Does the exposure mode to ENPs influence their toxicity to aquatic species? A case study with TiO2 nanoparticles and Daphnia magna

Recent studies suggest that the ecotoxicity of engineered nanoparticles (ENPs) is dependent upon the treatment of ENPs in suspensions (e.g. sonication or use of solvents) and on the mode of exposure to test organisms. We conducted several bioassays with Daphnia magna in order to determine how adverse effects of TiO₂nanoparticles (n-TiO₂) are influenced by experimental set-up. Several treatments were applied, including three test media, several treatments of n-TiO₂suspensions (stirring, sonication) and different exposure modes (exposure duration and volume of test suspension). No adverse effects were observed when D. magna were exposed to 50 mL of suspension, regardless of TiO₂concentration (up to 250 mg/L) and exposure duration. Conversely, adverse effects were observed when D. magna were exposed to 2 mL of suspension for 96 h with a 50 % effect concentration EC₅₀values ranging from 32 mg/L to 82 mg/L. Test media had no significant influence on the outcome of all treatments. For a better mechanistic understanding of the experimental set-up at which adverse effects were observed, the particle size of n-TiO₂in the test media was characterized throughout the test duration. These measurements revealed a fast and strong agglomeration with a secondary particle size in the order of magnitude of micrometers. Our study describes how the effects of n-TiO₂on D .magna are influenced by the duration of exposure and volume of media, highlighting the need for standardization of experimental methods.