Hepatic phase I drug metabolism is diminished in old age. It has been suggested that hepatocyte hypoxia and impaired bioenergetics in old age may contribute to this aging change. Therefore, we sought to determine whether old age was associated with in vivo hypoxia in the aged rat liver. Immunohistochemical studies with the nitroimidazole hypoxia marker, pimonidazole, were carried out in livers from young and old rats. Preliminary studies were performed on four young (4-month-old) and six old (2-year-old) F344 rats to directly visualize the distribution and intensity of pimonidazole staining. There were no significant differences in the distribution or in the intensity of pimonidazole immunohistochemical staining between young and aged rat livers. In conclusion, no major changes in hepatocyte oxygenation were seen in the aged rat liver, and the ATP changes are unlikely to be secondary to hepatocyte hypoxia or impaired oxygen diffusion into the liver. It is thus more likely that age-related reduction in liver ATP is attributable to mitochondrial dysfunction.