Both qualitative and quantitative changes in mitochondrial DNA (mtDNA) have been implicated in the pathogenesis of diabetes mellitus. It was previously found that decreased mtDNA content preceded the development of diabetes and mtDNA content correlated with the clinical parameters of insulin resistance syndrome, including diastolic blood pressure and waist-hip ratio. These results prompted one to look whether there are correlations between mtDNA content and the biochemical parameters of insulin resistance in non-diabetic subjects. MtDNA content of peripheral blood leukocytes was measured in Korean healthy young men, and this was correlated with various parameters of fuel metabolism at baseline and during euglycemic hyperinsulinemic clamps with indirect calorimetry. MtDNA content in peripheral blood leukocytes did not correlate with insulin sensitivity index or other metabolic variables such as body mass index (BMI), waist-to-hip ratio (WHR) and blood pressure. However, mtDNA content showed a positive significant correlation with fat oxidation rate during euglycemic clamps (r = 0.61, P < 0.05). Changes in fat oxidation rate and carbohydrate oxidation rate during the clamps were significantly correlated with mtDNA content (r = 0.65, P < 0.05, r = -0.65, P < 0.05, respectively). These results suggest that mtDNA content in peripheral blood may not correlate with insulin resistance per se but with some aspect of insulin resistance in healthy young men.