Agilent UPLC 1290II system combined with a 6540 QTOF (Agilent Technologies, Santa Clara, CA, USA) was used to determine the accurate mass of the metabolites. UPLC equipped with a binary solvent delivery system, autosampler, and column compartment was used in this study. Chromatographic separation was performed on a Waters BEH C18 column (2.1 × 100 mm, 1.7 μm), and the elution conditions were as follows: 0–15 min, 5%–95% B. A and B indicate 0.1% formic acid water (formic acid:water, 0.1:100, v/v) and acetonitrile, respectively.
Sample ionization was acquired in both positive and negative modes within the mass/charge (m/z) range of 50–1000. The electrospray ionization (ESI) source operating parameters in both the positive and negative modes and the ESI–MS conditions were as follows: gas temperature, 325 °C; gas flow, 5 L/min; nebulizer, 35 psig; and sheath gas temperature, 350 °C. Internal references (purine and HP-0921) were adopted to modify the measured masses in real time, and the reference masses were m/z 121.0509 and 922.0098 in the positive-ion mode and 119.0363 and 1033.9881 in the negative-ion mode. The accurate mass of each metabolite was used for quantification.
Sample ionization was acquired in both positive and negative modes within the mass/charge (m/z) range of 50–1000. The electrospray ionization (ESI) source operating parameters in both the positive and negative modes and the ESI–MS conditions were as follows: gas temperature, 325 °C; gas flow, 5 L/min; nebulizer, 35 psig; and sheath gas temperature, 350 °C. Internal references (purine and HP-0921) were adopted to modify the measured masses in real time, and the reference masses were m/z 121.0509 and 922.0098 in the positive-ion mode and 119.0363 and 1033.9881 in the negative-ion mode. The accurate mass of each metabolite was used for quantification.