Carbon monoxide (CO) is a ubiquitous environmental pollutant widely recognized for its ability to inhibit cytochrome P450-mediated metabolism of xenobiotics in vitro. In recent years, the importance of the lung in the metabolic disposition of certain airborne and systemically administered xenobiotics has been demonstrated. The purpose of this investigation was to establish a threshold for the CO-induced inhibition of cytochrome P450-mediated activity in the isolated perfused rabbit lung and to determine if hemoglobin would alter the carbon monoxide-cytochrome P450 interaction. On the basis of its half-life and the stoichiometry of its metabolism, aminopyrine was shown to be a good substrate for monitoring mixed function oxidase activity in the intact rabbit lung. First-order rate constants for aminopyrine metabolism were significantly lower in isolated rabbit lungs perfused with either artificial medium (39%) or whole blood (67%) and ventilated with a 7.5% CO/20% O2 mixture for 2.5 hr than in the respective control lungs ventilated with breathing air. The threshold level (7.5% CO) for this inhibition is the same in lungs perfused with artificial medium and in whole blood-perfused lungs and is well above environmentally relevant levels of exposure.