Cytochromes P450, 1A2, and 2C9 are responsible for the human hepatic O-demethylation of R- and S-naproxen

Biochem Pharmacol. 1996 Apr 26;51(8):1003-8. doi: 10.1016/0006-2952(96)85085-4.


A preliminary report implicated cytochrome P450 (CYP) 2C9 in the human liver microsomal O-demethylation of S-naproxen, suggesting that this pathway may be suitable for investigation of human hepatic CYP2C9 in vitro. Kinetic and inhibitor studies with human liver microsomes and confirmatory investigations with cDNA-expressed enzymes were undertaken here to define the role of CYP2C9 and other isoforms in the O-demethylation of R- and S-naproxen. All studies utilised a newly developed sensitive and specific HPLC assay that measured the respective O-desmethyl metabolites of R- and S-naproxen in incubations of human liver microsomes and in COS cell lysates. Microsomal R- and S-naproxen O-demethylation kinetics followed Michaelis-Menten kinetics, with respective mean apparent Km values of 123 microM and 143 microM. Sulfaphenazole, a specific inhibitor of CYP2C9, reduced the microsomal O-demethylation of R- and S-naproxen by 43% and 47%, respectively, and the CYP1A2 inhibitor furafylline decreased R- and S-naproxen O-demethylation by 38% and 28%, respectively. R,S-Mephenytoin was a weak inhibitor of R- and S-naproxen O-demethylation, but other CYP isoform specific inhibitors (e.g., coumarin, diethyldithiocarbamate, quinidine, troleandomycin) had little or no effect on these reactions. cDNA-expressed CYP2C9 and CYP1A2 were both shown to O-demethylate R- and S-naproxen. Apparent Km values (92-156 microM) for the reactions catalysed by the recombinant enzymes were similar to those observed for human liver microsomal R- and S-naproxen O-demethylation. The data demonstrate that CYP2C9 and CYP1A2 together account for the majority of human liver R- and S-naproxen O-demethylation, precluding the use of either R- or S-naproxen as a CYP isoform-specific substrate in vitro and in vivo.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Aryl Hydrocarbon Hydroxylases*
  • Cytochrome P-450 CYP1A2 / chemistry*
  • Cytochrome P-450 CYP2C9
  • Cytochrome P-450 Enzyme Inhibitors
  • Cytochrome P-450 Enzyme System / chemistry*
  • Humans
  • Kinetics
  • Methylation
  • Microsomes, Liver / enzymology*
  • Naproxen / chemistry*
  • Oxidoreductases, O-Demethylating / chemistry*
  • Recombinant Proteins / chemistry
  • Stereoisomerism
  • Steroid 16-alpha-Hydroxylase*
  • Steroid Hydroxylases / chemistry


  • Cytochrome P-450 Enzyme Inhibitors
  • Recombinant Proteins
  • Naproxen
  • Cytochrome P-450 Enzyme System
  • Oxidoreductases, O-Demethylating
  • Steroid Hydroxylases
  • CYP2C9 protein, human
  • Cytochrome P-450 CYP2C9
  • Aryl Hydrocarbon Hydroxylases
  • CYP1A2 protein, human
  • Cytochrome P-450 CYP1A2
  • Steroid 16-alpha-Hydroxylase