Spiramycin (SPM) and acetylspiramycin (ASPM) have been known to be potent macrolide antibiotics in vivo, although spiramycins had rather mild antimicrobial activities in vitro. The physiological disposition of SPM and ASPM could participate in their in vivo activities. 14C-labeled SPM-I or 3H-labeled ASPM was administered intravenously in rats (20 mg/kg), and plasma levels, excretion and distribution have been studied. The plasma levels of SPM-I and ASPM were low both in radioactivity and bioactivity. After intravenous administration, 14C-SPM-I was excreted by 48 hours into urine, bile and faeces at the rates of 39.6, 31.4% and 37.1%, respectively. And 27.8% of the dose was recovered into urine by 48 hours after administration of 3H-ASPM. Higher radioactivities were detected in the spleen, kidney cortex, submaxillary gland, liver and lung by whole body autoradiography 1 hour after intravenous administration, and the levels tended to be retained until 24 hours. After intravenous administration at the dose rate of 50 mg/kg, the biological half-lives (T 1/2) of ASPM,SPM-I, josamycin and midecamycin were 151, 103, 71 minutes and 54 minutes, respectively, and the apparent volumes of distribution (V beta) were 9.2, 8.9, 3.6 L/kg and 7.7 L/kg, respectively. From these results, it was suggested that the highest activity observed for ASPM in experimental mice infections might be correlated with high affinity for the tissues and the long biological half-life of ASPM.