Abstract: A factorial experiment was designed to evaluate the combined effects of temperature (20℃, 24℃ and 28℃) and salinity (10 and 30) on the osmotic pressure and antioxidant responses in the juvenile Japanese flounder (Paralichthys olivaceus). The results showed that the temperature and salinity caused no significant differences in the plasma cortisol of the juvenile Japanese flounder exposed in the experimental conditions for either 1 day (1 day after exposure, 1DAE) or 6 DAE. High temperature and low salinity (28℃, salinity10) significantly enhanced osmotic pressure level of the juvenile P. olivaceus in 1 DAE, but no significant difference in 6 DAE was observed. The Na+-K+-ATPase activity in the gill at 28℃ was significantly higher than that in 19℃ and 24℃ in 1 DAE, and the interactive effect between temperature and salinity on Na+-K+-ATPase activity was highly remarkable in 6 DAE. High temperature and low salinity increased SOD and CAT activity of juvenile P. olivaceus in 1 DAE, and the MDA content in the treatment (28℃, salinity 10) was significantly higher than the other treatments. The LPO content of juvenile P. olivaceus in salinity 10 was higher than that in the salinity 30 in 1 DAE at each temperature. There were no significant differences in liver SOD，CAT activity or MDA content in 1 DAE. Therefore，as euryhaline fish, P. olivaceus has very strong adaptability to environmental changes which allows them to tolerate conditions as high as 28℃ in temperature and as low as 10 in salinity. The changes of temperature and salinity may initially result in alterations of physiological activities and antioxidant responses, especially under high temperature and low salinity; and the physiological functions of flounders most likely recover to normal conditions after 6 days.