Towards Environmental Sustainability of Nanotechnology Through Improved Evaluations of Impact and Risk to Aquatic Species: Cross-Species Comparisons from the Laboratory and Environmentally Realistic Exposure Scenarios in the Field

Engineered nanomaterials (ENMs) are utilized in a wide variety of applications and products, including everything from toothpaste and personal care products to industries such as aerospace, defense, medicine, electronics, and agriculture. Depending on the application, release of ENMs to the environment may occur through both unintentional and deliberate routes. High surface-area-to-volume ratios make ENMs more volatile than bulk counterparts, and their small size allows translocation within biological systems that would not be possible with larger materials. These and other characteristics make ENM-biological system interactions unpredictable, creating uncertainty about their potential risks to environmental health. The purpose of this research is to address two current knowledge gaps in assessments of ENM toxicity, and to strive for more comprehensive ENM toxicity assessments. The first knowledge gap is the need to improve assessments of multi-species impacts. Given the copious amount of different ENMs that exist, read-across capabilities represent an important step towards the ability to predict nanotoxicity outcomes across species. However, many laboratory studies focus on impacts to a single species, and efforts to synthesize meaningful conclusions through the aggregation of results from individual studies is complicated by experimental designs that vary across studies. This work includes two different multi-species assessment of impact of lithium cobalt oxide nanosheets across three different freshwater model organisms that represent varied levels of sensitivities, different aquatic niches, and different taxa: Chironomus riparius (chironomids), Daphnia magna (daphnids) and Danio rerio (zebrafish). These assessments were carried out under uniform conditions, using the same media, coordinated developmental stages, and the same nanomaterial source. Additionally, comparing impacts in these species after two different exposure durations: 6- vs. 48-hours. It was found that species-specific responses (species-specific mechanisms or varying levels of differential gene expression response) may help to explain sensitivity levels across species, including adrenergic and cardiac-related pathways for zebrafish and daphnids, and the relatively high impact on iron-sulfur proteins in daphnids. Responses shared across species included some that have not commonly been described in impacts across a wide range of species, including impacts to neurological, visual, and reproductive systems. These findings expand mechanistic knowledge of ENM toxicity and provide a wealth of information that may be used to improve or develop new adverse outcome pathways. The influence of exposure duration was strongest for most-sensitive daphnids, which exhibited the strongest biochemical responses after the 6-hour exposure. Both duration exposure and concentration played significant roles zebrafish, which exhibited duration-dependent responses that showed opposite trends depending on exposure concentration. These results present opportunities for further study of the variation in biochemical responses across species and different time points, allowing the exploration of potentially novel mechanisms of ENM impact, as well as those that enhance understanding of variation in ENM toxicity across species. The second knowledge gap is the need to better understand post-application fate of agriculturally related ENMs, considering that direct release to the environment would be expected. The current work investigates how environmental transport and of potential nanoagrofertilizer copper phosphate ENMs (Cu3(PO4)2 ENMs) will compare with contemporary agrochemicals in terms of their release of copper to simulated agricultural runoff. It will also examine dissolution and transformation of Cu3(PO4)2 ENMs to better characterize exposure scenarios for aquatic species that would be exposed through contaminated runoff. Cu3(PO4)2 ENMs contributed less copper to the aqueous fraction of runoff than did Kocide 3000. Regardless of chemical, contributions of copper to particulate runoff fraction were substantially higher, leveling implications for benthic-dwelling species. Transformations of Cu3(PO4)2 ENMs included increased carbon and decreased copper detected on the particle surface, while other characteristics remained unchanged, including speciation of copper and phosphate; crystalline structure; and morphology. This indicates that environmentally relevant exposures in aquatic systems may involve particles with some of their original, as-synthesized characteristics