Elevated plasma homocysteine (tHcy) is considered as a risk factor for cardiovascular events, and has been associated with arterial stiffness and subclinical atherosclerosis in subjects with classical cardiovascular risk factors. The aim of this study is to investigate the association of plasma tHcy with functional and structural changes in the large arteries by measuring aortic pulse-wave velocity (PWV) and carotid artery intima-media thickness (CIMT). In a population-based sample of 376 middle-aged and elderly men, tHcy levels were measured by using fluorescence polarization immunoassay. Aortic stiffness was assessed non-invasively by measuring carotid-femoral PWV with the use of applanation tonometry. CIMT was measured by ultrasonography. We used multivariate linear regression analysis with the log-transformed value of tHcy as the determinant, and aortic PWV and CIMT as outcomes. In the univariate model, PWV increased with increasing tHcy concentration; PWV increased by 2.64 m/s (95% confidence interval (CI) 1.74; 3.54) per unit change in log tHcy. After adjustment for confounders, no statistically significant association remained; PWV increased by 0.42 m/s (95% CI -0.27; 1.11) per unit change in log tHcy. Furthermore, in the univariate model, CIMT increased significantly with increasing tHcy concentration; CIMT increased by 0.19 mm (95% CI 0.11; 0.26) per unit increase log tHcy. This association was attenuated and did not remain significant after additional adjustment for age and cardiovascular confounders (β = 0.06 (95%CI -0.01; 0.13)). The results of this study do not support the presence of an independent relationship between circulating tHcy levels and large artery stiffness and thickness.