Determination of trifolirhizin in rat plasma by UPLC: Application to a pharmacokinetic study

• A UPLC method was fully validated to determine trifolirhizin in rat plasma.
• This is a new analysis method for trifolirhizin.
• The plasma sample was prepared by liquid–liquid extraction with ethyl acetate.
• Trifolirhizin was analyzed in rat plasma up to 2 h after iv administration.

In this study, a simple, sensitive, and robust analytical method based on ultra-performance liquid chromatography (UPLC) has been developed for the determination of trifolirhizin in rat plasma using pirfenidone as internal standard (IS). After sample preparation by a simple liquid–liquid extraction, chromatography was performed on an Acquity UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm particle size) and ultraviolet detection set at a wavelength of 366 nm. The method was linear over the concentration range 25–1000 ng/mL with a lower limit of quantification (LLOQ) of 25 ng/mL. Inter- and intra-day precision (RSD%) were all within 10.2% and the accuracy (RE%) was equal or lower than 9.3%. The recovery was in the range of 78.5–86.4% for trifolirhizin and 87.4% for IS. Stability studies showed that trifolirhizin was stable under a variety of storage conditions. The method was successfully applied to a pharmacokinetic study involving oral administration of trifolirhizin to rats.