The chronic toxicity of josamycin was examined in Fischer 344 (F344) rats. Groups of 10 males and 10 females were given the test compound in the diet at concentrations of 0 (control), 0.02, 0.1, 0.5 or 2.5% for 52 weeks. Daily intake of josamycin was 0, 10, 50, 260 and 1310 mg/kg body weight in males and 0, 10, 60, 290 and 1460 mg/kg body weight in females, respectively. Body weight gain was significantly (P<0.05) reduced in the male 2.5% group but no noticeable changes were found in food intake. In hematological examination, the platelet count was significantly (P<0.01) lower in the male groups given 0.02% or more of josamycin and in the 2.5% female group as compared with the control group values in a dose-dependent manner. In serum biochemical examination, blood urea nitrogen was significantly (P<0.05 and P<0.01, respectively) higher in males given 0.5 and 2.5% and total bilirubin was significantly (P<0.05) higher in females receiving 2.5% as compared with those of the control group. No death occurred at any dose levels during the dosing period. At necropsy, with the exception of cecal enlargement in the groups given more than 0.1% josamysin and a significant (P<0.01) increase in the relative liver weight of females in the 2.5% group, no particular findings related to the administration were observed. Histopathologically, the incidence and severity of liver bile duct proliferation in female 2.5% group were significantly (P<0.01) greater than those of the control group. Other histological changes found in the treated and control groups were similar to the spontaneous lesions in this strain of rats in terms of the incidence and severity. Interestingly, the josamycin treatment reduced the development of altered liver cell foci in females in a dose-dependent manner. Thus, it is concluded that, under the present experimental conditions, josamycin induces bile duct proliferation in female F344 rats at a high dose of 1460 mg/kg body weight. Based on the decrease of platelet count found in males given 10 mg/kg body weight or more, the no-observed-adverse-effect level (NOAEL) was estimated to be less than 10 mg/kg body weight.