The total flavonoid content of the extracts was determined by a
UV–vis spectrophotometer, and the samples were prepared according
to the methodology proposed by Zhishen et al. (1999), based
on the flavonoid complexation with AlCl3, which dislocates the
absorption bands to higher wavelengths. An aliquot of 250 L of
the extract at all concentrations was mixed with 1.25 mL distilled
water and 75 L 5% NaNO2. Six minutes later, 150 L 10% AlCl3
solution was added. After 6 min, 0.5 mL 1 M NaOH was added and
then the total volume was brought to 2.5 mL with distilled water.
The samples were shaken in a vortex shaker and the absorbance
measured at 510 nm. All the tests were done in triplicate and the
results expressed in mg rutin/g extract.
2