<p>Perfluorooctanesulfonic acid (PFOS), a persistent and bioaccumulative substance, has emerged as a major environmental contaminant of global concern. PFOS, widely used in industry and consumer products, is common in aquatic ecosystems and may harm primary producers like phytoplankton, though its effects on their community structure and physiology remain unclear. Here, we investigated the impact of PFOS on freshwater physicochemical parameters, phytoplankton community dynamics, biochemical composition, and oxidative stress responses in a mesocosm experiment over a 28-day period. The water quality indicators generally were unaltered by PFOS exposure, but phosphate levels decreased initially in the control. Phytoplankton biomass (cell density) decreased at 10&#xa0;µg L⁻¹ and 10&#xa0;mg L⁻¹ PFOS exposures. Phytoplankton species richness, Shannon, and Simpson diversity indices declined at 10&#xa0;mg L⁻¹, with communities at this concentration becoming distinctly different from the control and lower PFOS treatments by day 28. This shift in community structure was primarily driven by taxa such as <i>Chlorogonium</i> sp. and <i>Microcystis aeruginosa</i>, which contributed strongly to the observed compositional dissimilarity. Proteins content in the phytoplankton increased under PFOS exposure, peaking on day 21. In contrast, carbohydrate and lipid contents were reduced under PFOS relative to the control. PFOS exposure elevated glutathione-S-transferase activity, malondialdehyde, and hydrogen peroxide levels, whereas peroxidase activity was reduced. The phytoplankton biomass and lipids were positively associated, while oxidative stress markers were closely linked with high PFOS concentrations. Overall, these results highlight that PFOS exposure suppressed phytoplankton biomass, altered their community composition, and induced oxidative stress and metabolic shift at higher concentrations, highlighting its potential ecological risk in freshwater systems.</p>