6520 accurate electrospray ionization quadrupole time of flight mass system

Each sample was processed by Agilent 1200 high performance liquid chromatography combined with a 6520 accurate electrospray ionization/quadrupole-time-of-flight mass system (Agilent Technologies, CA, USA). Plasma samples were separated on an Eclipse Plus C18 column (2.1 x 50 mm, 1.8 μm, Agilent Technologies), with the condition of 150 μL injection volume, 0.25 mL/min flow rate and 37°C column temperature, by using a gradient program of the mobile phase A 0.1% formic acid solution (ESI+)/water (ESI-) and mobile phase B was acetonitrile with 0.1% formic acid solution (ESI+)/acetonitrile (Merck, Darmstadt, Germany) (ESI-). The gradient program started from 2% B for 0–9 minutes, linearly increased from 2 to 60% B for 9–18 minutes, linearly increased from 60 to 100% B for 18–20 minutes, and kept at 100% B for 20–30 minutes. To avoid cross contamination from plasma samples, plusing an ultra-pure water elution after each sample. All the data were collected in ionization quadrupole time-of-flight mass spectrometry with both positive (ESI+) and negative (ESI-) full scan mode. The parameters of mass spectrometry were as following: GAS1, 80; GAS2, 70; CUR, 40; TEM, 375°C; and ISVF, 5500 V; a mass range of 50 to 1500; scan time of 0.3 seconds.