Title
Liquid chromatography/tandem mass spectrometry detection of covalent binding of acetaminophen to human serum albumin
Author
Damsten, M.C.
Commandeur, J.N.M.
Fidder, A.
Hulst, A.G.
Touw, D.
Noort, D.
Vermeulen, N.P.E.
TNO Defensie en Veiligheid
Publication year
2007
Abstract
Covalent binding of reactive electrophilic intermediates to proteins is considered to play an important role in the processes leading to adverse drug reactions and idiosyncratic drug reactions. Consequently, both for the discovery and the development of new drugs, there is a great interest in sensitive methodologies that enable the detection of covalent binding of drugs and drug candidates in vivo. In this work, we present a strategy for the generation and analysis of drug adducts to human serum albumin. Our methodology is based on the isolation of albumin from blood, its digestion to peptides by pronase E, and the sensitive detection of adducts to the characteristic cysteine-proline- phenylalanine (CPF) tripeptide by liquid chromatography/tandem mass spectrometry. We chose acetaminophen (APAP) as a model compound because this drug is known to induce covalent binding to proteins when bioactivated by cytochromes P450 to its reactive N-acetyl-p-benzoquinoneimine metabolite. First, by microsomal incubations of APAP in presence of CPF and/or intact albumin, in vitro reference adducts were generated to determine the mass spectrometric characteristics of the expected CPF adducts and to confirm their formation on pronase E digestion of the alkylated protein. When applying this methodology to albumin isolated from blood of patients exposed to APAP, we were indeed able to detect the corresponding CPF adducts. Therefore, this strategy could be seen as a potential biomonitoring tool to detect in vivo reactive intermediates of drugs and drug candidates, e.g., in the preclinical and clinical development phase. Copyright © 2007 by The American Society for Pharmacology and Experimental Therapeutics.
Subject
Cytochrome P450
Paracetamol
Serum albumin
Article
Biological monitoring
Covalent bond
Drug protein binding
Human
Liquid chromatography
Microsome
Priority journal
Tandem mass spectrometry
Acetaminophen
Acetylcysteine
Animals
Benzoquinones
Chromatography, Liquid
Cysteine
Humans
Imines
Microsomes, Liver
Models, Biological
Molecular Structure
Oligopeptides
Protein Binding
Rats
Reproducibility of Results
Serum Albumin
Tandem Mass Spectrometry
Chemicals
CAS: cytochrome P450, 9035-51-2
Paracetamol, 103-90-2
Serum albumin, 9048-46-8
Acetaminophen, 103-90-2
Acetylcysteine, 616-91-1
Benzoquinones
Cysteine, 52-90-4
Imines
N-acetyl-4-benzoquinoneimine, 50700-49-7
Oligopeptides
To reference this document use:
http://resolver.tudelft.nl/uuid:d861620b-3441-415b-add5-d4b30d41e937
DOI
https://doi.org/10.1124/dmd.106.014233
TNO identifier
240113
ISSN
0090-9556
Source
Drug Metabolism and Disposition, 35 (8), 1408-1417
Document type
article