Liquid chromatography-mass spectrometry for simultaneous determination of spironolactone and canrenone in plasma samples

In our study, we aimed to validate a method based on liquid chromatography-mass spectrometry (LC-MS) to quantify spironolactone (SPI) and its active metabolite canrenone (CAN) simultaneously in plasma samples to support in vivo experiments. Compounds were separated by using a C18 column with the isocratic elution of a mobile phase composed of 0.1% (v/v) formic acid in methanol-water (60:40 v/v) at a flow rate of 0.4 mL min-1. SPI and CAN were detected in an electrospray interface operating in a positive ionization mode and quantified using the selective ion mode monitoring of mass-charge ratios (m/z) of 439.0 for SPI and 363.1 for CAN. After calculating the matrix effect using theoretical equations, we observed the strong interference of plasma in the equipment-generated signal, which required creating analytical curves using the matrix as a solvent. The method was nevertheless linear (r2 > 0.999) in a concentration range of 0.4-5.0 mu g mL-1, as well as precise, with a coefficient of variation less than 5%. SPI's and CAN's recovery rates from the plasma ranged from 87.4% to 112.1%, while their limits of detection (i.e., 0.07 mu g mL-1 and 0.03 mu g mL-1, respectively) and quantification (i.e., 0.20 mu g mL-1 and 0.08 mu g mL-1, respectively) in the presence of plasma contaminants were low. Therefore, the bioanalytical method seems to be feasible for quantifying SPI and CAN in plasma.