Objective: To determine the relationship between aging, life-style factors and health-related factors and endogenous sex hormone levels. Design: Cross-sectional study of 400 independently living men between 40 and 80 years of age. Methods: After exclusion of subjects who were not physically or mentally able to visit the study center, 400 men were randomly selected from a population-based sample. Total testosterone (TT), bioavailable testosterone (BT) (i.e. not bound to sex hormone-binding globulin (SHBG)), SHBG, estradiol (E2) and dehydroepiandrosterone-sulfate (DHEA-S) were investigated for their relationship with age, body mass index (BMI), waist circumference, smoking, physical activity and general health status. Multivariate models using ANCOVA analyses were used to examine the contribution of life-style factors to sex hormone variability. Results: TT, BT and DHEA-S decreased with age; 0.2, 0.7 and 1.2%/year respectively. SHBG showed an increase with age of 1.1%/year. No changes with age were found for E2. General health status modified the association of TT and SHBG with age (P interaction 0.10 and 0.002 respectively). Increased BMI and waist circumference were associated with decreased TT, BT, SHBG and DHEA-S and increased E2 (all P < 0.01). Current smoking, lower alcohol intake and a higher physical activity score were associated with higher TT and SHBG levels. Conclusion: This study showed the important determinants of sex hormones were age, BMI, waist circumference, smoking, general health status and physical activity. Furthermore, it can be concluded that general health status modified the effect between sex hormones and age. For future observational studies it should be taken into account that the above-mentioned determinants may alter the association between sex hormones and diseases and related conditions.