Liquid Chromatography Tandem Mass Spectrometric Analytical Method for Study of Colchicine in Rats Given Low Doses

Abstract

A selective and sensitive assay was developed for colchicine in rat specimens. Colchicine and its deuterated analog (as internal standard, IS) were extracted from rat specimens (minimal 0.1 mL plasma, whole blood, or urine) using liquid-liquid extraction with n-hexane:dichloromethane:isopropanol. The mobile phase (formic acid: ammonium acetate: methanol) was pumped with uniform flow through an octadecylsilane analytical column. Detection was carried out by electrospray positive ionization in the multiple-reaction monitoring mode. The assay (total run time <3 min) had excellent linearity over a wide (400–800-fold) concentration range. The mean absolute recovery was >96.8%. The intra- and inter-day coefficients of variation were ˂15%, with lower limits of quantitation of 0.5 ng/mL in 0.1 mL of rat plasma. The method also provided the same lower limits of quantitation in urine and whole blood with 0.1 mL volumes, and 0.1 ng/mL using 0.5 mL of rat plasma. The blood-to-plasma ratio was >1. Rats had measurable colchicine blood concentrations for at least 24 h after intravenous doses of 0.1 mg/kg. The method possessed suitable measures of sensitivity and selectivity for detecting colchicine in several specimen types in rats given low doses.