A simple and sensitive LC-MS/MS method for determination of miltirone in rat plasma and its application to pharmacokinetic studies

A rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantification of miltirone concentration in rat plasma. The cytotoxic activity of miltirone was firstly evaluated by the MTT assay and compared with other tanshinones. Quantification was carried out on an Agilent triple quadrupole LC-MS system using multiple reaction monitoring (MRM) mode in positive mode. After simple protein precipitation with acetonitrile, the chromatographic separation of miltirone was achieved by using a Waters Symmetry C18 analytical column (2.1 mm × 100 mm, 3.5 μm) with a mobile phase of acetonitrile (A)-water (B) (75:25, v/v) containing 0.5% formic acid. The monitored transitions were set at m/z 283.1 → 223.1 and m/z 361.0 → 232.9 for miltirone and IS, respectively. The calibration curve was linear over the concentration range of 0.5-200 ng/mL with lower limit of quantification of 0.5 ng/mL. The intra- and inter-day accuracy and precision of miltirone were both within acceptable limits. The developed method was successfully applied to a pharmacokinetic study of following oral administration of 20, 40, 60 mg/kg and an intravenous administration of 0.5 mg/kg to rats. The results indicated that miltirone had linear pharmacokinetic properties within the tested dosage range and was poorly absorbed with an absolute bioavailability of approximately 3.4%.