The objective of this study was to test the hypothesis that the widely observed age-associated increase in the amounts of macromolecular oxidative damage is due to an elevation in the availability of redox-active iron, that is believed to catalyze the scission of H₂O₂ to generate the highly reactive hydroxyl radical. Concentrations of bleomycin-chelatable iron and nonheme iron were measured in various tissues and different regions of the brain of mice fed on ad libitum (AL) or a calorically restricted (to 60% of AL) diet at different ages. The concentrations of these two pools of iron varied markedly as a function of tissue, age, and caloric intake. There was no consistent ratio between the amounts of nonheme and the bleomycin-chelatable iron pools across these conditions. Nonheme iron concentration increased with age in the liver, kidney, heart, striatum, hippocampus, midbrain and cerebellum of AL animals, whereas bleomycin-chelatable iron increased significantly with age only in the liver. Amounts of both nonheme and bleomycin-chelatable iron remained unaltered during aging in the cerebral cortex and hindbrain of AL mice. Caloric restriction had no effect on iron concentration in the brain or heart, but caused a marked increase in the concentration of both bleomycin-chelatable and nonheme iron in the liver and the kidney. The results do not support the hypothesis that accumulation of oxidative damage with age, or its attenuation by CR, are associated with corresponding variations in redox-active iron.