Separation of steroid isomers by ion mobility mass spectrometry

•We report a reliable, fast and repeatable method to separate steroid isomers.•We compare two different drift gases in order to achieve optimum separation.•Different drift gas pressures were studied for optimum separation.•The separation efficiencies of the steroid isomers are increased by derivatization.

Ion mobility mass spectrometry performed in a compact traveling wave cell (TWIM-MS) is shown to provide a reliable, fast and repeatable method to separate derivatized steroid isomers. Three steroid isomer pairs were analyzed in their native form and as their p-toluenesulfonyl isocyanate derivatives. The native steroids were separated from each other, but no separation could be attained for the isomers. The derivatized steroid isomers were, however, properly separated by TWIM-MS with peak-to-peak resolutions close to or as high as baseline resolution (Rp − p = 0.77–1.08).