Simultaneous Determination of Glycyrrhizin, Glycyrrhetic Acid and Glycyrrhetic Acid Mono-Glucuronide in Shakuyaku-kanzo-to Incubated with Rat Feces by Semi-micro High-Performance Liquid Chromatogaphy

Abstract

A method for semi-micro high-performance liquid chromatography (HPLC) has been established for the simultaneous determination of glycyrrhizin (GL), glycyrrhetic acid (GA) and glycyrrhetic acid mono-glucuronide (GAMG) in incubation mixtures of rat feces with Shakuyaku-kanzo-to decoction (combination of licorice root and peony root). The analysis could be accomplished within 20 min with a TSKgel ODS-80TsQA (150×2.0 mm i.d.) column by linear gradient elution using a mobile phase containing aqueous phosphoric acid and acetonitrile at a flow rate of 0.2 ml·min^-1, a thermostatic oven at 25°C, and detection at 254 nm. The detection limits of these compounds were 0.1—0.85 pmol per injection (5 μl). The concentrations of GL and its metabolites in the incubation mixture after continuous consumption of Shakuyaku-kanzo-to were significantly different compared with those of untreated control. GL-hydrolysis of rat feces was enhanced by pre-consumption of Shakuyaku-kanzo-to.