The subacute toxicity test of cefoperazone (CPZ), a new cephalosporin antibiotic, was carried out in both sexes of Beagle dogs. CPZ was injected in Beagle dogs intramuscularly at dose levels of 500, 250 and 125 mg/kg/day for 3 months. The same amount of physiological saline was injected to control dogs intramuscularly for 3 months. The following results were obtained: 1) Sign of severe pain was shown immediately after injection in groups of CPZ at 500 and 250 mg/kg/day. In the same groups, the focal necrosis, hemorrhage, cell infiltration and fibrosis of muscles of injected site were noted microscopically. 2) In only one out of 6 dogs given CPZ at 500 mg/kg/day for 3 months, decrease of red blood cells and in values of hemoglobin and hematocrit an increase of reticulocytes in the peripheral blood were observed. In the same dog, splenomegaly and extramedullary hematopoiesis in the liver was found histopathologically, and in addition, the body weight and the food intake decreased during the administration period, associated with the development of anemia. 3) In dogs receiving up to 250 mg/kg/day, atrophy of the thymus was recognized at autopsy and slight decrease of cortical lymphocytes was seen histopathologically. 4) Based on these results, the maximum safety dose of CPZ was thought to be 125 mg/kg/day from the present experiment.