Objectives To investigate whether time spent watching television (a marker of sedentary behaviour) is associated with arterial stiffness, a major determinant of cardiovascular disease, and whether any such association could be explained by related deleterious levels of habitual physical activity (HPA) and/or other lifestyle and biological risk factors. Methods Prospective measures (ages 32 and 36 years) of television time and risk factors were retrieved from 373 participants (196 women) in whom stiffness of the carotid, brachial and femoral arteries was assessed by means of ultrasonography at age 36 years. Data were analysed with generalised estimating equations. Results Participants with stiffer carotid arteries spent more time (in min/day) watching television during the four preceding years than did those with less stiff arteries, as defined on the basis of the highest compared with the lowest gender-specific tertiles of the distensibility or compliance coefficients (reversed) or the Young's elastic modulus: +22.4 (95% CI 8.7 to 36.1), +18.4 (4.2 to 32.5) and +19.7 (6.0 to 33.4), respectively. These differences were independent of potential confounders, such as vigorous intensity HPA and other lifestyle risk factors, and could only in part (up to 31%) be explained by the adverse associations of television time with traditional biological risk factors. Qualitatively similar results were found for femoral, but not brachial, stiffness estimates. Conclusions Given the independent associations of time spent watching television and vigorous intensity HPA with arterial stiffness, our study suggests that not only promotion of physical activity, but also discouragement of sedentary behaviours should be targeted in younger adults to prevent arterial stiffening.