The total flavonoids were determined by the aluminum
chloride colorimetric method of Siddhuraju et al. [17]. Briefly,
0.5 mL of each sample with appropriate dilution was placed
in a 10 mL of graduated test tube, added with 3.5 mL of distilled
water and 0.3 mL of sodium nitrite (7.5 g/100 mL),
and then mixed well. 3 mL of aluminum chloride solution
(1 g/100 mL) was added 5 min later. After 6 min, 2 mL of sodium
hydroxide solution (4 g/100 mL) was added, and the
total volume was made up to 10 mL with distilled water,
and the mixture was vibrated for 20 s. After 15 min, the absorbance
was measured against a blank at 510 nm. The total
flavonoid content was calculated from a calibration curve of
rutin and expressed as rutin equivalent (mg RE/g).