Q exactive plus combined quadrupole orbitrap mass spectrometer

Rapid separation of DISS metabolites by UHPLC was performed on a Thermo Vanquish Flex Binary RSLC platform (Thermo Fisher Scientific, Waltham, MA, USA). Samples were separated on a Thermo Accucore aQ C18 (150 × 2.1 mm, 2.6 µm; Thermo Fisher Scientific, Waltham, MA, USA) maintained at 40 °C. The mobile phase consisted of 0.1 formic acid water (A) and methanol (B). Gradient elution was performed as follows: 0–3 min, 0–20% B; 3–8 min, 20–35% B; 8–14 min, 35–70% B; 14–18 min, 70–75% B; 18–21 min, 75–95% B; 21–28 min, 95–97% B; 28–30 min, 97–100% B and 30–45 min, 100% B. The flow rate was 0.3 mL/min and the injection volume was 3 µL.
Analysis of the metabolites was performed using a Q Exactive Plus combined quadrupole Orbitrap mass spectrometer (Thermo Fisher Scientific, Waltham, MA, USA). Analysis was performed using electron spray ionization in both positive and negative ion modes, and the specific parameters were set as follows: capillary temperature, 350 °C; spray voltage, 4 kV/3.5 kV (positive/negative); sheath gas, 50 arb; auxiliary gas, 10 arb; and probe heater temperature, 320 °C. Full-scan analysis, from m/z 150–1500 with a resolution of 70,000, was performed under positive and negative ion modes. Collision-induced dissociation was performed using an isolation width of 2 Da, and 30% maximum of collision energy.