Aim: Mitochondrial dysfunction has been suggested to play a significant role in obesity and insulin resistance. The aim of the present study was to investigate if changes in obesity and insulin resistance were related to similar changes in mitochondrial capacity for hydrogenperoxide release in Zucker diabetic fatty rats and their lean littermates. Methods: Thirty-four rats were used in this study. Rats were either lean or obese Zucker rats killed at 5-6 (young) or 12-14 (adults) weeks of age. Mitochondria were isolated from soleus muscles; respiration and release of hydrogenperoxide were determined and related to citrate synthase activity to determine intrinsic mitochondrial function. Mitochondrial-specific super-oxide dismuthase (MnSOD) protein content was determined in isolated mitochondria and muscle homogenate. Catalase protein content was determined in muscle homogenate. Results: Young lean and obese rats had a higher mitochondrial respiration when using palmitoyl-l-carnitine as substrate compared with adult lean and obese rats. The obese strain had higher mitochondrial hydrogenperoxide release but only in the adult animals. In both lean and obese animals, increased age was associated with increased mitochondrial hydrogenperoxide release. MnSOD tended to be higher in the obese strain in the isolated mitochondria. Regardless of age, catalase protein content was significantly lower in the obese rats. Conclusions: This study shows that the augmented increase in obesity and insulin resistance seen in Zucker diabetic fatty rats is associated with increased capacity for mitochondrial hydrogenperoxide release.