4.8. Determination of Phenols by HPLC

Samples were prepared as reported by Akrimi et al. [53]. Notably, each extract (2 mL) was filtered with a syringe PTFE filter (0.45 μm) (Albet, Barcelona, Murcia, Spain) and injected into the HPLC system (Thermo Finnigan Surveyor, Waltham, MA, USA), equipped with a UV–VIS detector. The column was a Kinetex^® C18 EVO (250 × 4.6 mm, 5 µm) (Phenomenex, Torrance, CA, USA), fitted with a guard cartridge packed with the same stationary phase. Solvents and HPLC programs were those described by Rizzo et al. [54]. Standards used in the experiment were gallic, syringic and chlorogenic acid in addition to quercetin. They were injected at different concentrations from 10 to 1000 mg kg⁻¹ to obtain suitable calibration curves.

Chromatograms were recorded at 280 and 320 nm; all samples were assayed in triplicate.