Nitric oxide (NO) unavailability plays an important role in the progression of macrovascular diseases in Type 2 diabetes (T2DM). C59038T polymorphism in GTP cyclohydrolase 1 (GCH1) gene is a functional mutation involved in NO metabolism and cardiovascular risk in a multiethnic population. To clarify the relationship between C59038T polymorphism and macrovascular disease in T2DM, an association study was performed among 611 unrelated T2DM patients. C59038T polymorphism was detected by polymerase chain reaction (PCR) restriction fragment length polymorphism. The PCR products after digestion displayed three genotypes, including CC, CT, and TT. The prevalence of cardiovascular disease, cerebrovascular disease, and peripheral vascular disease was significantly higher in T2DM patients with TT genotype than those with CC or CT genotype (P<.001). Compared with CC or CC+CT genotype, T2DM patients with TT genotype had a significantly increased risk of macrovascular disease (P<.001, P=.001), with odds ratio for 4.717 [95% confidence interval: 3.056-7.370] and 4.082 (2.716-5.868), respectively. Subjects with TT genotype showed lower levels of plasma NOx (nitrite and nitrate), flow-mediated artery dilatation and activities of superoxide dismutase but higher levels of plasma malonaldehyde and intima-media thickness of carotid artery than those with CC or CT genotype (P<.05). This study demonstrated that in Chinese T2DM population, C59038T polymorphism was associated with an increased risk of macrovascular disease, which was likely due to its effects on NO metabolism, oxidative stress, and subsequently vascular dysfunction.