A rapid and sensitive UPLC-MS/MS method for the determination of zanubrutinib in beagle plasma and its application in pharmacokinetics.

A reliable and sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed for the determination of zanubrutinib in the plasma of beagle dogs. The column used was an Acquity BEH C18 column (2.1 mm × 50 mm, 1.7 µm), maintained at 40°C with an injection volume of 2 µl. The gradient elution program was as follows: 0-1 min, 10-10% A; 1-1.1 min, 10-90% A; 1.1-2.1 min, 90-90% A; 2.1-2.2 min, 90-10% A; 2.2-3.0 min, 10-10% A. Mobile phase A was 0.1% formic acid, B was acetonitrile, and the total analysis time was 3 min. The mass spectrometry was performed in positive ion mode, and the scanning mode was multi-reaction monitoring mode with electrospray ionization as the ion source; m/z 472.2 → 455.01 for zanubrutinib and m/z 441.03 → 137.99 for ibrutinib (internal standard). The plasma samples were processed by protein precipitation. The standard curve showed good linearity (r2 = 0.999 8) in the range of 1.0-1,000 ng/ml (zanubrutinib) with a low limit of quantification of 1 ng/ml. Also, the intra-day and inter-day precision (RSD) was <5.88% and the accuracy (RE) ranged from -1.56 to 1.08%; the recoveries of zanubrutinib in beagle plasma ranged from 90.12 to 93.53% (RSD 1.67-6.42%) and the ME values of zanubrutinib were 98.70-101.06% (RSD 5.37-8.49%, n = 6). All values meet US Food and Drug Administration requirements. A rapid, highly selective and sensitive method for the determination of zanubrutinib concentration in plasma by UPLC-MS/MS was successfully developed. This method is suitable for pharmacokinetic studies in beagle dogs by following oral administration of zanubrutinib.

The Pharmacokinetic Effect of Itraconazole and Voriconazole on Ripretinib in Beagle Dogs by UPLC-MS/MS Technique.

BACKGROUND: A new UPLC-MS/MS technique for the determination of ripretinib in beagle dog plasma was developed, and the pharmacokinetic effects of voriconazole and itraconazole on ripretinib in beagle dogs were studied. METHODS: After extraction with ethyl acetate under alkaline conditions, ripretinib was detected using avapritinib as the internal standard (IS). The mobile phases were 0.1% formic acid-acetonitrile. The scanning method was multi-reaction monitoring using ESI+ source, and the ion pairs for ripretinib and IS were m/z 509.93→416.85 and 499.1→482.09, respectively. This animal experiment adopted a three period self-control experimental design. In the first period, ripretinib was orally administered to six beagle dogs at a dose of 5 mg/kg. In the second period, the same six beagle dogs were orally given itraconazole at a dose of 7 mg/kg, after 30 min, ripretinib was orally given. In the third period, voriconazole at a dose of 7 mg/kg was given orally, and then ripretinib was orally given. At different time points, the blood samples were collected. The concentration of ripretinib was detected, and the pharmacokinetic parameters of ripretinib were calculated. RESULTS: Ripretinib had a good linear relationship in the range of 1-1000 ng/mL. The precision, accuracy, recovery, matrix effect and stability met the requirements of the guiding principles. After erdafitinib combined with itraconazole, the Cmax and AUC0→t of ripretinib increased by 38.35% and 36.36%, respectively, and the t1/2 was prolonged to 7.53 h. After ripretinib combined with voriconazole, the Cmax and AUC0→t of ripretinib increased by 37.44% and 25.52%, respectively, and the t1/2 was prolonged to 7.33 h. CONCLUSION: A new and reliable UPLC-MS/MS technique was fully optimized and developed to detect the concentration of ripretinib in beagle dog plasma. Itraconazole and voriconazole could inhibit the metabolism of ripretinib in beagle dogs and increase the plasma exposure of ripretinib.