title: D-Isomer of gly-tyr-pro-cys-pro-his-pro peptide: a novel and sensitive in vitro trapping agent to detect reactive metabolites by electrospray mass spectrometry.
abstract: This paper describes a D-peptide isomer-based trapping assay using an LC/MS ion-trap spectrometer with an electrospray ionization (ESI) source as the analytical tool to study bioactivation of xenobiotics. Reactive metabolites were generated from parent compounds in in vitro incubations with different sources of CYP enzymes. A short D-isomer of gly-tyr-pro-cys-pro-his-pro proved to be a sensitive trapping agent and resistant to proteases. This method was tested with 16 probe substances. Acetaminophen, 1-chloro 2,4-dinitrobenzene, clozapine, diclofenac, imipramine, menthofuran, propranolol, pulegone and ticlopidine all formed D-peptide adducts, which were analogous to the GSH adducts previously described in the literature. New adducts were identified with clopidogrel (-Cl+peptide), nicotine (-CH(3+)H+peptide), nimesulide (+peptide) and tolcapone (+peptide), i.e., no GSH adducts of those drugs have been described in the literature. No adducts were identified with ciprofloxacin, ketoconazole and verapamil. In the literature no GSH adducts have been described with ciprofloxacin and verapamil. D-Peptide-based trapping proved to be a reliable and reproducible method to identify bioactivated intermediates. D-Peptide is a new and convenient protein trapping agent for use in early phase screening of bioactivation of new chemical entities and evaluation of toxic properties of chemicals.
authors.affiliation: University of Eastern Finland, Faculty of Health Sciences, School of Pharmacy, Kuopio, Finland. jaana.laine@uef.fi
authors.forename: Jaana E
authors.initials: JE
authors.lastname: Laine
authors.forename: Seppo
authors.initials: S
authors.lastname: Auriola
authors.forename: Markku
authors.initials: M
authors.lastname: Pasanen
authors.forename: Risto O
authors.initials: RO
authors.lastname: Juvonen
mesh_terms: D053000:Analytic Sample Preparation Methods; D000818:Animals; D001711:Biotransformation; D051544:Cytochrome P-450 CYP3A / Q000235:genetics / Q000378:metabolism; D004353:Drug Evaluation, Preclinical / Q000379:methods; D005978:Glutathione / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007202:Indicators and Reagents / Q000737:chemistry* / Q000378:metabolism; D007536:Isomerism; D051379:Mice; D008811:Mice, Inbred DBA; D008862:Microsomes, Liver / Q000378:metabolism; D009842:Oligopeptides / Q000737:chemistry* / Q000378:metabolism; D055550:Protein Stability; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000378:metabolism; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization; D015262:Xenobiotics / Q000737:chemistry / Q000493:pharmacokinetics*
chemical_list: D007202:Indicators and Reagents; D009842:Oligopeptides; D011994:Recombinant Proteins; D015262:Xenobiotics; C558053:glycyl-tyrosyl-prolyl-cysteinyl-prolyl-histidyl-proline; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human; D005978:Glutathione