Chemicals leaching from car tire rubber microplastics are the key drivers of toxicity towards early life stages of Atlantic cod (Gadus morhua)

Methods to accurately assess the toxicity of chemicals in plastics are needed. Importantly, there is a lack of studies successfully deconvoluting particle and leached chemical effects on organisms. A recent investigation by our group indicated elastomer chemicals are more potent than common thermoplastic chemicals in acute toxicity tests. The toxicity of car tire rubber (CTR) leachates and chemicals to marine species have recently been demonstrated in a range of studies, but other rubbers have been less studied. Here, we present a comprehensive study of chemicals leaching from rubber MPs, (rMPs) and their toxicity to cod (Gadus morhua) embryos. This study aimed to unravel the role of CTR (rMP model material) as a pollutant particle and as a carrier of chemicals, and thus distinguish between the effects derived from rMPs from those resulting from associated chemicals. To study the influence of experimental parameters on leachate composition, leachates of dishwashing (DG) and lab gloves (LG), balloons (BAL) and CTR were prepared and the effect of particle loading, size, water temperature, salinity and turbulence, as well as leaching time, on the chemical composition of the resulting leachate, were investigated. Early life stages of Atlantic cod (Gadus morhua) were used to study the effects of leachates, particles and a combination of the two. First, CTR, DG, BAL leachates were compared in terms of toxicity. Second, we compared 1) CTR particles pre-leached for 30 days to allow desorption of associated chemicals, 2) chemicals leached from the CTR over seven days, and 3) pristine CTR particles (pCTR) not subjected to pre-treatment, thus including all associated chemicals. Both experiments were conducted with 4 day exposures, followed by a recovery period of ~11 days. Endpoints included mortality, hatching, developmental alterations, and gene expression. DG and CTR exposure led to lower hatching success and increased mortality compared to BAL leachates. Embryonic development was impacted by exposure to CTR, resulting in smaller larvae at hatch. Leachate and pCTR exposures led to significantly lower hatching success and increased mortality compared to pre-leached particles. Larvae exposed to pCTR and leachates showed significant developmental alterations for the following morphometric parameters: eye diameter, eye to front distance, yolk/body ratio and myotome length.