2.7 LC-MS/MS analysis
The samples were analyzed in a 4°C autosampler using a SHIMADZU-LC30
ultra-high performance liquid chromatography (UHPLC) system with an
ACQUITY UPLC® HSS T3 (2.1×150 mm, 1.8 µm) (Waters, Milford, MA, USA)
column. The mobile phase A: 0.1% formic acid aqueous solution, B:
acetonitrile; the chromatographic gradient elution procedure was as
follows: 0 - 1.5 min, 0.3% B; 1.5 - 2 min, B linearly varied from 0.3%
to 0%; 2 - 6 min, B linearly varied from 0% to 48%; 6 - For 10 min, B
varied linearly from 48% to 10%; for 10-12 min, B was maintained at
100%; for 12-12.1 min, B varied linearly from 100% to 0%; for 12.1-15
min, B was maintained at 0%.
Each sample was detected in positive (+) and negative (-) ion modes by
electrospray ionization (ESI). The samples were separated by UPLC and
analyzed by mass spectrometry using a QE Plus mass spectrometer (Thermo
Scientific), and ionized using an HESI source with the following
ionization conditions: Spray Voltage: 3.8kv (+) and 3.2kv (-); Capillary
Temperature: 320 (±); Sheath Gas: 30 (±); Aux Gas: 5 (±); Probe Heater
Temp: 350 (±); S-Lens RF Level: 50.