The samples were then centrifuged (Heraeus Biofuge 13, Sepa-tech) at 10,000 g for 15 min. The resulting pellets were again re-suspended in each of the extraction solvents and water, sonicated, vortexed, shaken and centrifuged as before as a wash-step. The remaining pellets were discarded and the final supernatants were pooled and analysed by LC–MS/MS.