Quantifying Polyphenols in Cyanobacterial Extracts

The TPC of the cyanobacterial extracts was determined using the colorimetric assay of Folin–Ciocalteu, according to Barroso et al. [18 (link),48 (link)]. The acetonic extracts were solubilized in DMSO and the aqueous extracts in water. Briefly, a volume of 25 μL of each extract (10 mg mL⁻¹) was thoroughly mixed with 25 μL of Folin–Ciocalteu reagent (Sigma-Aldrich, St. Louis, MO, USA), 200 μL of Na₂CO₃ solution (75 g L⁻¹) and 500 μL of deionized water. After the incubation period (60 min at room temperature), the absorbance of the colored product was measured at 725 nm, using a Synergy HT Multi-detection microplate reader (Biotek, Bad Friedrichshall, Germany) operated by GEN5^TM software. Standard calibration curves (y = 2.097x + 0.01560, R² = 0.9989, for aqueous extracts and y = 2.204x + 0.01401, R² = 0.9982, for acetonic extracts) were obtained with seven concentrations of gallic acid (GA) (0.025 to 0.5 mg mL⁻¹). TPC in each extract was expressed in µg of gallic acid equivalents (GAE) per mg of dry biomass. Three independent determinations were carried out in triplicate.