G4220a quaternary pump

Experiments were carried out on an Agilent 1290 series high-performance liquid chromatography, including a G4220A quaternary pump, G4212A DAD detector and a G1316C column heater and a G4226A autosampler, a tandem Agilent 6460A triple-quadrupole mass spectrometer equipped with an electrospray ionization source (Agilent Inc., MA, United States) and operated with MassHunter Version B.06.00 workstation software. The separation was performed on an Agilent Zorbax SB-C18 column (3.5 μm, 2.1*100 mm) with the column temperature maintained at 55°C and the sample injection volume was 10 μl. The mobile phase consisted of a mixture of phase A (10 mM ammonium acetate buffer containing 0.1% formic acid) and phase B (methanol with 10 mM ammonium acetate buffer and 0.1% formic acid) with the flow rate at 0.25 mL min⁻¹. The gradient program was set as follows: 0 min, 80% B; 0–2 min, 80–100% B; 2–10 min, 100% B.
Quantification was performed using electrospray in the positive mode with the spray voltage set at 4,000 V. Nitrogen was used as nebulizer gas and nebulizer pressure was set at 20 psi. Desolvation gas (nitrogen) was heated to 325°C and delivered at a flow rate of 12 L/min.

An Agilent 1290 liquid chromatographic system includes a G4220A quaternary pump, a G1326C column incubator, a G4226A automatic sampler, and a G1330B degasser. The column temperature was set to 35°C. A measure of 5 µL of samples were separated in an Agilent SB-C18 column (2.1 mm ×50 mm, 1.8 μm) and eluted at 0.3 ml/min, followed by gradient elution with water (0.1% V/V formic acid) (A) and acetonitrile (B) (0–1 min, 20%–20% B; 1–3.5 min, 20%–80% B; 4–4.1 min, 80%-20% B; 4.1–6 min, 20% B). The total run time was 6 minutes.