2.4.2. Analysis of Phenolic Compounds

The dried extracts (10 mg) were dissolved in a 80:20 (v/v) ethanol:water mixture (2 mL) and filtered through 0.22-μm disposable syringe filters. The analysis was performed in a HPLC-DAD-ESI/MSn system (Dionex Ultimate 3000 UPLC, Thermo Scientific, San Jose, CA, USA), as previously described [27]. Chromatographic separation was made in a Waters Spherisorb S3 ODS-2 C18 column (3 μm, 4.6 mm × 150 mm; Waters, Milford, MA, USA). Double online detection was carried out with a diode array detector (DAD, using 280 and 370 nm as preferred wavelengths) and a Linear Ion

Trap (LTQ XL) mass spectrometer (MS, Thermo Finnigan, San Jose, CA, USA) equipped with an electrospray ionization (ESI) source. Phenolic compounds were identified by comparison of their retention times and UV-vis and mass spectra with those obtained from standard compounds, when available; otherwise, compounds were tentatively identified comparing the obtained information with available data reported in the literature. For quantitative analysis, a calibration curve for each available phenolic compound standard (aloin A (280 nm: *y* = 3859.4*x* + 21,770, *R*<sup>2</sup> = 0.9996; and 370 nm: *y* = 7184.4*x* + 17,013, *R*<sup>2</sup> = 0.9996); chlorogenic acid (*y* = 168,823*x* – 161,172, *R*<sup>2</sup> = 0.9999); *p*-coumaric acid (*y* = 301,950*x* + 6966.7, *R*<sup>2</sup> = 0.9999); apigenin-6-glucoside (*y* = 107,025*x* + 61531, *R*<sup>2</sup> = 0.9989); apigenin-7-glucoside (*y* = 10,683*x* – 45,794, *R*<sup>2</sup> = 0.9906); and luteolin-6-C-glucoside (*y* = 4087.1*x* + 72,589, *R*<sup>2</sup> = 0.9988)) was constructed based on the UV signal. Quantification of the phenolic compounds that are not commercially available as standards was performed by using the most similar available standard molecule. The results were expressed as mg per g of extract.
