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Objective: We recently reported a small increase in plasma nitrate/nitrite (NOx) following oral intake of the novel nitric oxide donor, 2-nitrooxy-ethyl-2-amino-3-methylbutanoate (2-NO). However, in that study subjects remained sedentary during the 60 minute data collection time. It is now believed that the 2-NO molecule may be activated via increased blood pressure, secondary to physical stress. We compared the effects of 2-NO on blood pressure, heart rate (HR), and NOx in normotensive men. Methods: 15 healthy men (28.5±0.8 yrs) reported to the lab on two different days separated by two weeks. They were randomly assigned in a double-blind, cross-over manner to receive either a 2-NO tablet or a placebo tablet before performing two, two-minute sets of sit-up exercise (exercise sets performed 20 minutes apart). Systolic (SBP) and diastolic (DBP) blood pressure and HR were measured before and for 20 minutes following each set of exercise. Blood was collected via an indwelling venous catheter at the same times and analyzed for NOx. Results: For both conditions, subjects performed a near identical number of repetitions (2-NO: 101; placebo: 100). HR was not different between conditions (p>0.05). However, SBP was lower with 2-NO compared to placebo (p=0.03), in particular during 20 minute recovery period following set 2 (p=0.04; ~6 mm Hg lower). 2-NO also resulted in a lower DBP during the same time period following set 2 (p=0.02; ~5 mm Hg lower). NOx was increased approximately 10% above baseline with 2-NO (p=0.09) but remained stable or decreased slightly with placebo. Conclusions: These data indicate that 2-NO can lower blood pressure in response to acute exercise in a sample of healthy, normotensive men. The change in blood pressure may be associated with an increase in circulating NOx.