Accucore c18 2

A Shimadzu LC (NEXERA X2) system coupled to a triple-quadrupole mass spectrometer was used (8060, Shimadzu, Kyoto, Japan). Chromatographic separation was carried out at 40 °C using an Accucore C18 150 mm × 2.1 mm × 2.6 μm column (Thermo Electron, France) with a pre-column (Accucore C18 2.1 mm × 2.6 μm). The methanol mobile phases were methanol + 2 mM ammonium acetate + 0.002% formic acid + 5 ml H₂O and H₂O + 2 mM ammonium acetate + 0.002% formic acid. The acetonitrile mobile phases were acetonitrile + 0.05% formic acid and H₂O + 0.05% formic acid. Injection volume was 5 μl, and the flow at 0.3 ml/min. ESI (electrospray ionization) was operated in a positive/negative polarity switching mode and data were acquired in multiple reaction monitoring mode (MRM). Interface voltage was at 1500 V, interface temperature, at 350 °C and CID Gas at 230 kPa.

A LC (Dionex Ultimate 3000) system coupled to a triple-quadrupole mass spectrometer was used (TSQ Quantiva, Thermo Scientific, France). Chromatographic separation was carried out at 40 °C using an Accucore C18 150 mm × 2.1 mm × 2.6 μm column (Thermo Electron, France) with a pre-column (Accucore C18 2.1 mm × 2.6 μm). The mobile phases were methanol + 0.5 mM ammonium acetate and H₂O + 0.5 mM ammonium acetate. Injection volume was 5 μl, and the flow at 0.3 ml/min. ESI was operated in a positive/negative polarity switching mode and data were acquired in MRM. The mass spectrometer parameters were: spray voltage of positive ion of 4000 V and negative of 2500 V; sheath gas of 40 (arbitrage); aux gas of 15 (arb); ion transfer tube temperature of 300 °C and vaporizer temperature of 300 °C.