Total flavonoid content was determined by a method described
by Zhishen et al. (1999) with some modifications. An aliquot of
the extracts (250 L) was mixed with distilled water (1.25 mL)
and 5% (w/v) NaNO2 solution (75 L). After stirring for 5 min, 10%
(w/v) AlCl3 solution (150 L) was added. After 6 min of stirring, 1 M
NaOH solution (500 L) and distilled water (275 L) were succes-
sively added, and then the absorbance was measured at 510 nm.
To calculate TFC, (+)-catechin standards at different concentrations
(100–400 g/mL) were used.