To study the UV-B effect on photosynthesis in winter wheat at different day/night temperatures, biologically effective UV-B radiation at 4.2 (L_UVB) and 10.3 (H_UVB) kJ m^–2 d^–1 was provided on the seedlings at 25/20°C or 10/5°C. UV-B radiation inhibited net photosynthesis rate (P_n) by enhanced intensity and decreased temperature without change of intercellular CO₂ concentrations (C_i). Decreased maximal quantum yield of Photosystem II (F_v/F_m) and increased minimum fluorescence (F_o) were observed in H_UVB at both temperatures and L_UVB at 10/5°C. H_UVB increased total pool size (VAZ) of xanthophyll cycle pigments, but decreased the de-epoxidation state (DEPS) of these pigments at both temperatures, while L_UVB only decreased DEPS at 10/5°C. The activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) and the redox states of ascorbate and glutathione (AsA/DAsA and GSH/GSSG) were enhanced at 25/20°C, while there were increased SOD and CAT, unaltered APX activities and AsA/DHA, as well as decreased GR activity and GSH/GSSG in L_UVB and H_UVB at 10/5°C. UV-B radiation resulted in higher H₂O₂ and thiobarbituric acid reactive substance (TBARS) concentrations at 10/5°C than 25/20°C. It appears that low temperature alone did not influence photosynthesis but aggravated UV-B induced photoinhibition, which was associated with PSII photochemistry rather than stomatal limitation. Xanthophyll cycle pigments failed to provide photoprotection through thermal dissipation. The antioxidant system was up-regulated in L_UVB and H_UVB at 25/20°C, but was impaired at 10/5°C. Low temperature intensified UV-B induced photoinhibition and damage by weakening the antioxidant system.