Development and validation of a rapid and sensitive assay for the determination of anisodamine in 50 µL of beagle dog plasma by LC-MS/MS

A simple, rapid, high-throughput, and highly sensitive LC-MS/MS was developed to determine anisodamine in a small volume (50 µL) of beagle dog plasma using atropine sulfate as the internal standard. The analyte and internal standard were isolated from 50 µL plasma samples after a one-step protein precipitation using Sirocco 96-well protein precipitation filtration plates. The separation was accomplished on a Hanbon Hedera CN column (100 × 4.6 mm, 5 µm) and the run time was 4 min. A Micromass Quatro Ultima mass spectrometer equipped with an ESI source was operated in the multiple reaction monitoring mode with the precursor-to-product ion transitions m/z 306.0[arrow right]140.0 (anisodamine) and 290.0[arrow right]123.9 (atropine) used for quantitation. The method was sensitive with a low LOQ of 0.05 ng/mL, and good linearity in the range 0.05-50 ng/mL for anisodamine (r2 ≥ 0.995). All the validation data, such as accuracy, intra- and interrun precision, were within the required limits. The method was successfully applied to the pharmacokinetic study of anisodamine hydrochloride injection in beagle dogs. [PUBLICATION ABSTRACT]