Agilent 6470 triple quadrupole tandem mass spectrometer

An Agilent 1290 UHPLC system (Agilent Technologies Inc., Palo Alto, CA, USA) coupled to an Agilent 6470 triple quadrupole tandem mass spectrometer with electrospray ionization (ESI) source was used for the quantitative analysis. An ACQUITY UPLC BEH C18 column (2.1 ×100 mm, 1.7 μm; Waters, Milford, MA, USA) was used for chromatographic separation, and the column temperature was maintained at 25°C. The binary gradient elution system consisted of 0.1% formic acid (solvent A) and acetonitrile (solvent B). The solvent gradient was as follows: 0–1 min, 25–33% B; 1–5 min, 33–33% B; 5–7 min, 33–41% B; 7–9 min, 41–41% B; 9–10 min, 41–59% B; 10–15 min, 59–59% B. The flow rate was 0.3 mL/min and the injection volume was 5 μL.
The mass spectrometer was operated in negative mode. The optimized mass conditions were as follows: gas temperature, 300°C; gas flow rate, 7 L/min; nebulizer, 35 psi; sheath gas temperature, 250°C; sheath gas flow, 12 L/min; capillary voltage, 4,000 V. Multiple reaction monitoring (MRM) mode was applied for the quantitative analysis of different compounds. An MRM diagram is shown in Supplementary Figure S2. The optimal mass spectral parameters and ion patterns are presented in Supplementary Table S2.

Quantitative analysis was performed on an Agilent 1290 UHPLC system (Agilent Technologies Inc., Palo Alto, CA, USA) equipped with an Agilent 6470 Triple quadrupole tandem mass spectrometer (Agilent Technologies, Singapore) with electrospray ionization (ESI) source. A Waters ACQUITY UPLC^®® BEH C18 column (2.1 × 100 mm, 1.7 µm) was used for chromatographic separation, and the column temperature was maintained at 20 °C. The binary gradient elution system consisted of 0.05% formic acid in water (A) and acetonitrile (B). The gradient profile started from 10% B, increased linearly to 23% B within 2 min, and then increased to 50% B within 3 min.
The mass spectrometer was operated in positive mode. The optimized mass conditions were as follows: gas temperature, 320 °C; gas flow rate, 8 L/min; nebulizer, 35 psi; sheath gas temperature, 250 °C; sheath gas flow, 12 L/min; capillary voltage, 4000 V. Multiple reaction monitoring (MRM) mode was applied for the quantitative analysis of different compounds. An MRM diagram is shown in Supplementary Figure S4. The optimal mass spectral parameters and ion patterns are presented in Table 4.