Toxicity to Embryo and Adult Zebrafish of Copper Complexes With Two Malonic Acids as Models for Dissolved Organic Matter

Abstract

Abstract-The toxicity to embryo and adult zebrafish (Danio rerio) of Cu complexes with two substituted malonic acids, benzyland n-hexadecyl-, chosen as models for low-molecular-weight natural dissolved organic matter, were investigated. Toxicity test solutions at pH 6.5 Ϯ 0.1 with the required Cu speciation were designed using the chemical speciation model MINTEQA2, and the speciation was confirmed by Cu ion-specific electrode. In the absence of malonic acids, concentrations of Cu 2ϩ up to 1.13 mol/ L increased the embryo hatching time from approx. 2 d in control solutions (no Cu or malonic acid) and solutions containing malonic acids without Cu to approx. 8 d. The Cu-benzylmalonic acid complex (n-octanol/water distribution coefficient [ ow ] ϭ 0.00) in the presence of inorganic Cu species did not delay hatching beyond that attributable to Cu 2ϩ . In contrast, 0.60 mol/L Cu-n-hexadecylmalonic complexes ( ow ϭ 6.89) delayed hatching by 5.5 d in excess of that attributable to 1.13 mol/L Cu 2ϩ , assuming that the hatching delays caused by the different Cu species were additive, possibly because of Cu entry into the embryo as the lipophilic Cu-n-hexadecylmalonic complex. None of the Cu-malonic acid complexes was acutely toxic to adult zebrafish at concentrations up to 1.4 mol/L, possibly because Cu was removed from the Cu-malonic acid complexes by stronger chelating groups at the gill surface. Substituted malonic acids with similar proton and Cu association constants can be readily prepared with a variety of simple substituents, radiolabeled if required. Our results show that these acids could be useful ligands for investigating intracellular transport and metabolism of metal-organic complexes.

Intra‐ and intertreatment variability in reference toxicant tests: Implications for whole effluent toxicity testing programs

“…Similarly, test conditions with respect to temperature, light regime and intensity, and dissolved oxygen conformed to the narrowly defined ranges specified in U.S. Environmental Protection Agency protocols. Small changes in environmental variables, such as dissolved organic matter and hardness, however, can have significant effects on observed toxicity with some reference toxicants (e.g., CuSO 4 used in the M. beryllina studies) [18]. Studies are required to determine important contributors to test result variability so that laboratories can take the required steps to reduce variability.…”

Formation of copper complexes in landfill leachate and their toxicity to zebrafish embryos

“…The aquatic toxicities of solutions containing inorganic Cu (no organic ligands), the leachate, the fractions and mixtures of Cu, and the leachate and the fractions, were tested with zebrafish embryos. Embryo hatching delay tests followed the Standardiseringskommissionen i Sverige Standardisation Group protocol [25] and the ring test [26] as described previously [17]. Zebrafish (one to two years old) for embryo production were maintained in glass aquaria (28 ± 1°C, 14:10 h light: dark cycle) in aerated, conditioned Auckland tap water and fed twice daily except just before spawning, when they were fed three times daily to maximize egg production.…”

“…The test solution compositions necessary to achieve the desired Cu 2+ concentrations were calculated using MINTEQA2 and the composition of the dilution water, the ligand concentrations, and the H and Cu constants determined from the titration data. The correction to I = 0 for the Cu constants was done by the procedure described previously for malonic acids [17]. The accuracy of the calculated Cu speciation in the test solutions and the stability of the speciation during the tests were checked by measuring the concentrations of Cu 2+ in the embryo test solutions after embryo exposure.…”

“…We have found that the Cu complex formed with hexade‐cylmalonic acid, an alkyl‐substituted dicarboxylic acid, is lipophilic and toxic to zebrafish embryos [17]. Organic acids are a major product of the early stages of anaerobic decomposition and up to 95% of the total organic compounds in leachate from young landfills, a common feature in urban environments, can be organic acids [18,19].…”

“…In view of the high concentrations of organic ligands that can occur in these leachates we hypothesized that either metabolite or lipophilic Cu complexes or both would be formed. We used embryo zebrafish ( Danio re‐rio ; family Cyprinidae) to test the toxicity of the complexes because of our previous observations that these embryos are sensitive to lipophilic Cu organic acid complexes [17].…”