in vitro Chemical and physical toxicity of polystyrene microplastics in human-derived cells

<title>Abstract</title> Background With the increase of plastics production, a variety of toxicological studies regarding the microplastics have been reported since the microplastics could be ingested by the human body and cause serious diseases. However, the previous studies have been mainly focused on the toxicity of sphere type microbeads, which may be different from that of the randomly-shaped microplastics in real environment. Here, we have conducted the in vitro toxicology for randomly-shaped microplastics following the hypothesis that (1) physical cytotoxicity is affected from nano-/micro-size roughness in polystyrene (PS) microplastics and (2) chemical toxicity is caused by chemical reagents from microplastics.Results To prepare random shape of PS microplastics, we produced microfragments by ball mill grinding, then analyzed them via various toxicity tests in chemical and physical aspects with various kinds of human-derived cells. Ground PS microplastics were sorted in 3 ranges: 5-25 μm, 25-75 μm and 75-200 μm, and treated up to 1 mg/mL to cells based on weekly human intake of microplastics. We have confirmed that the PS microfragments induced 20 times increased acute inflammation for immune cells, production of reactive oxygen species and cell-death for fibroblasts and cancer cells by release of chemical reagents from microplastics. In addition, when the PS microfragments were in direct contact with the fibroblast and red blood cells, they lead to the lactose dehydrogenase release caused by a cell membrane damage and hemolysis by physical stress of microfragments. This phenomenon was amplified as microfragments concentration and roughness increases, we quantitatively analyzed roughness differences between microplastics, demonstrating that there are strong relationship physical damage of cells and roughness of microplastics.Conclusion We found that the PS microfragments have chemical toxicity. Furthermore, the physical toxicity by PS resulted in cellular membrane damage and correlated with statistically quantified-shape roughness. Therefore, we newly suggested the additional physical toxicity of random shape of microplastics. This provides the evidence of environmental and biological risks on random shape of microplastics.