The mitochondrial theory of aging was tested with optimised preparation techniques. Mitochondria were isolated from approximately 90 mg quadriceps muscle from healthy humans at age 70+ and 20+. The content of mitochondrial protein was approximately 10 mg g(-1) muscle and the yields were approximately 40%. The mitochondrial integrity was high as judged from the respiratory control and P/O ratios. No general membrane alterations or changes in the cytochrome contents were observed. BSA decreased the non-phosphorylating rates of respiration equally in both age groups. Thirteen different enzyme activities were assayed and normalised to protein content and citrate synthase activity. Most of the critical levels for detection of declines were <10%. In the 70+ group, the activity for fatty acid oxidation was decreased by approximately 20%. Two inherently low activities associated with oxidation of sarcoplasmic NADH were also decreased, probably related to the age change of fibre types. The remaining activities measured, e.g. those of pyruvate dehydrogenase, tricarboxylic acid cycle, respiratory chain, and ATP synthesis, were not observed to be lowered. Thus, the central bioenergetic systems appeared unaltered with age. The obvious discord with reported age declines of human skeletal muscle mitochondrial function is discussed. It is concluded that the present results are incompatible with the mitochondrial theory of aging.