On the Role of ROS and Glutathione in the Mode of Action Underlying Nrf2 Activation by the Hydroxyanthraquinone Purpurin

Purpurin is a major anthraquinone present in the roots of Rubia cordifolia (madder). known to activate Nrf2 (Nuclear transcription factor erythroid 2-related 2) EpRE (electrophile responsive element) mediated gene expression as potential beneficial effect. This study aimed elucidate balance between electrophilicity or pro-oxidant activity purpurin underlying induction. For this, activation with modified intracellular glutathione (GSH) levels was measured an CALUX reporter assay. In addition, both cell-free and ROS formation (intracellular) GSH at different pH were quantified using DCF-DA adduct evaluated by UPLC LC-TOF-MS analysis. GSSG following incubations LC-MS/MS. We show that induction significantly increased cells buthionine sulfoximine depleted levels, while decreased upon incubation N-acetylcysteine being precursor GSH. incubations, increasing pointing role for deprotonated form purpurin. Upon physiological pH, appeared negligible (<1.5% added purpurin). The addition resulted conversion reduced formation. Together these results demonstrate originates from not its electrophilicity, which becomes especially relevant when can no longer scavenge ROS. demonstrated efficiency related anthraquinones will depend on (i) their pKa level deprotonation (ii) oxidation (iii) levels. Thus, depends electrophilicity.