Background: To date, no longitudinal studies have examined the influence of sedentary behavior on metabolic syndrome development while accounting for cardiorespiratory fitness. Purpose and Methods: This prospective study examined the relationship between sedentary behavior and incident metabolic syndrome while considering the effects of physical activity and cardiorespiratory fitness on the association among 930 men enrolled in the Aerobics Center Longitudinal Study. Results: A total of 124 men developed metabolic syndrome during 8974 person-years of exposure. After adjusting for covariates, men with middle and high sedentary behavior had 65% and 76% higher risks of developing metabolic syndrome, respectively, than men with low sedentary behavior (linear trend P = .011). This association remained significant after additional adjustment for activity status and cardiorespiratory fitness. Cardiorespiratory fitness and physical activity were also inversely associated with metabolic syndrome, even after adjustment for sedentary behavior. Conclusions: The findings highlight the importance of reducing sedentary behavior, increasing physical activity, and improving cardiorespiratory fitness for preventing metabolic syndrome.