Structural elucidation of in vitro metabolites of emodin by liquid chromatography–tandem mass spectrometry

Abstract Liquid chromatography–electrospray ionization tandem mass spectrometry (LC‐ESI‐MS/MS) was employed to investigate the in vitro metabolism of emodin. Emodin was incubated with rat liver microsomes in the presence of a NADPH‐generating system, followed by extraction with ethyl acetate. After separation on a reversed‐phase C 18 analytical column with a linear gradient elution of methanol and 0.1% formic acid in water, negative electrospray ionization tandem mass spectrometry experiments were performed. As a result, the parent drug and its six metabolites were detected from rat liver microsomal incubations. The identification of the metabolites and elucidation of their structure were performed by comparing the changes in molecular masses (Δ M ), retention times and MS 2 spectral patterns of metabolites with those of parent drug. Besides three mono‐hydroxylated metabolites ( ω ‐hydroxyemodin, 2‐hydroxyemodin, 4‐hydroxyemodin), three other metabolites were identified, which were emodic acid, 3‐carbomethoxy‐6‐methoxy‐1,8‐dihydroxyanthraquinone and physcion, respectively. Copyright © 2008 John Wiley & Sons, Ltd.