The interaction between prostaglandin F2α (PGF) and reactive oxygen species (ROS) is crucial for regulating the life span of the corpus luteum (CL). The local accumulation of ROS is mainly controlled by copper/zinc superoxide dismutase (SOD1). Thus, PGF may induce luteolysis by decreasing the expression of SOD and its bioactivity. Here, we examined whether SOD1 is involved in the luteolytic action of PGF in the bovine corpus luteum (CL). SOD activity gradually increased from the early to late luteal stage and then decreased to the lowest level at the regressed luteal stage. SOD1 protein expression and SOD activity increased at 2 h but decreased at 24 h after administration of a luteolytic dose of PGF. In addition, PGF and H2O2 increased SOD1 protein expression and SOD activity at 2 h but suppressed it at 24 h in cultured luteal cells. Furthermore, H2O2 increased PGF production by luteal cells in a dose- and time-dependent manner. PGF, in turn, induced ROS production. These results indicate that PGF via interaction with ROS regulates bovine luteal SOD1 in a biphasic manner with an initial increase at 2 h followed by a decrease at 24 h. The down regulation of SOD1 during structural luteolysis may enhance ROS production and luteal cell death to ensure the regression of the bovine CL.