title: Flavin-containing monooxygenase activity in hepatocytes and microsomes: in vitro characterization and in vivo scaling of benzydamine clearance.
abstract: Liver microsomes, and more recently cryopreserved hepatocytes, are commonly used in the in vitro characterization of the metabolism of new xenobiotics. The flavin-containing monooxygenases (FMO) are a major non p450 oxidase present in liver microsomes and hepatocytes. Since FMO is known to be thermally labile, and this enzyme may be involved in the metabolic clearance of some drugs, we sought to more completely characterize the metabolic competency of this enzyme in cryopreserved hepatocytes and in liver microsomes preincubated under various conditions using benzydamine as an in vitro and in vivo probe. The metabolism of benzydamine to its major metabolite, the N-oxide, is mediated by FMO3 in humans. We found that the in vitro microsomal t(1/2) was 70% longer when incubations were prewarmed at 37 degrees C in the absence of NADPH compared with prewarming in the presence of an NADPH-regenerating system, and N-oxide formation was inhibited >99%. Interestingly, the in vivo clearance predicted from these incubations and from human hepatocytes overpredicted the observed clearance of benzydamine in humans (>10.5 versus 2.4 ml/min/kg). In contrast, rat hepatocytes successfully predicted rat in vivo benzydamine clearance to within approximately 30% (>68 versus 48 ml/min/kg). Benzydamine N-oxidation in liver microsomes from all common preclinical species demonstrated heat sensitivity. This information should be considered when extrapolating metabolism data of xenobiotics from these in vitro systems.
authors.affiliation: Discovery Pharmacokinetics, Dynamics and Metabolism, Pfizer Global Research and Development, Pfizer Inc, Eastern Point Rd, Groton, Connecticut 06340, USA. michael_fisher@groton.pfizer.com
authors.forename: Michael B
authors.initials: MB
authors.lastname: Fisher
authors.forename: Kwansik
authors.initials: K
authors.lastname: Yoon
authors.forename: Marcie L
authors.initials: ML
authors.lastname: Vaughn
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Strelevitz
authors.forename: Robert S
authors.initials: RS
authors.lastname: Foti
mesh_terms: D000818:Animals; D001591:Benzydamine / Q000737:chemistry / Q000378:metabolism*; D001711:Biotransformation / Q000187:drug effects / Q000502:physiology; D004285:Dogs; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D022781:Hepatocytes / Q000201:enzymology*; D006801:Humans; D008252:Macaca fascicularis; D008657:Metabolic Clearance Rate / Q000187:drug effects / Q000502:physiology; D051379:Mice; D008862:Microsomes, Liver / Q000201:enzymology*; D010105:Oxygenases / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D001591:Benzydamine; D010105:Oxygenases; C020066:dimethylaniline monooxygenase (N-oxide forming)