Human neutrophil elastase (HNE) is the predominant elastolytic enzyme in the sputum of cystic fibrosis (CF) patients. However, a variably small portion of the activity can be ascribed to Pseudomonas aeruginosa elastase (PaE). The purpose of these studies was to evaluate the activities of the two elastases in an in vivo model of acute lung injury (ALI). The elastolytic activity of Pseudomonas aeruginosa elastase (MW = 39K) and human neutrophil elastase (MW = 33K) were also examined using insoluble bovine neck and lung elastin. The ability of hamster serum to inhibit elastinolysis by the two elastases was also examined. On a per milligram protein basis, PaE was the more potent elastase, regardless of substrate, and it preferentially hydrolyzed lung relative to neck elastin. PaE is poorly inhibited by hamster serum compared to HNE. In vivo, PaE is much more efficient than HNE in inducing an acute lung injury in hamsters. The duration of effects induced by doses of the two proteases that produce similar acute biological effects are essentially identical. The increases of lung weight and total lavagable WBCs persist for at least 7 days. All other parameters return to baseline between 3 and 5 days. The predominant cells in the lavage 1 and 2 days post insult are PMNs. By day 7, the predominant cell is the macrophage. These data suggest that even though PaE is a minor component of the elastolytic activity in CF patients, it may still contribute significantly to the pathology of the disease.