Nanoplastic Aggregation Driven by Environmental Components Reshapes Hormone Signaling and Energy Metabolism in Lactuca sativa

Nanoplastics seldom exist as isolated particles in the environment; they interact with dissolved organic matter (DOM) and metal ions, altering their aggregation and interfacial properties. Such aggregation is often overlooked in plant-toxicity assessments. Using polystyrene nanoplastics (PSNPs) as a model, we studied how Ca²⁺ and molecular-weight-fractionated DOM regulate aggregation and Lactuca sativa responses. Ca²⁺ induced rapid homoaggregation via charge screening. High-molecular-weight DOM stabilized PSNP dispersions, while low-molecular-weight DOM promoted compact heteroaggregation. These distinct aggregation states caused statistically significant differences in phytotoxic outcomes in lettuce, with large aggregates (Dh ≈ 1300 nm) inducing elevated oxidative stress (O₂⁻ and H₂O₂ increased by up to 71.0%) and growth inhibition, whereas dispersed or small aggregates (Dh ≈ 160 nm) elicited only mild adaptive stress responses. Integrated metabolomic and transcriptomic analyses revealed aggregation-dependent molecular responses, with large aggregates triggering a defense-prioritized regulatory shift. Thus, environmental component-mediated aggregation governs plant toxicity and should be considered in practical risk assessments.