2.3.2. Antioxidant Activity Evaluation

The DPPH (2,2-diphenyl-1-picrylhydrazyl) assay was performed on the extracts according to the procedure of Tarantino et al. [44]. Each extract (50 μL) was combined with 950 μL DPPH solution (0.08 mM in ethanol). The decrease in absorbance was read at 517 nm using a Cary 60 UV-Vis spectrophotometer (Agilent, Cernusco, Milan, Italy). The results were expressed in μmol Trolox equivalents g<sup>−</sup><sup>1</sup> dry weight for all vine shoot samples (μmol TE g<sup>−</sup><sup>1</sup> DW). All determinations were carried out in duplicate. Antioxidant activity was also determined by ABTS-TEAC assay [44]. For spectrophotometry, the reaction took place directly in cuvettes by adding 50 μL of each sample to 950 μL of final ABTS•<sup>+</sup> solution. After 8 min, the decrease in absorbance was measured at 734 nm, using a Cary 60 UV-Vis spectrophotometer (Agilent, Cernusco, Milan, Italy). The results were expressed in μmol TE g<sup>−</sup><sup>1</sup> dry weight for all vine shoot samples (μmol TE g<sup>−</sup><sup>1</sup> DW). Each sample was analysed in duplicate.

### 2.3.3. Quantification of Rsv and Vf by HPLC-DAD

The analysis of the stilbenes was performed according to the method of Ewald et al. [38] using high-performance liquid chromatography (UltiMate 3000 HPLC, Thermo scientific, Munich, Germany) that included an HPG-3200RS binary pump, WPS-3000RS/TRS autosampler, TCC-3000RS column oven, and a DAD-3000RS photodiode array detector. HPLC separation was achieved on AcclaimTM 120 C18 columns (120 A 3 ˚ × 150 mm, 3 μm) maintained at 25 ◦C using a mobile phase consisting of 1% aqueous acetic acid (*v*/*v*) (A) and methanol (B). The separation was carried out at 25 ◦C with a flow rate of 0.6 mL min−<sup>1</sup> under the following conditions: 0 min (20% B), 10 min (20% B) 6.5 min (37% B), 12.6 min (50% B), and 21.0 min (100% B). Under these conditions, Rsv and Vf were eluted with a retention time of 14.7 min and 17.8 min and monitored at 306 and 324 nm, respectively. Calibration curves were prepared using the endotoxin standards (Sigma-Aldrich, Steinheim, Germany) of Rsv (R<sup>2</sup> = 0.9993) and Vf (R<sup>2</sup> = 0.9994) in the concentration range 1–500 mg L−1. The amount of Rsv and Vf found in each extract was expressed as mg of compound kg−<sup>1</sup> of DW. Each sample was analysed in duplicate.
