With more information becoming available on the presence of plastic waste in the environment, there is a growing societal concern for potential hazards that these plastics may pose to both human health and the environment. Microplastics, defined as pieces of plastic that are ≤5 mm, may cause adverse effects in small, aquatic invertebrates that filter feed in contaminated habitats via direct or indirect physical effects or via intrinsic toxicity. From the perspective of a company that produces plastics, there is a desire to investigate potential hazards of plastic waste, inform our product stewardship practices, and be a part of effective solutions. Previous investigations have indicated a need for improved methods for evaluating microplastics (e.g. see Connors KA, Dyer SD, Belanger SE. 2017. Advancing the quality of environmental microplastic research. Environmental Toxicology and Chemistry 36(7):1697-1703), and this information was considered in designing our own study.

Our ecotoxicity lab began planning a chronic toxicity study with the freshwater invertebrate, Daphnia magna, following the methods in the Daphnia magna Reproduction Test Guideline (OECD 211). In order to adapt this standard test guideline to microplastics research, several design aspects were modified, including the addition of a “particle control group” (in this case, fumed silica). This control group was added to the study design to assess whether any observed effects of the test material could be attributed to the particulate nature of the material or to intrinsic toxicity specific to the test material. Particles of an ethylene acrylic acid copolymer were evaluated in the daphnid chronic study at two concentration levels. These particles were approximately 100 nm in size and contained no additives that were expected to be toxic to the daphnids at the concentrations tested. Significant pre-work to characterize the agglomeration, homogeneity and stability of the particle dispersion was undertaken prior to the study. The test design was also adapted to include sufficient replication to assess not only D. magna growth, survival, and reproduction, but also transcriptome information in response to the test material exposure. Pre-work to extract high quality mRNA from the daphnids was performed prior to the study. The transcriptome analyses were conducted using RNA-seq, and this capability was performed in collaboration with researchers at U.S. Army Engineer Research and Development Center. The addition of transcriptomic endpoints allowed for the potential to investigate sub-organismal, molecular responses of exposed D. magna.

Results of the 21-day D. magna toxicity study with the ethylene acrylic acid copolymer particles indicated no significant effects on D. magna growth, survival, or reproduction in comparison with both particle and untreated control groups. However, significant transcriptomic alterations were induced at the highest treatment level of the test material (10¹² particles/L; 1.2 mg/L). These transcriptomic alterations occurred in multiple biological pathways, including central metabolism and energy reserves, oxidative stress, as well as ovulation and molting. In our opinion, this response indicated a non-specific, global, transcriptomic pattern. Upon reflection, the results were difficult to put into perspective of risk assessment at this time, because, to date, microplastic environmental monitoring approaches have largely not been equipped to detect particles in the nanosize range. However, our results indicate that ethylene acrylic acid copolymer particles in the upper nanosize range are not expected to adversely affect D. magna growth, survival, or reproductive outcomes at relatively high concentrations of up to 10¹² particles/L.

Partnering with the U.S. Army Engineer Research and Development Center as well as internal company experts in Formulation and Materials Sciences was critical to conducting this research. My learnings, as an ecotoxicologist interested in future microplastics research, are to continue to collaborate with scientists that have an expertise in polymer chemistry and particle behavior as well as collaborators that can expand upon the traditional endpoints in ecotoxicology. Studies with complex designs and data interpretation do “take a village” of scientists with different areas of expertise. For more details, I invite interested readers to take a look at our recently published article that summarizes this work: http://dx.doi.org/10.1002/etc.4745 (Coady et al. 2020. Assessment of Transcriptomic and Apical Responses of Daphnia magna Exposed to a Polyethylene Microplastic in a 21‐d Chronic Study. Environmental Toxicology & Chemistry 39:1578-1589.)

Published August 3, 2020.