2.2. Chromatographic Separation of Phenolic Compounds

An Agilent 1200 HPLC system (Agilent Technologies, Waldbronn, Germany) equipped with a vacuum degasser, autosampler, a binary pump, and a DAD detector was used for the chromatographic determination. Phenolic compounds were separated by using an Agilent Eclipse Plus C18-column (4.6 × 150 mm, 1.8 mm particle size) operating at 25°C and a flow rate of 0.8 mL/min. The mobile phases used were water with acetic acid (0.5%) (phase A) and acetonitrile (phase B), and the solvent gradient changed according to the following conditions: 0 to 10 min, 5–30% B; 10 to 12 min, 30–33% B; 12 to 17 min, 33–38% B; 17 to 20 min, 38–50% B; and 20 to 23 min, 50–95% B. Finally, the B content was decreased to the initial conditions (5%) in 2 min, and the column reequilibrated for 10 min. A volume of 10 mL of the extracts was injected. The separated compounds were monitored in sequence first with a diode-array detector (DAD) at 240 and 280 nm and then with a mass spectrometry (MS) detector.