Simultaneous determination of senecionine, adonifoline and their metabolites in rat serum by UPLC–ESIMS and its application in pharmacokinetic studies

A rapid, selective and sensitive ultra-performance liquid chromatography–electrospray ionization mass spectrometry (UPLC–ESIMS) method was firstly developed and validated for the simultaneous determination of two hepatotoxic pyrrolizidine alkaloids (HPAs), senecionine (SEN), adonifoline (ADO), and their N-oxides (SENNOX and ADONOX), the main metabolites in rat serum. The whole analysis was achieved within 4.5 min by gradient elution on an ACQUITY UPLC BEH C18 column (50 mm × 2.1 mm, i.d. 1.7 μm) following a solid phase extraction for serum samples. Good linearity was achieved using weighted (1/x2) least squares linear regression over a 1600-fold dynamic range for SEN and ADO (LLOQ was about 0.006 μg/ml) and 800-fold dynamic range for SENNOX and ADONOX (LLOQ was about 0.012 μg/ml). The R.S.D. of intra- and inter-day precision was below 4.91% and 11.15% respectively, while the R.E. of accuracy was within 4.52%, 6.81%, 2.69%, and 7.12% for SEN, SENNOX, ADO, and ADONOX, respectively. The developed method was successfully applied to the in vivo pharmacokinetic study in rats after intravenous administration of SEN and ADO.