Inactivation of CYP2D6 by Berberrubine and the Chemical Mechanism

Berberrubine (BRB), belonging to the benzylisoquinoline alkaloid, is a main metabolite of berberine <i>in vivo</i>. BRB was previously proven to undergo metabolic activation mediated by P450s. In this study, the chemical interactions between BRB and CYP2D6 enzyme were investigated. First, a variety of P450s participated in the metabolism of berberine transformed to BRB, but CYP2D6 was the most involved enzyme. A time-, concentration-, and nicotinamide adenine dinucleotide phosphate (NADPH)-dependent inhibition of CYP2D6 was caused by BRB. The inhibitory effect of BRB on CYP2D6 was irreversible. The maximum reaction rate constants of inactivation (<i>k</i>_inact) and half-maximal inactivation (<i>K</i>_I) of BRB on CYP2D6 were 0.0410 min^-1 and 3.798 μM, respectively. Metoprolol, a classic substrate of CYP2D6, attenuated CYP2D6 from inactivation by BRB. Glutathione (GSH) and catalase/superoxide dismutase failed to protect against the inactivation of CYP2D6 caused by BRB. Three cys-based adducts derived from the reaction of electrophilic metabolites of BRB with CYP2D6 were detected by ultra performance liquid chromatography-mass spectrometry (UPLC-MS)/MS. The reactive metabolites derived from BRB might be responsible for the inactivation of CYP2D6. In summary, BRB was characterized as a mechanism-based inactivator of CYP2D6.