Mimicking the reactivity of drug metabolites: Biomolecule conjugation of an electrochemically-generated, reactive oxidation product of the antibiotic minocycline

Abstract

Minocycline is an antibiotic of the tetracycline family which is widely used to treat a range of medical conditions. Although it has been in use for more than 50 years, little information is available on its metabolism in the human body. In this study, we simulate the biotransformation of minocycline by means of electrochemistry coupled to mass spectrometry. This analytical technique has already been used successfully in several cases to imitate enzyme-catalyzed reactions. Using this approach, we could show the generation of multiple electrochemical oxidation products which were characterized by tandem mass spectrometry. A N-dealkylated product was found to correspond to a literature-known in vivo metabolite. Two further oxidation products were detected, one of which exhibiting a reactive quinone moiety formed through electrochemical oxidation. The reactivity of this transformation product was assessed by conjugation reactions with glutathione, human hemoglobin and human serum albumin as model biomolecules. For all three peptides, conjugation reactions took place within minutes, corresponding to the number of free cysteine residues in the respective molecule, which are particularly susceptible to electrophiles like quinones. For glutathione, serum albumin and α-hemoglobin, a single conjugation of the reactive transformation product took place, whereas a twofold conjugation was detected for β-hemoglobin. This project showcases the capability of the purely instrumental approach to simulate the metabolism of xenobiotics without an interfering matrix to screen for reactive transformation products and to assess the reactivity of these products with regard to biomolecules.