Determination of rifalazil in dog plasma by liquid-liquid extraction and LC-MS/MS: Quality assessment by incurred sample analysis

Liquid-liquid extraction was used for isolation of rifalazil and an isotope labeled internal standard from plasma. A manual procedure, based on an 8 × 12 array format, was used for sample extraction. Extracts were analyzed by reversed-phase liquid chromatography using octylsilica column with gradient elution (1 mM ammonium acetate + 0.01% (v/v) acetic aid in water and methanol). Mass spectrometric detection was made with positive ion electrospray ionization and LC-MS/MS analysis in a triple-quadrupole mass spectrometer. The lower limit of quantification was 50 pg/ml. MS characteristics of rifalazil are presented. In particular, two different sets of ionization and selected reaction monitoring (SRM) conditions, where in-source fragmentation was used for precursor ion formation in one of the sets, were compared. The good correlation found between the two sets of results for authentic sample extracts indicated that either condition could be used for quantification of rifalazil in dog plasma.