Development of an LC‐MS/MS method for determination of bicalutamide on dried blood spots: application to pharmacokinetic study in mice

A rapid and highly sensitive assay method has been developed and validated for the estimation of bicalutamide (BCL) on mouse dried blood spots (DBS) using liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the negative-ion mode. The assay procedure involves a simple liquid extraction of BCL and tolbutamide (internal standard, IS) from mouse blood DBS cards using tert-butyl methyl ether. Chromatographic separation was achieved with 5 mm ammonium acetate (pH 6.5)-acetonitrile (35:65, v/v) at a flow rate of 0.60 mL/min on an Atlantis dC18 column with a total run time 3.0 min. The MS/MS ion transitions monitored were 428.80 → 254.70 for BCL and 269.00 → 169.60 for IS. Method validation was performed as per regulatory guidelines. A linear response function was observed from 0.92 to 1911 ng/mL for BCL in mouse blood. The intra- and inter-day precisions were in the ranges of 1.86-12.5 and 3.19-10.8%, respectively. This novel DBS method has been applied to a pharmacokinetic study in mice.