Aim To investigate the time-dependent changes in serum concentrations of endogenous inhibitor of nitric oxide synthase (NOS) asymmetric dimethylarginine (ADMA) in streptozotocin-induced diabetic rats and age-matched control rats, and to determine whether elevated endogenous ADMA is related to metabolic control and is implicated in the endothelium-dependent vasodilation dysfunction of diabetic rats at different stage of diabetic duration. Methods Serum levels of ADMA and L-arginine were measured by high-performance liquid chromatography at stages of 2-, 4- and 8-week diabetic duration. Thoracic aortic rings from diabetic rats with different diabetic duration and their age-matched control rats were studied for endothelium-dependent relaxation response to acetylcholine in vitro. Serum concentrations of glucose, glycosylated serum protein, and malondialdehyde, derived from lipid peroxidation were also examined to estimate metabolic control. Results Serum levels of ADMA significantly elevated in diabetic rats with 2-, 4-, and 8-week diabetic duration compared with their age-matched control rats(3.71±0.44 μmol/L vs 1.04±0.46 μmol/L for 2-week,3.54±1.70 μmol/L vs 0.95 ±0.13 μmol/L for 4-week,3.21±1.13 μmol/L vs 1.03±0.20 μmol/L for 8-week,n=5～6,all p<0.01), whereas serum levels of L-arginine were not different between control and diabetic rats at stages of 2-, 4-, and 8-week diabetic duration. Accordingly, the ratio of L-arginine and ADMA in diabetic rats was lower than that in their age-matched control rats. This elevation of ADMA was accompanied by impairment of relaxation response to acetylcholine of aortic rings in diabetic rats. Serum levels of glucose, glycosylated serum protein, and malondialdehyde were significantly increased in parallel with the elevation of ADMA in diabetic rats with 2-, 4-, and 8-week diabetic duration compared with their age-matched control rats. Conclusion These results reveal that the extent of elevation in serum ADMA in streptozotocin-induced diabetic rats with 2-, 4-, and 8-week diabetic duration is not correlated with the length of diabetic duration. This elevation of endogenous inhibitor of NOS in diabetic rats may relate to metabolic control of the disease and may be impli-cated in the endothelium-dependent vasodilation dysfunction associated with diabetes.