the monomers undergo re-association in the presence of the substrates and that the active species is a dimer. Modification of Pfk-2 with eosin-5-maleimide (EM) results in the labelling of Cys-295. This modified enzyme is inactive and is not able to bind to the allosteric effector, remaining as a dimer in its presence. Nonetheless, Cys-295-labelled Pfk-2 is able to bind to the substrate fru-6-P in an hyperbolic fashion with a K(d) value that is 6-fold higher than the one determined for the native enzyme. These are the first residues to be implicated in the activity and/or structure of the Pfk-2.
Assuntos
Cisteína/química , Cisteína/fisiologia , Amarelo de Eosina-(YS)/análogos & derivados , Escherichia coli/enzimologia , Fosfofrutoquinase-2/química , Fosfofrutoquinase-2/metabolismo , Regulação Alostérica , Sequência de Aminoácidos , Dimerização , Amarelo de Eosina-(YS)/química , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Cinética , Maleimidas/química , Dados de Sequência Molecular , Estrutura Quaternária de Proteína , Solventes/química , Reagentes de Sulfidrila/químicaRESUMO
The unfolding and dissociation of the tetrameric enzyme fructose-1,6-bisphosphatase from pig kidney by guanidine hydrochloride have been investigated at equilibrium by monitoring enzyme activity, ANS binding, intrinsic (tyrosine) protein fluorescence, exposure of thiol groups, fluorescence of extrinsic probes (AEDANS, MIANS), and size-exclusion chromatography. The unfolding is a multistate process involving as the first intermediate a catalytically inactive tetramer. The evidence that indicates the existence of this intermediate is as follows: (1) the loss of enzymatic activity and the concomitant increase of ANS binding, at low concentrations of Gdn.HCl (midpoint at 0.75 M), are both protein concentration independent, and (2) the enzyme remains in a tetrameric state at 0.9 M Gdn.HCl as shown by size-exclusion chromatography. At slightly higher Gdn.HCl concentrations the inactive tetramer dissociates to a compact dimer which is prone to aggregate. Further evidence for dissociation of tetramers to dimers and of dimers to monomers comes from the concentration dependence of AEDANS-labeled enzyme anisotropy data. Above 2.3 M Gdn.HCl the change of AEDANS anisotropy is concentration independent, indicative of monomer unfolding, which also is detected by a red shift of MIANS-labeled enzyme emission. At Gdn.HCl concentrations higher than 3.0 M, the protein elutes from the size-exclusion column as a single peak, with a retention volume smaller than that of the native protein, corresponding to the completely unfolded monomer. In the presence of its cofactor Mg(2+), the denaturated enzyme could be successfully reconstituted into the active enzyme with a yield of approximately 70-90%. Refolding kinetic data indicate that rapid refolding and reassociation of the monomers into a nativelike tetramer and reactivation of the tetramer are sequential events, the latter involving slow and small conformational rearrangements in the refolded enzyme.
Assuntos
Frutose-Bifosfatase/química , Rim/enzimologia , Naftalenossulfonato de Anilina/química , Animais , Anisotropia , Cromatografia em Gel , Frutose-Bifosfatase/metabolismo , Guanidina/química , Cinética , Magnésio/química , Magnésio/farmacologia , Naftalenossulfonatos/química , Desnaturação Proteica , Dobramento de Proteína , Renaturação Proteica , Espectrometria de Fluorescência , Compostos de Sulfidrila/química , Reagentes de Sulfidrila/química , Suínos , Tirosina/químicaRESUMO
Binding of MgATP to the allosteric site of phosphofructokinase-2 promotes a dimer to tetramer conversion. In the presence of Fru-6-P the enzyme remains as a dimer. Limited proteolysis in the presence of MgATP completely protects the enzyme against inactivation and cleavage, while Fru-6-P provides a partial protection. A 28-kDa proteolytic fragment containing the N-terminus of the protein is inactive, but retains the ability to bind Fru-6-P and the allosteric effector MgATP. The fragment remains as a dimer but does not form a tetramer in the presence of MgATP. The results suggest major conformational changes of the enzyme upon ligand binding that confer a higher degree of compactness to the monomers in the dimer and in the tetramer, demonstrate the presence of the active and allosteric sites in this N-terminus fragment, and stress the importance of the C-terminus region of the protein for catalytic activity and ligand-induced oligomerization.