2.11. Individual Polyphenol Analysis

Analysis of phenolic compounds was conducted using a Hitachi LaChroma Ultra HPLC system coupled with a photodiode array detector (Hitachi, Tokyo, Japan), according to Abountiolas et al. [31], with minor adjustments regarding retention times and dry weight conversions. Samples were injected at 40 °C into a reverse-phase Hypersil Gold C18 column (100 × 2.1 mm; particle size, 1.9 µm) (Thermo Fisher Scientific Inc., Waltham, MA, USA). The two mobile phases consisted of acidified water containing 0.5% formic acid (mobile phase A) and 0.1% formic acid in acetonitrile (mobile phase B) in an isocratic mixture. The flow rate was 0.3 µL/min, the wavelength detection included 250, 280, 360, and 520 nm, and the sample injection volume was 10 µL. Retention times and spectra were compared with pure standards of 17 compounds (at concentrations of 0.005 mg/mL, 0.01 mg/mL, and 0.1 mg/mL) from different polyphenol classes: flavonoids (cyanidin, pelargonidin, cyanidin 3-glucoside, pelargonidin 3-glucoside, quercetin, kaempferol, quercetin 3-glucoside, kaempferol 3-glucoside, myricetin, catechin, and epicatechin), phenolic acids (p-coumaric acid, ferulic acid, caffeic acid, gallic acid, and chlorogenic acid), and hydrolyzable tannins (ellagic acid). The flavonoids can be further classified into anthocyanidins (cyanidin and pelargonidin), anthocyanins (cyanidin 3-glucoside, and pelargonidin 3-glucoside), flavonols (quercetin, kaempferol, quercetin 3-glucoside, kaempferol 3-glucoside, and myricetin), and flavanols (catechin and epicatechin).