Resorufin ether O-dealkylase activities in the isolated perfused rat lung

Previous studies have shown that ethoxyresorufin O-deethylase (EROD), methoxyresorufin O-demethylase (MROD), and benzyloxyresorufin O-debenzylase (BROD) are sensitive assays for determining the activities of cytochromes P-450 (CYP) 1A1 (CYP1A1), 1A2 (CYP1A2), and 2B1/2B2 (CYP2B1/2B2), respectively, in microsomes. The objectives of the current study were to determine these activities in the isolated perfused rat lung, assess the effect thereon of animal pretreatment with the cytochrome P-450 inducer pyridine, and determine the CYP isozyme specificity of the reactions. In perfused lungs from untreated rats, EROD, MROD, and BROD activity (pmol resorufin formed/g lung/min) was 40, 4.1, and 5.0, respectively. Pyridine pretreatment increased EROD and MROD activity 3 times and. 3.2 times, respectively, but did not affect BROD activity. The CYP1A2-specific substrate phenacetin inhibited MROD activity but not EROD or BROD activity, and was metabolized to N-acetyl p-aminophenol (APAP) in the perfused, lung. This O-deethylation of phenacetin was induced 2.5 times by animal pretreatment with pyridine, and was inhibited by methoxyresorufin (MR) but not by ethoxyresorufin (ER) or benzyloxyresorufin (BR). No evidence was seen of further biotransformation of the major O-dealkylation product resorufin. The results show that (1) CYP1A2 is present in the isolated perfused rat lung as are CYP1A1 and CYP2B1/2; (2) pyridine induces both forms of CYP1A in the lung; and (3) ER and MR are sensitive substrates for the rapid and sensitive determination of the catalytic activities of CYP1A1 and CYP1A2, respectively, in the intact lung.