Detecting and characterizing reactive metabolites by liquid chromatography/tandem mass spectrometry

Abstract

Metabolic activation of a drug leading to reactive metabolite(s) that can covalently modify proteins is considered an initial step that may lead to drug-induced organ toxicities. Characterization of reactive metabolites is critical to designing new drug candidates with an improved toxicological profile. High performance liquid chromatography (HPLC) coupled with mass spectrometry (MS) predominates over all analytical tools used for screening and characterization of reactive metabolites. In this review, a brief description of experimental approaches employed for assessing reactive metabolites is followed by a discussion on the reactivity of acyl glucuronides and acyl coenzyme A thioesters. Techniques for high-throughput screening and quantitation of reactive metabolite formation are also described, along with proteomic approaches used to identify protein targets and modification sites by reactive metabolites. Strategies for dealing with reactive metabolites are reviewed. In conclusion, we discuss the challenges and future needs in this field of research.

The Phosphoinositide-Specific Phospholipase C Inhibitor U73122 (1-(6-((17β-3-Methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione) Spontaneously Forms Conjugates with Common Components of Cell Culture Medium

“…2C). MS 2 analysis resulted in loss of 129 Da, a common neutral loss observed for thiollinked glutathione conjugates (Ma and Subramanian, 2006). MS 3 analysis (Fig.…”

CPY3A4-Mediated Lopinavir Bioactivation and Its Inhibition by Ritonavir

“…dmd.aspetjournals.org (Evans et al, 2004;Li et al, 2011). GSH is commonly used to trap soft electrophiles, such as epoxides, ␣,␤-unsaturated carbonyls, quinones, quinone imines, quinone methides, imine methide, isocyanate, isothiocyanates, aziridinium, and episulfonium (Evans et al, 2004;Ma and Subramanian, 2006). Semicarbazide is used to trap hard electrophiles, such as aldehydes (Evans et al, 2004;Li et al, 2011).…”

“…4). DMP is further oxidized to form O_DMP, which can be transformed into reactive intermediates, such as quinone and/or quinone methide, and then be trapped by GSH (Evans et al, 2004;Ma and Subramanian, 2006). In the incubation of LPV in HLM containing GSH, two GSH-conjugated DMP adducts were identified (Fig.…”

Nucleophile addition of reduced glutathione on 2-methyl-2-nitroso compound: A combined electron paramagnetic resonance spectroscopy and electrospray tandem mass spectrometry study

“…In both types of conjugation, reduced glutathione can add to a double bond of aromatic compounds [6], to an epoxide group, to an isocyanate carbon [7], or substitute a halogen atom in alkyl-or aryl-halides [8]. Methods for the detection and characterization of metabolites arising from reactions involving GSH were thus developed, in particular using a mass spectrometric approach [9].…”