Objective: To establish a rapid and sensitive HPLC-MS /MS method for the determination of the concentration of gestrinone in Bama minipig plasma, and to study the pharmacokinetic characteristics of gestrinone Capsules in Bama minipigs. Methods: Bama minipigs were used as experimental animals for 12 weeks and twice a week, the blood samples were collected at different time points after the administration. The pharmacokinetic parameters were calculated by measuring the concentration of gestrinone in plasma of minipigs after multiple administrations. The plasma samples were extracted by cyclohexane and analyzed by LC-MS / MS. Gestodene was used as the internal standard(IS). Column: Agilent ZORBAX Extend-C18(2.1 mm×50 mm, 5 μm);mobile phase: water(containing 0.1% formic acid)-acetonitrile(containing 0.1% formic acid), gradient elution;flow rate: 0.40 mL·min-1, Amount: 20 μL;column temperature: 35 ℃. Mass spectrometry was performed using ESI positive ion mode, scanning mode was multiplex reaction monitoring(MRM)mode, and ion reaction pairs were selected to be m/z 309.2-241.1(gestrinone), and m/z 311.2-109.1(gestodene). Results: The concentration of gestrinone in the range of 0.1-10 ng·mL-1 was linear(r=0.999 9)and the lower limit of quantification was 0.1 ng·mL-1. The recovery rate was higher than 83.1%, the intra-day and intra-day RSD were less than 11.3%, the specificity was good, and the samples were stable under the conditions involved in this experiment. The first administration pharmacokinetics parameters for gestrinone capsule in 8 minipigs were as followed: Cmax(2.85±0.96)μg·L-1, Tmax(2.06±0.62)h, t1/2(8.43±4.13)h, AUC(0-85)(23.81±8.62)μg·L·h-1, AUC(0-∞)(24.36±8.73)μg·L·h-1, MRT(0-85)(8.40±2.44)h, MRT(0-∞)(9.81±3.88)h;the last administration parameters were as followes: Cmax(2.70±0.55)μg·L-1, Tmax(1.69±0.65)h, t1/2(8.51±3.72)h, AUC(0-85)(17.71±4.89)μg·L·h-1, AUC(0-∞)(18.51±4.48)μg·L·h-1, MRT(0-85)(8.54±3.50)h, MRT(0-∞)(9.98±4.41)h. Conclusion: The method is accurate, rapid, sensitive and specific, and is suitable for animal experiment and clinical determination of plasma concentration of gestrinone and its pharmacokinetic study. After repeated administration of gestrinone capsules, there was no difference in 2 h plasmg concentration, and no adverse reaction after treatment, and the safety was higher.
