Simultaneous determination of three triterpenes in rat plasma by LC–MS/MS and its application to a pharmacokinetic study of Rhizoma Alismatis extract

• A sensitive and specific LC–MS/MS method was developed for simultaneous determination of alisol A (A), alisol A 23-acetate (A23) and alisol A 24-acetate (A24) in rat plasma.
• In view of the existence of mutual transformation A23 and A24in vivo, simultaneous determination of A23 and A24 is more accurate in rat plasma.
• The instability of A23 and A24 in solution have been solved
• The LC–MS/MS used in this paper was achieved in 100 μL plasma which was more suitable for small animal pharmacokinetic study due to small blood volume.
• We used this method for the pharmacokinetic study of RAE in rat.

We have developed a sensitive and specific LC–MS/MS method for the simultaneous determination of alisol A (A), alisol A 23-acetate (A23) and alisol A 24-acetate (A24), the major active components in Rhizoma Alismatis extract (RAE), in rat plasma. In brief, plasma samples were extracted by methyl tert-butyl ether and chromatographically separated by using a C18 column. A tandem mass spectrometric detection with an electrospray ionization (ESI) interface was conducted via multiple reaction monitoring (MRM) under positive ionization mode. This method was validated for specificity, linearity, accuracy (within ±15.4%), intra- and inter-day precision (CV < 11.4%) over the concentration range of 25–5000 ng/mL for A, and 5–1000 ng/mL for both A23 and A24. The significantly lower detection limit was determined as 25 ng/mL for A, 5 ng/mL for A23 and A24. This validated method of ours was then used to study the pharmacokinetics of RAE in rat. The elimination half-lives (t1/2) of A, A23 and A24 was determined as 0.75, 0.83 and 0.82 h respectively after intravenous injection, and the oral absolute bioavailability of A, A23 and A24 was 43.1 ± 18.1%, 6.3 ± 1.5% and 7.9 ± 1.2%. This new determination method of us for alisols is proven to very useful to study the pharmacological activities of RAE in future.