Purpose This study aimed to investigate whether a reduction in daily sedentary behavior (SB) improves insulin sensitivity in adults with metabolic syndrome in 6 months, without adding intentional exercise training. Methods Sixty-four sedentary inactive middle-age adults with overweight and metabolic syndrome (mean (SD) age, 58 (7) yr; mean (SD) body mass index, 31.6 (4.3) kg·m-2; 27 men) were randomized into intervention and control groups. The 6-month individualized behavioral intervention supported by an interactive accelerometer and a mobile application aimed at reducing daily SB by 1 h compared with baseline. Insulin sensitivity by hyperinsulinemic euglycemic clamp, body composition by air displacement plethysmography, and fasting blood samples were analyzed before and after the intervention. SB and physical activity were measured with hip-worn accelerometers throughout the intervention. Results SB decreased by 40 (95% confidence interval, 17-65) min·d-1, and moderate-to-vigorous physical activity increased by 20 (95% confidence interval, 11-28) min·d-1 on average in the intervention group with no significant changes in these outcomes in the control group. After 6 months, fasting plasma insulin decreased (1 mU·L-1) in the intervention group compared with the control group (time-group, P = 0.0081), but insulin sensitivity did not change in either group. The changes in body mass or adiposity did not differ between groups. Among all participants, the changes in SB and body mass correlated inversely with the change in insulin sensitivity (r = -0.31, -0.44; P = 0.025, 0.0005, respectively). Conclusions An intervention aimed at reducing daily SB resulted in slightly decreased fasting insulin, but had no effects on insulin sensitivity or body adiposity. However, as the change in insulin sensitivity associated with the changes in SB and body mass, multifaceted interventions targeting to weight loss are likely to be beneficial in improving whole-body insulin sensitivity.