Mixture toxicity study of two metal oxide nanoparticles and chlorpyrifos on Eisenia andrei earthworms.

Co-exposure soil studies of pollutants are necessary for an appropriate ecological risk assessment. Here, we examined the effects of two-component mixtures of metal oxide nanoparticles (ZnO NPs or goethite NPs) with the insecticide chlorpyrifos (CPF) under laboratory conditions in short-term artificial soil assays using Eisenia andrei earthworms. We characterized NPs and their mixtures by scanning electron microscopy, atomic force microscopy, dynamic light scattering and zeta potential, and evaluated effects on metal accumulation, oxidative stress enzymes, and neurotoxicity related biomarkers in single and combined toxicity assays. Exposure to ZnO NPs increased Zn levels compared to control in single and combined exposure (ZnO NPs + CPF) at 72 h and 7 days, respectively. In contrast, there was no indication of Fe increase in organisms exposed to goethite NPs. One of the most notable effects on oxidative stress biomarkers was produced by single exposure to goethite NPs, showing that the worms were more sensitive to goethite NPs than to ZnO NPs. Acetylcholinesterase and carboxylesterase activities indicated that ZnO NPs alone were not neurotoxic to earthworms, but similar degrees of inhibition were observed after single CPF and ZnO NPs + CPF exposure. Differences between single and combined exposure were found for catalase and superoxide dismutase (goethite NPs) and for glutathione S-transferase (ZnO NPs) activities, mostly at 72 h. These findings suggest a necessity to evaluate mixtures of NPs with co-existing contaminants in soil, and that the nature of metal oxide NPs and exposure time are relevant factors to be considered when assessing combined toxicity, as it may have an impact on ecotoxicological risk assessment.

Combined effects of goethite nanoparticles with metallic contaminants and an organophosphorus pesticide on Eisenia andrei.

The effects of mixtures of nanoparticles (NPs) and other chemicals have been poorly studied in terrestrial invertebrates. In this study, we investigated the effects of binary mixtures of goethite (α-FeOOH) NPs and metallic (Cd and Pb) or organic (chlorpyrifos, CPF) contaminants in Eisenia andrei earthworms. We used the filter paper contact test to evaluate (i) the uptake of NPs in organisms exposed to the mixtures of NPs+Metals and NPs+CPF and (ii) the potential effects of the mixture of NPs+CPF on the CPF-induced inhibition of the biomarker enzymes acetylcholinesterase (AChE) and carboxylesterases (CES). We used the artificial soil test to deepen the study on joint effects of NPs+CPF. All compounds were applied separately and in binary mixtures. In the single exposure treatment, Fe levels decreased significantly in organisms exposed to NPs on filter paper, suggesting systemic effects aimed at eliminating Fe incorporated through NPs. Conversely, earthworms exposed to binary mixtures showed Fe levels similar (NPs+Metals) to or higher (NPs+CPF) than controls. The earthworms single exposed to NPs presented no changes in AChE and CES activities. In the artificial soil test, the only treatment that showed AChE inhibition after 72 h was single CPF exposure, while no significant changes were observed in CES activity. However, after 7-day exposure in artificial soil or 72-h exposure on filter paper, the mixture of NPs+CPF induced a similar degree of AChE and CES inhibition as single CPF exposure. All these suggested that NPs did not produce neurotoxic effects, and that the inhibition of the enzymes' activities in all cases was due to the presence of the pesticide. On the other hand, the differences in the pattern of Fe accumulation in the earthworms indicate that the presence of other contaminants in the exposure media can modify the uptake and/or the excretion of Fe and evidence the interactions that may be found in binary mixtures of metal oxide NPs and other pre-existing contaminants in the soil ecosystem.