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

Abstract

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 μg 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 μg mL−1 and 0.03 μg mL−1, respectively) and quantification (i.e., 0.20 μg mL−1 and 0.08 μg 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.