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This study aimed to compare ambulatory blood pressure (ABP) response to accumulated standing (STAND), cycling (CYCLE), and walking (WALK) to a sitting-only (SIT) day in adults.Nine overweight or obese (body mass index, 28.7 ± 2.7 kg·m−2) adults (30 ± 15 yr) participated in this randomized crossover full-factorial study. Four conditions (WALK, STAND, CYCLE, and SIT) were randomly performed 1 wk apart. WALK, STAND, and CYCLE conditions consisted of progressively increasing activity time to accumulate 2.5 h during an 8-h simulated workday. WALK (1.0 mph) and STAND (0.0 mph) were completed on a treadmill placed underneath a standing-height desk. During CYCLE, participants pedaled on a Monark cycle ergometer at a cadence and energy expenditure equivalent to WALK. Participants remained seated during the SIT condition. Participants wore an ABP cuff from 0800 h until 2200 h on all conditions. Linear mixed models were used to test condition differences in systolic (SBP) and diastolic (DBP) blood pressure. Chi-square was used to detect frequency difference of BP load.There was a whole-day (during and after work hours) SBP and DBP treatment effect (P < 0.01). Systolic blood pressure during STAND (132 ± 17 mm Hg), WALK (133 ± 17 mm Hg), and CYCLE (130 ± 16 mm Hg) were lower compared with that during SIT (137 ± 17 mm Hg) (all P < 0.01). CYCLE was lower than STAND (P = 0.04) and WALK (P < 0.01). For DBP, only CYCLE (69 ± 12 mm Hg) was lower than SIT (71 ± 13 mm Hg; P < 0.01). Compared with SIT, WALK, STAND, and CYCLE reduced SBP load by 4%, 4%, and 13%, respectively (all P < 0.01).Compared with sitting, accumulating 2.5 h of light-intensity physical activity or standing during an 8-h workday may reduce ABP during and after work hours.