Dionex dx 600 hplc

The concentration of organic acids (citric, tartaric, malic, succinic and lactic) was determined as described previously [22 (link)] using an Aminex HPX-87H Bio-Rad 7.8 × 300 mm column (Bio-Rad, Hercules, CA, USA) fitted on a Dionex DX-600 HPLC equipped with a DAD detector (Thermo Scientific, Waltham, MA, USA).

The determination of phenolic compounds was conducted, as previously reported [21 (link)], using a Dionex DX-600 HPLC equipped with a DAD detector (Thermo Scientific, Waltham, MA, USA). All samples were degassed and sterile-filtered (0.22 μm) prior to injection. The separation of benzoic acids, hydroxycinnamic acids and flavonoids was performed by using an Inersustain C18 column (5 μm, 4.6 × 250 mm, CPS Analitica, Milano, Italy) as the stationary phase. The different compounds were eluted by applying a binary gradient (buffer A: 5% CH₃COOH in 5% water, v/v; buffer B: 80% acetonitrile: 20% water, v/v). The injection volume was 10 μL. The identification and quantification of different phenolic compounds was performed by using different standard calibration curves with protocatechuic acid, vanillic acid, hydroxybenzoic acid, gallic acid, syringic acid, (+)-catechin, (−)-epicatechin, p-coumaric acid, caffeic acid, chlorogenic acid, ferulic acid, rutin and quercetin. All standards were sourced from Sigma-Aldrich (St. Louis, MO, USA). The compounds for which a standard was unavailable (caftaric, sinapic, coutaric and ferulic acids) were quantified as coumaric (CUE) or caffeic (CAE) acid equivalents.