A new anthracycline antitumor antibiotic, aclacinomycin A, was labeled with 3H uniformly or with 14C simultaneously at the anthracycline nucleus and L-rhodosamine. These labeled drugs were administered intravenously to normal dd mice, solid type Sarcoma 180 tumor-bearing ICR mice, normal or pregnant Wistar rats and normal rabbits, respectively. 14C-Aclacinomycin A given to rabbits (5 mg/kg) was rapidly cleared from the blood and transferred to tissues. But low level of radioactivity (equivalent to about 0.5 mcg/ml) was remained in the blood even 8 approximately 10 hours after administration. About 45% of the radioactivity were recovered from the urine and 20% from the feces by 72 hours after administration. Tissue levels of 3H-14C-aclacinomycin A given to normal and tumor-bearing mice were highest in the lungs and spleen. Higher distribution was observed also in the liver and kidneys 2 hours after administration. Bioassay revealed that the drug was present in the lungs and spleen in biologically active form and in the liver and kidneys in inactive form, respectively. In the tumor tissue the radioactivity was low but it persisted for 48 hours. Autoradiography with 14C-aclacinomycin A in rats demonstrated that radioactivity due to the drug distributed in the lungs, spleen, kidneys, thymus, intestine, lymph nodes, bone marrow, salivary gland, hypophysis and pineal body but it was rapidly cleared. About 0.2% of radioactivity given to a pregnant rat were transferred to a fetus when 14C-aclacinomycin A was administered intravenously on the 18 approximately 19th day of pregnancy.