To determine the most sensitive early indicator of toxicity from exposure to O2, we measured respiratory clearance of 99mTc-diethylenetriamine pentaacetate (Tc-DTPA), pulmonary function, serum Factor VIII antigen, and plasma fibronectin values and monitored symptoms in six normal volunteers who breathed 21, 40, and 100% O2, administered in random order, for 17 h at least 1 wk apart. Twenty minutes after beginning O2, arterial PO2 differed among the three exposure groups. After exposure to the three concentrations of O2, there were no differences in Tc-DTPA clearance, vital capacity, FEV1/FVC, diffusing capacity, Factor VIII antigen, or fibronectin concentration. In contrast, all subjects complained of retrosternal pain during and after breathing 100% O2 (p less than 0.001). We conclude that with exposure to 100% O2, retrosternal pain, presumably from tracheal inflammation, occurs before detectable abnormalities of epithelial solute permeability (Tc-DTPA clearance), endothelial O2 injury (fibronectin concentration and Factor VIII), or pulmonary function. These findings indicate that symptoms are more sensitive than signs in detecting early O2 toxicity.