Natural Organic Ligands Research Articles

Desorption of Tributyltin, Dibutyltin and Zinc from Resuspended Sediment

Adsorption–desorption equilibrium and de- sorption kinetic studies were conducted in order to assess desorption of tributyltin (TBT), dibutyltin (DBT) and zinc from resuspended sediment. Equilibria were described in terms of linear adsorption. Desorption kinetics were represented by a two-site model which assumed that desorption consisted of rapid desorption and first-order kinetic desorption. TBT desorbed the most easily. Desorption kinetic parameters were related to adsorption mechanisms. Some experimental results did not correspond with the theoretical model. This was thought to be caused by natural organic ligands originally held in the sediment. Zinc deviated the most, because it was affected by the natural organic ligands more strongly than butylins. For example the partition coefficient of zinc varied by up to 1000-fold, and re-adsorption to sediment of the zinc that had been discharged to the water took place. Release of chemicals induced by remedial dredging was assessed. Desorbed TBT may pose a high risk. Among remediation alternatives, capping or leaving the sediment to stand prior to in situ degradation are preferred. © 1997 by John Wiley & Sons, Ltd.

Influence of Natural and Anthropogenic Ligands on Metal Transport during Infiltration of River Water to Groundwater

The behavior of dissolved metals, the anthropogenic ligands EDTA and NTA, and the natural ligands for Cu and Zn during infiltration of river water to the adjacent aquifer was investigated at the Glatt River field site (Switzerland). The speciation of EDTA was determined by a combination of experimental methods and of equilibrium calculations. The speciation of EDTA that behaves conservatively with respect to its total concentration changes on the infiltration path. The main EDTA species in the river water are Zn−EDTA, Ca−EDTA, and Fe(III)−EDTA and in some of the groundwaters Mn(II)−EDTA. The speciation of EDTA is strongly influenced by the presence of natural organic ligands for Cu and Zn that bind a large fraction of these metals, even in groundwater. Ni−EDTA is a less significant species than expected from equilibrium calculations. Fe(III)−EDTA is not in equilibrium with other metals due to its slow exchange kinetics. The fraction of EDTA that is present as Fe(III)−EDTA decreases from the river with 35%...

Effects of ligand speciation on determinations of the complexing abilities of strong ligands in natural waters

The presence of organic ligands (L) whose conditional stability constants (K′ML) are strong enough to allow them to form complexes with copper has been reported in oceanic waters. However, there is no general agreement on the distributional characteristics of such strong ligands in the water column. We present that these inconsistencies are derived from not only different analytical methods employed for their detection but also different oceanographic conditions. In particular, the nature and quantities of detectable natural ligands are affected by what kind of form the ligands are presentin situ in different marine environments, that is, chemical speciation of natural ligands (ligand speciation), which depends strongly on the variations in concentrations of coexisting trace metals. Using published data from observations in the laboratory and the field, we provide limits to the ranges of conditional stability constants of organic ligands for copper, zinc and cadmium that are detectable by the extensively used direct metal-titration method. For example, our model indicates that organic ligands for copper with log K′CuL(Cu′)>12.4 in surface water and with log K′CuL(Cu′)>9.9 in deep water might not have been detected because stronger ligands had formed complexes with ambient metalsin situ at a station in the North Pacific. The estimation suggests that there is a basinscale difference in speciation of natural organic ligands and, moreover, that this difference influences metal speciation. It is postulated that comparisons of the occurrence and levels of organic ligands might not be possible among spatially and temporally different observations without reconciliation of the effects of speciation of the ligands, even if an identical method is used in every case.

Natural Organics and Environmental Stability of Mercury: Electrochemical Considerations

The behavior of elemental mercury in aqueous systems containing organics has been examined thermodynamically in terms of the complexing effect of a natural organic ligand species on aqueous electrochemical equilibria in the system, and supplemented by tests on liquid mercury exposed to solutions containing natural organics. The implications of the study are serious in relation to the likely effect of organic‐containing aquatic systems on mercury solubility in the mercury contaminated regions of the world.

Voltammetry of labile heterogeneous complexes with low diffusion coefficients

The heterogeneity of natural organic ligands in the presence of a metal ion affects its concentration profile in the diffusion layer. In fact, owing to heterogeneity, the ratio of complexed metal to free metal increases in the diffusion layer with proximity to the electrode surface. The diffusing species are then decelerated because the diffusion coefficient of the macromolecular complex is lower than that of the metal ion, and so the total amount of reduced metal is decreased compared with that obtained in a homogeneous system with the same bulk speciation. Therefore, the voltammetric current, which is directly related to the concentration profile in the diffusion layer, is affected by heterogeneity. In the work reported in this paper the concentration profile in the diffusion layer was determined on the basis of the diffusion process and a heterogeneity model describing the concentration of the different complexing sites and their complexing affinity for the metal. The mean stability parameters for the complexation of natural organic ligands with a metal ion were determined from this profile, simulating the value that should be determined from voltammetric currents. The difference between the simulated value and the mean stability parameter related to bulk speciation was analysed.

Complexation of iron(III) by natural organic ligands in the Central North Pacific as determined by a new competitive ligand equilibration/adsorptive cathodic stripping voltammetric method

A highly sensitive voltammetric technique was developed to examine Fe speciation in seawater. The technique involves adding an Fe(III)-complexing ligand, salicylaldoxime, which competitively equilibrates with inorganic and organic Fe(III) species in ambient seawater. The Fe(III)-salicylaldoxime complex then is measured by adsorptive cathodic stripping voltammetry (ACSV). This new method revealed that 99.97% of the dissolved Fe(III) in central North Pacific surface waters is chelated by natural organic ligands. The total concentration of Fe-binding ligands is approximately 2 nM, a value greatly in excess of ambient dissolved iron concentrations. The titration data can be modeled as consisting of two classes of Fe-binding ligands, a strong ligand class (L 1 ) with an average surface-water concentration equal to 0.44 nM with a conditional stability constant K L1 Fe′ cond = 1.2 × 10 13 M −1 , and a weaker ligand class (L 2 ) with an average concentration equal to 1.5 nM with K L2 Fe′ cond = 3.0 × 10 11 M −1 . The low concentration of dissolved Fe present in surface waters (~ 0.2 nM), coupled with the excess of strong Fe-chelators, results in extremely low equilibrium concentrations of dissolved inorganic iron, [Fe′] ≈ 0.07 pM. In the deeper waters there is a 2 nM excess of Fe-binding ligands with a stability constant similar to that of the L 2 class of ligands observed in surface waters, resulting in dissolved Fe(III) existing primarily in the chelated form in deep waters as well. The stability constants of the natural ligands are comparable to the model ligands desferal, a siderophore, and the prosthetic heme group, protoporphyrin-IX. The high degree of organic complexation of iron makes it critically important to reevaluate our perceptions of the marine biogeochemistry of iron and the mechanisms by which biota can access this chelated Fe.

Complexation of Fe(III) by natural organic ligands in the Northwest Atlantic Ocean by a competitive ligand equilibration method and a kinetic approach

Total and labile Fe measurements, and Fe3+ titrations were carried out both at sea and in the laboratory with adsorptive cathodic stripping voltammetry (CSV) methods using 1-nitroso-2-naphthol (1N2N) as a complexing ligand to study Fe(III) speciation and the kinetic interaction of Fe3+ with naturally occurring organic ligands. On the continental slope and at the shelf/slope front of the Northwest Atlantic ocean, the total dissolved (< 0.4 μm) Fe was predominantly 1N2N nonlabile, with 60% nonlabile at the mouth of Delaware Bay. The exact chemical speciation of this nonlabile Fe is not known; although some of this Fe is likely in strong organic complexes with a KFeL1023.22 as determined by a competitive ligand equilibration/cathodic stripping voltammetry (CLE/CSV) method. To obtain kinetic information on Fe3+ uptake by natural ligands in excess of ambient Fe3+ concentrations in seawater, Fe3+ was added to UV irradiated seawater samples with and without the model ligands, EDTA and enterobactin. Measurable Fe in UV seawater with model ligands decreased with time much faster than that without model ligands. Similar faster decreases of measurable Fe with time were observed for field samples, suggesting the existence of Fe(III) complexing organic ligands. The reaction kinetics of the added Fe3+ with natural organic ligands and of the Fe(III)-organic complexes with 1N2N suggested that there were different organic ligands in different oceanographic regimes. Two classes of ligands are distinguished from these studies; one class which binds to Ca and Mg and another which does not. The latter assessment could not be unambiguously determined by a CLE/CSV method. Our kinetic experiments indicate that equilibrium is not achieved at the same rate in all natural samples studied and that different ligands occur in different water masses. The maximum second-order rate constant determined for Fe3+ uptake by natural ligands in our field samples is 106 M−1s−1. Using a CLE/CSV method at pH 6.9 with complete equilibrium established for the samples, a KFeLCL value of < 1012.04 ± 0.09 is calculated for the excess ligands in these samples. These data are in agreement with our detailed kinetic analysis using the steady state approximation based on natural ligand dissociation from FeL and recovery of that Fe by 1N2N at pH 8. KFeL (1020.6 M−1) estimated from kinetic data is within the range of estimates made by researchers using only the CLE/CSV approach.

Bioavailable iron species in seawater measured by macroalga (Laminaria japonica) uptake

The bioavailability of iron in seawater filtered through a 0.025-μm filter was investigated using 59Fe-labeled iron uptake by the macroalga Laminaria japonica (Areschoug: Phaeophyta) (collected in the northern Japan Sea 1993) as an assay. About 80% of the iron in the 0.025-μm filtered coastal seawater was soluble and/or small colloidal organically bound iron, associated with natural organic ligands forming complexes with ferric ion. After decomposition of the organic matter by ultraviolet (UV) irradiation, 55% of the iron addition [or 0.6 nM, nearly the concentration of Fe(OH)2+in equilibrium with amorphous hydrous ferric oxide in seawater at pH 8.0] in the 0.025-μm filtered coastal seawater was taken up by the macroalga. Since the iron concentrations in the 0.025-μm filtered coastal seawater are 0.1 to 2.0 nM and only 0.6 nM of the iron is likely available to biota over 1 to 2 d, we suggest that only small amounts of bioavailable iron exist in coastal seawater not affected by inflow from land and that a significant fraction of dissolved (<0.025 μ) iron occurs in forms, such as organic iron complexes, other than the simple hydroxo-complex species predicted by thermodynamic models.

Determination of complexation of iron(III) with natural organic complexing ligands in seawater using cathodic stripping voltammetry

A method is presented to determine the extent of iron complexation by natural organic ligands in seawater. Catalytic cathodic stripping voltammetry (CSV) is used to take advantage of ligand competition between the added ligand, 1-nitroso-2-napthol (NN), and natural ligands present in seawater. The conditional stability constant for the complexation of iron by NN was calibrated for salinities between 1 and 36 using ligand competition with EDTA. The values of log K′ Fe(NN) 3 (valid for pH 6.9 seawater) were found to vary linearly with log salinity according to log K′ Fe(NN) 3 = −1.04 ± 0.08 log(salinity) + 30.12 ± 0.09. The detection window, defined by the iron complexing ability of NN was altered by varying the concentration of NN. Preliminary measurements of iron complexing ligands in samples from coastal and open oceanic origin revealed the presence of natural complexing ligands at concentrations higher than that of total dissolved iron. The stability constants for the complexes were high, log K′ FeL falling within the range of 18.8–21.2, indicating that by far the greatest component (99%) of the dissolved iron occurs organically complexed in pH 6.9 seawater. Model calculations showed that it is possible that the organic fraction may be somewhat less at a pH value near 8.

Zinc speciation in lake waters and its determination by ligand exchange with EDTA and differential pulse anodic stripping voltammetry

A technique to determine free zinc ion concentration and zinc speciation in lake water was developed. The technique involves ligand exchange of added EDTA with natural organic ligands and measurement by differential pulse anodic stripping voltammetry (DPASV); pretreatment of samples and separation are not necessary. The validity of the ligand exchange approach was demonstrated in model waters containing glycine and CDTA (or EDTA), as representatives of weak and strong organic ligands. The ligand exchange and DPASV measurements were applied to water samples from Lake Greifen, a small eutrophic lake in Switzerland. The average fractions of dissolved zinc species in this lake were 7.8% free zinc ions, 33.5% weak organic complexes and 50.5% strong organic complexes. Total dissolved Zn in the lake water column ranged between 18 to 25 nM, free zinc ion concentrations 1.3–1.8 nM, with an average pZn of 8.8 and an average ratio of total Zn to [Zn2+] ≈ 13. The extent of complexation of Zn in the lake was much less than in some sea waters (ratios 60–100 in the North Pacific), probably due to higher ratios of total Zn concentration relative to organic ligands in the lake.

Conditional stability constants of metal complexes of organic ligands in sea water: past and present, and a simple coordination chemistry model

The conditional stability constants of complexes of organic ligands with trace metals (Cu and Zn) that have been determined by solution chemistry techniques since 1980 are summarized. The present common understanding on the chemical speciation of trace metals is that > 99% of copper dissolved in sea water is complexed with natural organic ligands; more than strongly binding organic ligand exists in sea water. A simple coordination model, including two non-specific discrete strong organic ligands such as EDTA, is introduced to understand the complex features of recent metal speciation. The model, consistent with the result of the ligand titration, permits the wide range of conditional stability constants determined by metal titrations and the present concentration levels of several trace metals in sea water to be explained. These findings suggest that organic metal complexes play a significant role in the geochemical cycle of reactive trace metals.

Effects of root exudates and humic substances on weathering kinetics

The effect of complex natural organic ligands on the weathering kinetics of aluminum oxide was investigated in laboratory experiments. A peat-derived humic substance and root exudates obtained from ectomycorrhizal (Picea abies — Hebeloma crustuliniforme) and non-mycorrhizal Norway Spruce trees; and γ-Al2O3 were used as a model system. The experimental weathering rates are in accordance with a surface-controlled dissolution mechanism. The effect of the humic material on dissolution rates appears to depend on the degree of protonation of the humic (macro)molecules: we observed dissolution-enhancement or -inhibition at pH 3 and 4, respectively. Ectomycorrhizal exudates proved to be effective weathering agents at pH 4, as opposed to humic material and non-mycorrhizal exudates. Our results suggest that (i) the role of humic materials in mineral weathering and podzolization is different from what is commonly thought, and (ii) mineral weathering rates in the rhizosphere may be higher than in the bulk soil.

The relation between salinity and copper complexing capacity of natural estuarine waters and the uptake of dissolved64Cu byMacoma balthica

The radiotracer 64Cu was used to assess the influence of natural organic ligands on the bioavailability of copper. Biological availability of the 64Cu-complexes was measured by accumulation in the bivalve Macoma balthica. The experiments were carried out in April as well as in February with water from the relatively clean Oosterschelde Sea arm and the relatively polluted Westerschelde estuary. Adsorption onto shells, as well as uptake in tissues was assessed at salinities of 10‰ and 30‰. Simultaneously with the exposure experiments, ligand characteristics of the natural waters were assessed. High ligand concentrations, as occurring in the Westerschelde around February, reduced 64Cu (320 nM) uptake by more than 50%, in spite of the much lower salinity in the Westerschelde water. At the low salinity, uptake was increased slightly in Westerschelde water, but considerably in Oosterschelde water. This implies that at low ambient ligand concentrations (during the whole year in Oosterschelde water and in the summer period also in Westerschelde water) the influence of salinity on 64Cu uptake is more pronounced.

Adsorption of lead and cadmium ions on calcite in the Krka estuary

The adsorption of Pb 2+ and of Cd 2+ ions from calcareous Krka river water of various salinities (3, 14, 20 and 38 psu) on calcite (CaCO 3, Merck p.a.) was investigated. Simultaneous adsorption of Pb 2+ and Cd 2+ ions was studied as well. The results suggest that the two ions are adsorbed at different calcite surface sites; Pb 2+ remained firmly bound to calcite at all salinities, whereas Cd 2+ was firmly bound at low salinities and released at high salinities. Dissolved natural organic ligands at the freshwater-seawater interface (FSI; at 14 psu) promoted and below the FSI (at 20 psu) decreased the adsorption of Cd 2+. The experiments were performed at metal concentrations of 8 × 10 −8 mol l −1 and at natural pH values around pH 8. Adsorbed amounts of trace metal ions were determined in filtered samples by differential pulse anodic stripping voltammetry (DPASV) with a three-electrode system, after 24 h of equilibration with calcite. Several adsorption models were tested, such as Freundlich's, Langmuir's and Schindler-Stumm's surface coordination model. Adsorption isotherms belong to S-1 class of empirical adsorption isotherms. None gave reasonable values of adsorption constants. The fractional partitioning of adsorbent to the solid phase when normalized to the quantity of adsorbent present ( K d) declined as solid concentration increased. It was found that the surface charge is not responsible for the observed effect. Instead, aggregate size increased, which effected a loss in surface area. This is a reasonable explanation for the observed S-shaped adsorption isotherms curves.

Complexation of cadmium by natural organic ligands in the central North Pacific

Complexation of cadmium by natural organic ligands in the central North Pacific

A new cathodic stripping voltammetric method for determining organic copper complexation in seawater

Different analytical methods may determine copper-complexing organic ligands of different strengths and concentrations in seawater, both because a variety of natural ligands may exist and because different methods may have different detection windows. Measurement of organic complexation by any single method at any single detection window may give only partial information on the true concentration of organically complexed and other forms of copper in seawater. Differential pulse cathodic stripping voltammetry (DPCSV) methods for studying dissolved trace metal complexation in seawater rely upon establishment of a competing equilibrium for the metal between the complexing ligands naturally present and a competing ligand added to the sample, followed by detection of the metal-added ligand complexes by DPCSV. The detection window can be varied by varying the concentration of the added competing ligand, or by adding a competing ligand which forms complexes of different strength with the metal of interest. We have developed a new DPCSV method for determination of organically complexed copper using tropolone as the added competing ligand because tropolone forms copper complexes that have different strength from those formed with catechol, the competing ligand added in a similar method. (The tropolone-copper complexes are weaker.) Thus, the tropolone-DPCSV method allows determination of ligands that form weaker copper complexes but that may exist at a higher concentration than those determined using the catechol-DPCSV method. In addition, tropolone is relatively stable and can be equilibrated with copper and natural ligands in a sample for extended periods (at least 12–15 h). We have demonstrated that this method can accurately determine total dissolved copper concentrations in standard seawater reference materials. Then, after calibrating the strength of the interaction between copper and tropolone in seawater of various salinities, we determined the concentration of natural copper-complexing organic ligands and the conditional stability constants of their copper complexes in surface water samples from the Indian Ocean and the North Sea. Over the range of copper concentrations employed in the titrations, we detected one copper-complexing organic ligand class in each sample. For the Indian Ocean sample, we determined a ligand concentration of 4.13 ± 0.05 nM, and copper complexes formed with this ligand had a conditional stability constant of 10 12.6 ± 0.5. For the North Sea sample, we determined an average ligand concentration of 16.2 ± 0.8 nM and an average conditional stability constant of 10 12.4 ± 0.8. Comparison of these results for the North Sea sample with those using the catechol-DPCSV method indicated that, as a result of the weaker copper complexes formed with tropolone relative to those with catechol, the tropolone-DPCSV technique determined copper-complexing organic ligands that exist at a higher concentration but form weaker copper complexes than those determined using the catechol-DPCSV technique. These results indicate that greater than 99.7% (in the Indian Ocean sample) and essentially 100% (in the North Sea sample) of the dissolved copper in these samples was organically complexed.

Sorption behavior of cobalt on illitic soil

Sorption behavior of cobalt has been studied on illitic soil samples from dilute nitrate solutions using a batch technique. 60Co was used as tracer. Distribution coefficients ( k d) were determined for the soil-cobalt solution system as function of contact time, pH, adsorbent, adsorbate and complementary cation concentrations. The effect of organic complexing ligands was also investigated. Desorption experiments were carried out to understand the sorption mechanism. The sorption was found to be rapid initially with slow rise later on. It was observed to increase markedly between pH 7–8 with maximum at pH 10. Percent adsorption increased with adsorbent concentration, while k d values first increased and then started decreasing. There was little effect observed with adsorbate concentrations in the concentration range studied. The data fit the Freundlich and Langmuir adsorption equations. Complementary cations, especially Ca and Mg, reduced the adsorption of cobalt. A 10 −3 mol dm −3 concentration of synthetic organic chelating agents strongly inhibited the sorption of cobalt, while the natural organic ligands had little effect. The desorption experiments showed that sorption was irreversible. The soil under investigation has got the capability of fixation of cobalt and would be useful for the disposal of radioactive waste solutions.

The use of chemiluminescence and ligand competition with EDTA to measure copper concentration and speciation in seawater

A batch chemiluminescence method for measurement of copper in seawater was developed from an existing flow injection technique. The method involved the reaction of copper in the sample with 1,10-phenanthroline, and subsequent release of photons from the oxidation of copper(II)-phenan-throline chelates by H 2O 2 at alkaline pH. The method was extremely sensitive with a detection limit in the range of 0.05–for 200 μl samples. Because of its sensitivity, it could be used to measure copper directly in seawater without a preconcentration step. To measure total copper, samples had to be acidified to pH 2 and stored for weeks to months (and in some cases also photo-oxidized by UV light) to release copper from kinetically inert species. Total copper concentrations measured by the chemiluminescence method in natural seawater agreed well with those obtained by more traditional methods. The chemiluminescence method was sensitive to copper speciation due to the slow dissociation of many bound copper species. This property was utilized to develop a ligand competition technique for measurement of free cupric ion concentrations. The method involved additions of EDTA and copper to seawater and was based on the lack of detectability of CuEDTA chelates by the chemiluminescence method. Free cupric ion concentrations measured by this method ranged from 10 −12.9 M for a relatively clean estuarine sample containing 5 nM total dissolved copper to 10 −10.9 M for a sample from a polluted harbor which contained 38 nM copper. Cupric ion concentrations measured by the new method were in good agreement with those measured in the same samples by a previously published method: EDTA competition combined with copper adsorption on to C 18 Sep—Pak cartridges. Copper was found to be heavily (>99%) complexed by natural organic ligands in both shelf and estuarine water samples and the concentrations of these ligands generally increased with dissolved copper concentration and decreasing sample salinity. These results agree with those of several previous studies.

Chemical environment for red tides due toChattonella antiqua

In order to assess the roles of Fe and Cu in outbreaks ofChattonella antiqua red tide, concentrations of these metals in the surface seawater were monitored around the Ie-shima Islands in the Seto Inland Sea during the summers of 1986–1988. Bioassay of the surface seawater with respect to Fe and Cu was also conducted using a cultured strain ofC. antiqua. Concentrations of Fe and Cu in the filtered seawater (FeF and CuF) were in the range of 3.9–10.0 and 9.3–11.2 nM, respectively. The bioassay with respect to Fe revealed that Fe at the surface layer was usually insufficient to support the maximum growth rate ofC. antiqua, except whenC. antiqua was dominant in the field. However, correlations between FeF and the growth rate of the control cultures (Fe, EDTA=not enriched; N, P, B12=enriched at optimum levels) were not apparent, probably because FeF did not reflect the concentration of available Fe. The bioassay with respect to Cu was coupled with the CuF values obtained. The results indicated that Cu at the surface layer was detoxified by complexation with natural organic ligand(s), and that pCu (=minus log of cupric ion activity) was 11.5–11.7, optimum for the growth ofC. antiqua, throughout the survey period. It is suggested that Fe, but not Cu, is a potentially important factor in regulating the natural populations ofC. antiqua in the Seto Inland Sea.

Effects of the detection window on the determination of organic copper speciation in estuarine waters

The complexation of copper by natural organic ligands in sea water was measured by cathodic stripping voltammetry with ligand (catechol and quinolin-8-ol) competition. Two methods to determine copper complexation in estuarine waters were compared, one based on a complete titration of the complexing capacity of the sample and the other on measurement of the labile and total dissolved metal concentrations only. Values for log α CuL (ihe α-coefficient for complexation of Cu 2+ by natural organic ligands) ranging from 3.2 to 7.7 and from 3.3 to 7.8 could be detected by varying long α CuAL (the α-coefficient for complexation of Cu 2+ by the added competing ligand) from 3.4 to 8.9 in samples from the Tamar estuary and from the Channel. The two methods gave comparable results and showed that the type of sites detected depends on the detection window of the technique. This effect is due to the sea and estuarine water samples containing a series of complexing ligands forming complexes of greatly varying strength, thus causing a rnage of complex stabilities to be measured as a function of the detection window of each technique. A comparison showed that lower values for α CuL are obtained by anodic stripping voltammetry as a result of that technique having a lower detection window. A detailed study of the Tamar estuary revealed a decrease in log α CuL from 10.8 to 8.3 with increasing salinity, demonstrating that major cations compete with copper for the complexing sites. The free Cu 2+ concentrations were very low throughout the estuary (16.2 < pCu 2+ < 18.2) even though the total measurements to establish potential toxic effects of copper in natural waters.