Cytotoxicity of paracetamol and 3,5-dihalogenated analogues: Role of cytochrome P-450 and formation of GSH conjugates and protein adducts.
The effect of 3,5-dihalogenation of paracetamol (PAR) on the cytotoxicity in rat hepatocytes isolated from beta-naphthoflavone pretreated, non-fasted rats, and the role of cytochrome P-450 in this regard, were studied. On incubation, 3,5-difluoro-PAR, 3,5-dichloro-PAR and 3,5-dibromo-PAR, as well as PAR, caused severe leakage of lactate dehydrogenase (LDH) which was preceded by a rapid concentration- and time-dependent depletion of intracellular glutathione (GSH). IC(50) values, representing the concentration of compound that caused 50% GSH depletion after 30 min of incubation, varied from 0.1 to 0.5 mM. This LDH leakage and GSH depletion could be inhibited by 1-ethynylpyrene. In hepatocytes from uninduced rats, GSH depletion was much less prominent and the concomitant LDH leakage almost completely absent. HPLC analysis of soluble metabolites and gas chromatography-mass spectrometry analysis, after alkaline peralkylation of the protein fraction, revealed (a) that 3,5-dihalogenated PAR analogues were liable to structure-related detoxification by glucuronidation, and (b) analogous to PAR, a substantial amount of each 3,5-dihalogenated PAR analogue was bioactivated by cytochrome P-450, ultimately leading to GSH-conjugates as well as (for 3,5-dichloro-PAR and 3,5-dibromo-PAR), protein adducts at regio-specific aromatic positions.
Bessems
J G
JG
Leiden/Amsterdam Centre for Drug Research, Division of Molecular Toxicology, Department of Pharmacochemistry, Vrije Universiteit, De Boelelaan 1083, 1081 HV Amsterdam, The Netherlands.
Van Stee
L L
LL
Commandeur
J N
JN
Groot
E J
EJ
Vermeulen
N P
NP