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Though there are many toxicological studies on metal nanoparticles (NPs), it remains difficult to explain discrepancies observed between studies, largely due to the lack of positive controls and disconnection between physicochemical properties of nanomaterials with their toxicities at feasible exposures in a specified test system. In this study, we investigated effects of particle size and surface charge on in vitro mutagenic response and in vivo embryonic toxicity for newly synthesized silver nanoclusters (AgNCs) at human or environmental relevant exposure and compared the new findings with one of the most common nanoscale particles, titanium dioxide NPs (TiO 2 NPs as a positive control). We hypothesized that the interaction of the test system and physicochemical properties of nanomaterials are critical in determining their toxicities at concentrations relevant with human or environmental exposures. We assessed the mutagenicity of the AgNCs (around 2 nm) and two sizes of TiO 2 NPs (i.e., small: 5-15 nm, big: 30-50 nm) using a Salmonella reverse mutation assay (Ames test). The smallest size of AgNCs showed the highest mutagenic activity with the Salmonella strain TA100 in the absence and presence of the S9 mixture, because the AgNCs maintained the nano-size scale in the Ames test, compared with two other NPs. For TiO 2 NPs, the size effect was interfered by the agglomeration of TiO 2 NPs in media and the generation of oxidative stress from the NPs. The embryonic toxicity and the liver oxidative stress were evaluated using a chicken embryo model at three doses (0.03, 0.33, and 3.3 μg/g egg), with adverse effects on chicken embryonic development in both sizes of TiO 2 NPs. The non-monotonic response was determined for developmental toxicity for the tested NPs. Our data on AgNCs was different from previous findings on AgNPs. The chicken embryo results showed some size dependency of nanomaterials, but they were more well correlated with lipid peroxidation (malondialdehyde) in chicken fetal livers. A different level of agglomeration of TiO 2 NPs and AgNCs was observed in the assay media of Ames and chicken embryo tests. These results suggest that the test nanotoxicities are greatly impacted by the experimental conditions and the nanoparticle's size and surface charge.

Effects of Particle Size and Surface Charge on Mutagenicity and Chicken Embryonic Toxicity of New Silver Nanoclusters