2.4. Phenolic Compounds Analysis by LC-MS

The analysis of phenolic compounds was carried out following a procedure previously described by the authors [43] with some modifications. Briefly, the phenolic extract was analyzed by liquid chromatography–mass spectrometry (LC-MS) (Thermo Scientific, Waltham, MA, USA). Chromatographic separation was accomplished using an Acclaim™ 120 (Thermo Scientific, Waltham, MA, USA) reverse phase C18 columns (3 µm 150 × 4.6 mm) thermostatted at 35 °C, and peaks were detected at 280 nm as the preferred wavelength. The mobile phase used was composed of 1% acetic acid in water and 100% acetonitrile. The elution gradient established was from 10% to 15% B over 5 min, from 15% to 25% B over 5 min, from 25% to 35% B over 10 min, from 35% to 50% B over 10 min, isocratic 50% B for 10 min, and re-equilibration of the column, using a flow rate of 0.5 mL/min. The identification of phenolic compounds in the samples was characterized according to their UV-Vis spectra and identified by their mass spectra and retention times in comparison with commercial standards. Quantification was made from the areas of the peaks recorded at 280 nm by comparison with calibration curves obtained from the standard of each compound. The results were expressed in µg per gram of dw.