Oxidative stress (OS) increases during the human aging process, and the sedentary lifestyle could be a prooxidant factor. In this study, we determine the effect of sedentary lifestyle on OS during the aging process in Mexican women. A longitudinal study of two-year follow-up was carried out with 177 community-dwelling women (40-69 y) from Mexico City. We measured as OS markers plasma malondialdehyde, erythrocyte glutathione peroxidase (GPx) and superoxide dismutase (SOD), total plasma antioxidant status, uric acid level, antioxidant gap, and SOD/GPx ratio. To define OS using all the markers, we defined cut-off values of each parameter based on the 90th percentile of young healthy subjects and, we calculated a stress score (SS) ranging from 0 to 7, which represented the intensity of the marker modifications. All the women answered a structured questionnaire about prooxidant factors, including physical activity specially the type of activity, frequency, and duration, and they answered Spanish versions of self-assessment tests for establishing dysthymia and insomnia as potential confounders. Principal component and Poisson regression analysis were used as statistical tools, being two-year OS the primary outcome. The OS was considerate as
SS
≥
4
and sedentary lifestyle as <30 min/day of physical activity, beside several prooxidant factors and age that were covariables. SS is higher in sedentary lifestyle women after the two-year follow-up; although, the difference was statistically significant only in older women. Four principal components were associated with the OS, and 7 out of 8 prooxidant factors were important for the analysis, which were included in the Poisson model. The predictive factors for OS were the sedentary lifestyle (adjusted PR = 2.37, CI95%: 1.30–4.30,
p
<
0.01
), and age, in which the risk increases 1.06 (CI95%:1.02–2.11,
p
<
0.01
) by each year of age. Our findings suggest that a sedentary lifestyle increases the OS during the aging in Mexican women.