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

OBJECTIVE
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.


METHODS
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 (MRM) mode with ESI as the ion source. m/z 472.2→455.01 for zanubrutinib and m/z 441.03→137.99 for ibrutinib (IS). The plasma samples were processed by protein precipitation.


RESULTS
The standard curve showed good linearity (r2 =0.999 8) in the range of 1.0-1000 ng/mL (zanubrutinib) with a low limit of quantification of 1 ng/mL. Also, the intra-day and inter-day precision (RSD %) was less than 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 FDA requirements.


CONCLUSION
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.