Total flavonoid content (TFC) was determined using colorimetric method described by Abu Bakar, Mohamed, Rahmat, and Fry (2009) with slight modification. Briefly, 0.5 mL of the extract was
mixed with 2.25 mL of distilled water in a test tube followed by addition of 0.15 mL of 5% NaNO2 solution. After 6 min, 0.3 mL of a 10% AlCl36H2O solution was added and allowed to stand for another
5 min before 1.0 mL of 1 M NaOH was added. The mixture was mixed by vortex mixer. The absorbance was measured immediately at 510 nm using spectrophotometer. Results were expressed as mg rutin equivalents in 1 g of dried sample (mg RE/g).