Simultaneous determination of icariin, naringin and osthole in rat plasma by UPLC–MS/MS and its application for pharmacokinetic study after oral administration of Gushudan capsules

• Icariin, naringin and osthole were simultaneously determined by UPLC–MS/MS method.
• Compared with former methods, it reached to a shorter run time of 6 min.
• LLOQ of icariin, naringin and osthole were 1.06, 2.10 and 1.05 ng/ml, respectively.
• Pharmacokinetic parameters of the analytes in male and female rats were compared.

A rapid, sensitive and selective ultra performance liquid chromatography tandem mass spectrometry (UPLC–MS/MS) method was developed and validated for the simultaneous determination of icariin, naringin and osthole in rat plasma. Plasma samples pretreatment involved a one-step liquid–liquid extraction with a mixture of ethyl acetate-methyl tert-butyl ether (3:1, ν/ν). The separation was performed on an ACQUITY UPLC™ BEH C18 column with a gradient mobile phase system of methanol and water. The detection was performed on a triple quadrupole tandem mass spectrometer equipped with electrospray ionization (ESI) by multiple reactions monitoring (MRM), with the transitions at m/z 513.3 → 366.8 (icariin), m/z 579.3 → 150.9 (naringin), m/z 245.1 → 189.0 (osthole) and m/z 237.1 → 194.1 (IS), respectively. A good linear response was observed over the concentration ranges of 1.06–424 ng/ml, 2.10–525 ng/ml and 1.05–1.05 × 103 ng/ml with lower limit of quantification (LLOQ) of 1.06, 2.10 and 1.05 ng/ml for icariin, naringin and osthole, respectively. The intra- and inter-day precisions (R.S.D.) were within 14.3%, and the accuracy (R.E.) ranged from −4.1% to 4.6% at three quality control levels. The sensitive and selective method was applied to a pharmacokinetic study of icarrin, naringin and osthole in rats after oral administration of Gushudan capsule.