Analysis of the intracellular localization of amiodarone using live single-cell mass spectrometry.

Amiodarone is a well-known antiarrhythmic drug with side effects including phospholipidosis. However, it is not clear how amiodarone and its metabolites are localized in the cell. In the present study, the localization of amiodarone in the cytosol, vacuoles, and lipid droplets of a single HepG2 human hepatocellular carcinoma cell was determined directly using live single-cell mass spectrometry. The cytosol, vacuoles, and lipid droplets of a single HepG2 cell treated with amiodarone were separately captured using a nano-spray tip under a fluorescence microscope after visualizing the lipid droplets using a fluorescent probe. This assay showed a linearity in the measurement of amiodarone levels with R2 values of 0.9996 and 0.9998 in the cell lysates and serum, respectively. The peak intensities of amiodarone and its metabolites in lipid droplets and vacuoles were significantly higher than those in the cytosol, while those in lipid droplets were higher than those in vacuoles. Amiodarone metabolites were detected in both lipid droplets and the cytosol. Live single-cell mass spectrometry combined with fluorescence imaging demonstrated clear localization of amiodarone and its metabolites in lipid droplets separately from the vacuole. This assay system combined with fluorescence imaging could be useful for investigating the intracellular localization of various drugs and their metabolites.