The influence of free radicals on iodothyronine 5'-monodeiodinase activity, the enzyme responsible for the deiodination of thyroxine to most active thyroid hormone 3,3',5-triiodothyronine (T3), was examined in rabbit's liver. Incubation of the liver homogenate with the xanthine oxidase based free radical generating system (FRGS) caused a reduction in 5'-monodeiodinase activity to the 53.9% of initial value taken as 100%, and on increase (52.9% over the control value) in the level of lipid peroxidation by-product malondialdehyde. The inhibitory effect of FRGS on 5'-monodeiodinase activity was blocked by free radical scavengers: catalase (91.2%), thiourea (88.8%), superoxide dismutase (85%) and by some antioxidants; Trolox (the water soluble alpha-tocopherol analog, 81.4%) and glutathione (77.7%). These results suggest that oxygen radicals, hydrogen peroxide and hydroxyl radicals were involved in the inhibition of the 5'-monodeiodinase activity. The same scavengers significantly decreased the malondialdehyde formation. In the presence of the FRGS the amount of total SH groups (the cofactor of the deiodination reaction) was decreased in the liver homogenate to 51% of the initial value, and a positive relationship between the total SH groups levels and the 5'-monodeiodinase activity in the presence of free radical scavengers was observed. It suggests, that active oxygen radicals generated by FRGS may inactivate 5'-monodeiodinase, at least in part, by reduction of thiol cofactors.