Total phenols were determined according to
Singleton and Rossi (1965), with some modifications.
Sample of 50 μL were taken from a 2:8 dilution with
80% methanol) and 3 mL of HPLC-grade water and
250 μL of Folin-Ciocalteu 1N (1:1) reactive were
added. After 5 min 750 μL of 20% Na2CO3 was added,
followed by 950 μL of HPLC-grade water; shaken in a
vortex and kept in the dark for 30 minutes. Absorbance
was read using an UV-VIS VARIAN CARY 50 BIO
spectrophotometer, at a wavelength of 765 nm. Results
were expressed in mg of Gallic Acid Equivalents
(GAE)/100 g of Fresh Weight (FW). Analyses were
performed in triplicate per each RS.