1525 binary pump solvent management system

Analyses were primarily performed by using a Waters HPLC System (Waters Crop, Milford, MA, USA) equipped with a 1525 binary pump solvent management system, 2998 PDA detector, 2707 automatic sampling device, and Breeze 2 workstation. Two additional different HPLC instruments were used: One was high performance liquid chromatography (Waters Crop, Milford, MA, USA) equipped with 1525 binary pump solvent management system, 2489 UV detector, and Breeze 2 workstation. Another was a Waters Alliance e2695-2998 HPLC system (Empower workstation, Waters Crop, Milford, MA, USA). HPLC separation was carried out on a CAPCELL PAK C₁₈ column (4.6 mm × 150 mm, 5 μm). Column temperature was set at 25°C, and inject volume was 10 μL. The mobile phase consisted of methanol (A) and 0.3% formic acid aqueous solution (B). The gradient elution was programmed at a flow rate of 1.0 mL·min⁻¹ as follows: 0–12 min, 32% A; 12–40 min, 32%–50% A; 40–50 min, 50%–70% A; 50–52 min, 70% A. The detection wavelength was set at 254 nm.