Quantitation and metabolism of mitoquinone, a mitochondria-targeted antioxidant, in rat by liquid chromatography/tandem mass spectrometry

Mitoquinone (MitoQ10 mesylate) is a mitochondria-targeted antioxidant undergoing development for the treatment of neurodegenerative diseases. The aim of this study was to develop and validate an assay based on liquid chromatography/tandem mass spectrometry (LC/MS/MS) to determine mitoquinone and to detect and identify the metabolites of MitoQ10 in rat plasma after an oral dose. After a simple protein precipitation step, plasma samples were analyzed by reversed-phase liquid chromatography using gradient elution with acetonitrile/water/formic acid. Electrospray ionization in the positive ion mode with multiple reaction monitoring (MRM) was used to analyze mitoquinone employing the deuterated compound (d3-MitoQ10 mesylate) as internal standard. The calibration curve for mitoquinone was linear over the concentration range 0.5-250 ng/mL with a correlation coefficient>0.995. The method was sensitive (limit of quantitation 0.5 ng/mL) and had acceptable accuracy (relative error<8.7%) and precision (intra- and inter-day coefficient of variation<12.4%). Recoveries of mitoquinone at concentrations of 1.5, 20 and 200 ng/mL were in the range 87-114%. The method was successfully applied to a pharmacokinetic study in rat after a single oral dose in which four metabolites of MitoQ10 were tentatively identified as hydroxylated MitoQ10, desmethyl MitoQ10 and the glucuronide and sulfate conjugates of the quinol form of MitoQ10.