Surface functionalization of polystyrene nanoparticles modulates nanoparticle-induced phytotoxicity in Chlorella vulgaris

: Polystyrene nanoparticles (NPs) are increasingly released into aquatic systems, but how surface formulations shape their interactions with primary producers remains insufficiently understood. We examined the effects of nominally non-functionalized polystyrene NPs (PS-NPs) and amino- (PS-NH 2 -NPs) or carboxyl-functionalized NPs (PS-COOH-NPs) on Chlorella vulgaris after short-term 72 h exposure to a comparative high concentration (40 mg L -1 ). All polystyrene NPs remained stable in the culture medium and were detected in EPS-associated and cell-associated fractions, with stronger relative retention of PS-NPs and PS-NH 2 -NPs than PS-COOH-NPs. Because Py-GC-MS data were qualitative and relative/semi-quantitative, cell-associated signals were interpreted as retention rather than direct evidence of intracellular uptake. All NPs induced ultrastructural alterations, including plasmolysis and thylakoid disorganization, without substantially affecting growth or cell viability. PS-NPs did not increase detectable ROS at the 72 h endpoint but caused lipid, protein and DNA damage. PS-NH 2 -NPs increased H 2 O 2 and lipid peroxidation and enhanced pigment accumulation, whereas PS-COOH-NPs showed the most pronounced total ROS response and protein oxidation. Antioxidant responses were characterized mainly by peroxidase activation and depletion of non-enzymatic antioxidants, reflecting redox imbalance. All NPs reduced oxygen evolution at growth light intensity, although unchanged F v /F m and increased PI abs did not indicate impairment of PSII functionality. Overall, commercial surface functionalization modulated NP retention and phytotoxic responses in C. vulgaris , highlighting the need to consider NP surface properties and exposure context in nanoplastic ecotoxicology.