The impacts of plastic particles are being extensively studied, especially in agro-ecosystem. In the simulating experiment, elevated atmospheric CO2 concentration (e[CO2], 800 μmol mol−1) increased nanoplastics (NPs, red fluorescently labeled polystyrene, 95 nm) concentration in NPs-treated roots from 0.0082 mg g−1 to 0.0275 mg g−1. The H2O2 concentrations in NPs-treated leaves and roots were decreased by 28.41 % and 43.29 % responding to e[CO2], due to the intervention of antioxidant enzymes. In NPs-treated plants, e[CO2] increased Pn, Ci and carotenoid concentration by 10.97 %, 118.07 % and 40.83 %, while decreased gs and Tr by 36.36 % and 32.55 %. e[CO2] weakened NPs-induced modulation of photosynthetic light reaction via causing less electron production, but induced higher Ci promoting the Calvin-Benson cycle in NPs-treated plants. In the presence of NPs exposure, e[CO2] aggravated the reduction in glycolysis by decreasing the activities of cytInv, vacInv, cwInv, AGPase, UGPase, PGM, PGI, G6PDH, PFK and Ald, while suppressed the efficiency of tricarboxylic acid and biological oxidation due to the decreased activities of CS, ICDH, α-KGDH, NADH dehydrogenase, succinate dehydrogenase, cytochrome c oxidase and ATP synthase. The results showed the landscapes by which e[CO2] sculpt the eco-physiological response of barley plant to NPs, and provided key insights into the agricultural sustainability in the future under emerging pollution and environmental changes.