Abstract
An analytical methodology appropriate for the determination of the novel drug candidate salicylaldehyde isonicotinoyl hydrazone (SIH) in rabbit plasma has been developed and validated. Desirable chromatographic separation was achieved on a C~18~ column employing a mixture of phosphate buffer (0.01 M NaH~2~PO~4~·2 H~2~O with 2 mM EDTA, pH 6.0) and methanol (53:47; v/v) as the mobile phase. In order to develop a suitable sample preparation procedure, different methods have been tested (solid‐phase extraction, liquid‐liquid extraction, and protein precipitation). Protein precipitation using 0.1 M HClO~4~ and acetonitrile allowed the highest recoveries of the analyte to be reproducibly attained. The analytical methodology developed in this study was validated with respect to linearity (0.26–30.0 μg/mL), accuracy, precision, selectivity, recovery, and stability. A concentration of 0.26 μg/mL was determined as the LLOQ. The chromatographic method was applied to a preliminary plasma pharmacokinetic study. This study has provided the first information about the concentrations of SIH in plasma of a living subject. These results could have a significant impact on further progress in the development of this promising compound.