LC-MS/MS based detection and characterization of covalent glutathione modifications formed by reactive drug of abuse metabolites

Abstract

1. Conjugation with the tripeptide glutathione (GSH) is a common mechanism of detoxification of many endogenous and exogenous compounds. This phenomenon typically occurs through the formation of a covalent bond between the nucleophilic free thiol moiety of GSH and an electrophilic site on the compound of interest.

2. While GSH adducts have been identified for many licit drugs, there is a lack of information on the ability of drugs of abuse to adduct GSH. The present study utilized a metabolic assay with GSH as a nucleophilic trapping agent to bind reactive drug metabolites formed in situ.

3. Extracted ion MS spectra were collected via LC-QqQ-MS/MS for all potentially significant ions and examined for fragmentation common to GSH-containing compounds, followed by confirmation of adduction and structural characterization performed by LC-QTOF-MS/MS.

4. In addition to the two positive controls, of the 14 drugs of abuse tested, 10 exhibited GSH adduction, with several forming multiple adducts, resulting in a total of 22 individual identified adducts. A number of these are previously unreported in the literature, including those for diazepam, naltrexone, oxycodone and Δ⁹-THC.

Keywords:

• Glutathione
• LC-MS/MS
• drugs of abuse
• adducts

Disclosure statement

No potential conflict of interest was reported by the authors.

Additional information

Funding

This project was supported by Award No. 2015-NE-BX-K001 (to APD), awarded by the National Institute of Justice, Office of Justice Programs, U.S. Department of Justice. The opinions, findings, and conclusions or recommendations are those of the authors and do not necessarily reflect those of the Department of Justice.