Abstract
Autoxidation of para-aminophenol (PAP) has been proposed to account for the selective nephrotoxicity of this compound. However, other studies suggest that hepatic metabolites of PAP rather than the parent compound may be responsible for renal damage. These studies were designed to investigate PAP metabolism in isolated hepatocytes. We synthesized several proposed metabolites for analysis by HPLC/mass spectrometry and compared those results with HPLC/mass spectrometric analyses of metabolites found after incubating hepatocytes with PAP. Hepatocytes prepared from male Sprague-Dawley rats were incubated in Krebs-Henseleit buffer at 37°C for 5 h with 2.3 mM PAP under an atmosphere of 5% CO2/95% O2. Aliquots were withdrawn at 0.1 h of incubation and then hourly through 5 h of incubation. Reactions were terminated by the addition of acetonitrile. Hepatocyte viability was unaltered with PAP present in the incubation medium. We found that hepatocytes converted PAP to two major metabolites (PAP-GSH conjugates and PAP-N-acetylcysteine conjugates) and several minor metabolites [PAP-O-glucuronide, acetaminophen (APAP), APAP-O-glucuronide, APAP-GSH conjugates, and 4-hydroxyformanilide]. Preincubating hepatoyctes with 1-aminobenzotriazole, an inhibitor of cytochromes P450, did not alter the pattern of PAP metabolism. In conclusion, we found that PAP was metabolized in hepatocytes predominantly to PAP-GSH conjugates and PAP-N-acetylcysteine conjugates in sufficient quantities to account for the nephrotoxicity of PAP.
Footnotes
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Send reprint requests to: Joan B. Tarloff, Ph.D., Dept. of Pharmaceutical Sciences, University of the Sciences in Philadelphia, 600 South 43rd Street, Philadelphia, PA 19104-4495. E-mail:j.tarlof{at}usip.edu
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↵1 Current address: American Cyanamid Company, Agricultural Products Research Division, P.O. Box 400, Princeton, NJ 08543-0400.
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These studies were supported by U.S. Public Health Service Grant GM51361 (J.B.T.).
- Abbreviations used are::
- PAP
- para-aminophenol
- MS
- mass spectrometry
- FPAP
- 4-hydroxyformanilide
- ABT
- 1-aminobenzotriazole
- PAP-NACys
- PAP-S-(N-acetylcysteine) conjugates
- TSP
- thermospray
- GC
- gas chromatography
- FIA
- flow injection analysis
- ESI
- electrospray ionization
- PBQI
- para-benzoquinoneimine
- APAP
- acetaminophen
- APAP-Gluc
- acetaminophen-O-glucuronide
- PAP-GSH
- PAP-S-glutathione conjugates
- ACN
- acetonitrile
- PAP-Gluc
- 4-aminophenyl-O-glucuronide
- PAP-Cys
- PAP-S-cysteine conjugates
- Received October 27, 1999.
- Accepted April 18, 2000.
- The American Society for Pharmacology and Experimental Therapeutics
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