Size-Dependent Toxicity of Polystyrene Nanoplastics to <i>Tetrahymena thermophila</i>: A Toxicokinetic–Toxicodynamic Assessment

Nanoplastic (NP) pollution poses a growing threat to aquatic ecosystems. Yet, accurate risk assessment based on their bioaccumulation and toxicity remains limited. Here, we synthesized polystyrene (PS) NPs with sizes of 30 nm (PS₃₀), 100 nm (PS₁₀₀), and 200 nm (PS₂₀₀), labeled with aggregation-induced emission fluorogens. This labeling approach enabled precise tracking of NP uptake and elimination in the protozoan <i>Tetrahymena thermophila</i>, thereby circumventing issues of low fluorescence intensity and label leakage associated with conventional fluorescence labeling methods. Significant uptake and elimination of the differently sized PS NPs were observed with multiple endocytosis and exocytosis pathways involved. Then we evaluated the effects of PS NPs on the growth of <i>T. thermophila</i> and explored the toxicity mechanisms. Transcriptomic analysis revealed that PS NPs disrupted energy metabolism, lipid metabolism, and cellular uptake pathways, with PS₃₀ even inducing genotoxicity. Using toxicokinetic-toxicodynamic modeling, we predicted median inhibitory concentrations (IC₅₀) and no-effect concentrations (NEC) of the differently sized PS NPs across exposure durations. Under chronic exposure conditions, the NECs were 0.52, 2.1, and 3.9 mg L^-1 for PS₃₀, PS₁₀₀, and PS₂₀₀, respectively, which have been detected in aquatic environments. Overall, our study provides a robust framework to evaluate the risks of NPs based on their toxicokinetic-toxicodynamic processes.