Bioactivation of 4-Methylphenol (p-cresol) via CYP-mediated Aromatic Oxidation in Human Liver Microsomes

Abstract

ABSTRACT:It has previously been proposed that 4-methylphenol (p-cresol) is metabolically activated by oxidation of the methyl group to form a reactive quinone methide. In the present study a new metabolism pathway is elucidated in human liver microsomes. Oxidation of the aromatic ring leads to formation of 4-methyl-ortho-hydroquinone, which is further oxidized to a reactive intermediate, 4-methylortho-benzoquinone. This bioactivation pathway is fully supported by the following observations: 1) one major and two minor glutathione (GSH) adducts were detected in microsomal incubations of p-cresol in the presence of glutathione; 2) a major metabolite of p-cresol was identified as 4-methyl-ortho-hydroquinone in microsomal incubations; 3) the same GSH adducts were detected in microsomal incubations of 4-methyl-ortho-hydroquinone; and 4) the same GSH adducts were chemically synthesized by oxidizing 4-methyl-ortho-hydroquinone followed by the addition of GSH, and the major conjugate was identified by liquid chromatography-tandem mass spectrometry and NMR as 3-(glutathione-S-yl)-5-methyl-ortho-hydroquinone. In addition, it was found that 4-hydroxybenzylalcohol, a major metabolite derived from oxidation of the methyl group in liver microsomes, was further converted to 4-hydroxybenzaldehyde. In vitro studies also revealed that bioactivation of p-cresol was mediated by multiple cytochromes P450, but CYP2D6, 2E1, and 1A2 are the most active enzymes for formation of quinone methide, 4-methyl-ortho-benzoquinone, and 4-hydroxybenzaldehyde, respectively. Implications of the newly identified reactive metabolite in p-cresol-induced toxicity remain to be investigated in the future.

Metabolism and Disposition of Eltrombopag, an Oral, Nonpeptide Thrombopoietin Receptor Agonist, in Healthy Human Subjects

“…8). The formation of methides has been reported as a bioactivation pathway for a number of drugs, although quinone methides are more typically involved (Bolton et al, 1990;Fan and Bolton, 2001;Kassahun et al, 2001;Zhang et al, 2003;Yan et al, 2005). An imine methide intermediate has been proposed as one of the bioactivation pathways for diclofenac (Grillo et al, 2008) and for 3-methylindole (Nocerini et al, 1985).…”

Rat Cytochrome P450 2C11 in Liver Microsomes Involved in Oxidation of Anesthetic Agent Propofol and Deactivated by Prior Treatment with Propofol

“…This is the first report, to our knowledge, of the deactivation of P450, especially CYP2C isoforms, by propofol. With regard to bioactivation via secondary oxidation of hydroxylated metabolites, methylphenol has been reported to be activated as evidenced by its glutathione adducts (Yan et al, 2005). In our preliminary study, effects of glutathione (3 mM) on propofol metabolism by rat liver microsomes were not seen under the present standard conditions.…”

Oxidative ipso Substitution of 2,4-Difluoro-benzylphthalazines: Identification of a Rare Stable Quinone Methide and Subsequent GSH Conjugate

“…are reported to undergo metabolic activation via formation of quinone methides. Quinone methides have also been reported as reactive intermediates of a number of xenobiotics including alkyl-substituted phenols such as pcresol (Yan et al, 2005), eugenol , polyphenols such as resveratrol (Steenwyk and Tan, 2010), catechols (Iverson et al, 1995), and bis-hydroxy toluene (Tajima et al, 1985;Bolton et al, 1990;Bolton and Thompson, 1991) and at times may even be formed by spontaneous rearrangement of o-quinones with 4-allylic substituents (Bolton et al, 1994).…”