2.6 Total Phenolic Content
Total phenolic content in extracts exhibiting the strongest antimicrobial activity was estimated using the
Folin-Ciocalteu colorimetric oxidation/reduction method described by Skerget (2005), with some modifications.
Aliquots (0.5 ml) of appropriate dilutions of extracts were oxidized for 4 min with 2.5 ml of Folin-Ciocalteu’s
reagent (Sigma-Aldrich, Germany) diluted 10 times with water, and the reaction was neutralized with 2 ml of
sodium carbonate (75 g l-1). The samples were incubated for 1h at 25 °C, and the absorbance of the resulting blue
colour was measured at 760 nm with a UV-vis mini - 1240 spectrophotometer (Shimadzu Corporation, Kyoto,
Japan). For a control sample, 0.5 ml of distilled water was used. A standard curve was generated using various
concentrations of gallic acid (0-200mgl-1), which resulted in an R2 value of 0.99, and results were expressed as
mg of gallic acid equivalent (GAE) per gram of extract. All values are presented as means of triplicate analyses
with duplicate samples analyzed per replicate.