title: Aromatic hydroxylation of salicylic acid and aspirin by human cytochromes P450.
abstract: Aspirin (acetylsalicylic acid) is a well-known and widely-used analgesic. It is rapidly deacetylated to salicylic acid, which forms two hippuric acids-salicyluric acid and gentisuric acid-and two glucuronides. The oxidation of aspirin and salicylic acid has been reported with human liver microsomes, but data on individual cytochromes P450 involved in oxidation is lacking. In this study we monitored oxidation of these compounds by human liver microsomes and cytochrome P450 (P450) using UPLC with fluorescence detection. Microsomal oxidation of salicylic acid was much faster than aspirin. The two oxidation products were 2,5-dihydroxybenzoic acid (gentisic acid, documented by its UV and mass spectrum) and 2,3-dihydroxybenzoic acid. Formation of neither product was inhibited by desferrioxamine, suggesting a lack of contribution of oxygen radicals under these conditions. Although more liphophilic, aspirin was oxidized less efficiently, primarily to the 2,5-dihydroxy product. Recombinant human P450s 2C8, 2C9, 2C19, 2D6, 2E1, and 3A4 all catalyzed the 5-hydroxylation of salicylic acid. Inhibitor studies with human liver microsomes indicated that all six of the previously mentioned P450s could contribute to both the 5- and 3-hydroxylation of salicylic acid and that P450s 2A6 and 2B6 have contributions to 5-hydroxylation. Inhibitor studies indicated that the major human P450 involved in both 3- and 5-hydroxylation of salicylic acid is P450 2E1.
authors.affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232-0146, USA. Electronic address: mbojic@pharma.hr.
authors.forename: Mirza
authors.initials: M
authors.lastname: Bojić
authors.affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232-0146, USA. Electronic address: carl.a.sedgeman@vanderbilt.edu.
authors.forename: Carl A
authors.initials: CA
authors.lastname: Sedgeman
authors.affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232-0146, USA. Electronic address: cellavporte@gmail.com.
authors.forename: Leslie D
authors.initials: LD
authors.lastname: Nagy
authors.affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37232-0146, USA. Electronic address: f.guengerich@vanderbilt.edu.
authors.forename: F Peter
authors.initials: FP
authors.lastname: Guengerich
mesh_terms: D001241:Aspirin / Q000378:metabolism*; D001547:Benzaldehydes / Q000737:chemistry; D001711:Biotransformation; D065607:Cytochrome P-450 Enzyme Inhibitors / Q000494:pharmacology; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D003676:Deferoxamine / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D006900:Hydroxylation; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010755:Organophosphates / Q000737:chemistry; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species / Q000737:chemistry; D011994:Recombinant Proteins / Q000378:metabolism; D020156:Salicylic Acid / Q000378:metabolism*
chemical_list: C105855:2,4-dihydroxybenzaldehyde 4-phosphate; D001547:Benzaldehydes; D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D010755:Organophosphates; D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; D009249:NADP; D003577:Cytochrome P-450 Enzyme System; D003676:Deferoxamine; D020156:Salicylic Acid; D001241:Aspirin
keywords: Aromatic hydroxylation; Aspirin (acetylsalicylic acid); Cytochromes P450; Salicylic acid
references.citation: Biochem Biophys Res Commun. 1988 Jan 15;150(1):436-43
references.pmid: 3337723
references.citation: J Biol Chem. 1974 Oct 10;249(19):6302-10
references.pmid: 4153601
references.citation: Drug Metab Dispos. 2006 Feb;34(2):199-202
references.pmid: 16258079
references.citation: J Am Pharm Assoc Am Pharm Assoc. 1958 Aug;47(8):602-5
references.pmid: 13563283
references.citation: Biochem Pharmacol. 1988 Jan 15;37(2):271-80
references.pmid: 3342084
references.citation: Arch Biochem Biophys. 1995 Jun 1;319(2):540-50
references.pmid: 7786040
references.citation: J Biol Chem. 2008 Nov 28;283(48):33698-707
references.pmid: 18818195
references.citation: Clin Pharmacol Ther. 2010 Jun;87(6):693-8
references.pmid: 20445534
references.citation: Clin Pharmacol Ther. 2003 Mar;73(3):264-71
references.pmid: 12621391
references.citation: J Vet Pharmacol Ther. 2000 Apr;23(2):73-9
references.pmid: 10849251
references.citation: Allergy Asthma Immunol Res. 2011 Oct;3(4):273-6
references.pmid: 21966608
references.citation: PLoS Biol. 2008 May 6;6(5):e107
references.pmid: 18462017
references.citation: Arch Biochem Biophys. 1998 Feb 15;350(2):324-32
references.pmid: 9473308
references.citation: Biochem J. 1991 Jun 15;276 ( Pt 3):753-7
references.pmid: 2064611
references.citation: Agents Actions. 1975 Dec;5(5):549-52
references.pmid: 1220558
references.citation: Drug Metab Dispos. 2000 Nov;28(11):1361-8
references.pmid: 11038165
references.citation: Anal Biochem. 1995 May 1;227(1):101-11
references.pmid: 7668368
references.citation: Xenobiotica. 1986 Mar;16(3):239-49
references.pmid: 3705620
references.citation: Biochemistry. 1992 Jul 7;31(26):6063-9
references.pmid: 1627548
references.citation: J Biol Chem. 2013 Dec 6;288(49):35210-21
references.pmid: 24129570
references.citation: Arch Biochem Biophys. 1994 Jul;312(1):59-66
references.pmid: 8031147
references.citation: Biochemistry. 2000 May 23;39(20):5929-39
references.pmid: 10821664
references.citation: Arch Biochem Biophys. 1993 Nov 1;306(2):443-50
references.pmid: 8215449
references.citation: J Chromatogr. 1983 Feb 11;272(2):325-31
references.pmid: 6833429
references.citation: Arch Biochem Biophys. 2005 Aug 15;440(2):204-11
references.pmid: 16055078
references.citation: Biochem J. 1986 Jul 15;237(2):499-504
references.pmid: 3026319
references.citation: Curr Drug Metab. 2009 Nov;10(9):998-1008
references.pmid: 20214591
references.citation: J Biol Chem. 1986 Jan 15;261(2):909-21
references.pmid: 3079764
references.citation: Drug Metab Dispos. 2001 Sep;29(9):1176-82
references.pmid: 11502724
references.citation: Clin Pharmacol Ther. 1978 Jun;23(6):635-43
references.pmid: 417892
references.citation: Free Radic Biol Med. 1992 Dec;13(6):621-6
references.pmid: 1459481
references.citation: Arch Biochem Biophys. 1993 Aug 15;305(1):123-31
references.pmid: 8342945
references.citation: J Biol Chem. 1984 May 25;259(10):6447-58
references.pmid: 6327680
references.citation: Arch Biochem Biophys. 2001 Sep 15;393(2):255-61
references.pmid: 11556812