Nitrogen oxide (NO) species are markers for oxidative stress that may be pathogenic in acute lung injury (ALI). We tested two hypotheses in patients with ALI: (1) higher levels of urine NO would be associated with worse clinical outcomes, and (2) ventilation with lower VT would reduce urine NO as a result of less stretch injury. Urine NO levels were measured by chemiluminescence in 566 patients enrolled in the National Heart Lung and Blood Institute Acute Respiratory Distress Syndrome Network trial of 6 ml/kg versus 12 ml/kg VT ventilation. The data were expressed corrected and uncorrected for urine creatinine (Cr). Higher baseline levels of urine NO to Cr were associated with lower mortality (odds ratio, 0.43 per log(10) increase in the ratio), more ventilator-free days (mean increase, 1.9 d), and more organ-failure-free days (mean increase, 2.3 d) on multivariate analysis (p < 0.05 for all analyses). Similar results were obtained using urine NO alone. NO to Cr levels were higher on Day 3 in the 6 ml/kg than in the 12 ml/kg VT group (p = 0.04). Contrary to our hypothesis, higher urine NO was associated with improved outcomes in ALI at baseline and after treatment with the 6 ml/kg VT strategy. Higher endogenous NO may reflect less severe lung injury and better preservation of the pulmonary and systemic endothelium or may serve a protective function in patients with ALI.