Uhplc supergradient

The mobile phase was prepared with formic acid (>96%, Sigma-Aldrich, St. Louis, MO, USA), methanol (UHPLC-Supergradient, Panreac, Barcelona, Spain) and water (Elix3, Millipore, Bedford, MA, USA). Polyphenols of analytical quality were purchased from Sigma-Aldrich (St. Louis, MO, USA): gallic, homogentisic, protocatechuic, caftaric, gentisic, vanillic, caffeic, syringic, ferulic, p- coumaric acids, (+)-catechin, (−)-epicatechin, ethyl gallate, resveratrol, rutin, myricetin and quercetin. These compounds were selected according to their abundance and relevance in white and rosé cavas. Stock standard solutions of 1000 mg L⁻¹ of each polyphenol were prepared in MeOH. Working standard solutions consisting of a mixture of analytes at concentrations ranging from 20 to 0.05 mg L⁻¹ were prepared in MeOH:water (1:1, v:v).

The chemicals used in the extraction process, the spectrophotometric assays, and the HPLC method, were methanol and acetonitrile (99.9%, UHPLC Supergradient, PanReac, Barcelona, Spain), formic acid (≥95%, Sigma-Aldrich, St. Louis, MO, USA), hydrochloric acid (37%, PanReac, Barcelona, Spain), Fe (III) chloride and sodium carbonate (Merck, Darmstadt, Germany), FC reagent (PanReac, Barcelona, Spain), and 2,4,6-tripyridyl-S-triazine (TPTZ) (Alfa Aesar, Kandel, Germany). Purified water was generated with an Elix 3 coupled to a Milli-Q system (Bedford, MA, USA). The extraction solvent consisted of MeOH/H₂O/HCl (70:29:1, v:v:v).
The standards used for spectrophotometric assays were: gallic acid (97.5%) purchased from Sigma Aldrich (St. Louis, MO, USA) and 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox) purchased from Carbosynth (Berkshire, UK). Solutions of the standards were prepared at 1000 mg L⁻¹ (stored in amber vials at 4 °C) using dimethyl sulfoxide (DMSO) as the solvent. The solutions for the calibration were prepared with water to obtain concentrations in the range of 1 to 18 mg L⁻¹ gallic acid and 0.2 to 9 mg L⁻¹ Trolox, for FC and FRAP, respectively.