2.5. Measurement of flavonoid content
The flavonoid content of the apple juice was measured using an aluminum chloride colorimetric method. A volume of apple juice (25 ul) was added to a 96-well plate containing 125 ul of distilled water and 10 ll of 5% sodium nitrite solution. After 6 min, 15 ul of 10% (w/v) aluminum chloride was added to each well of the 96 well plate. The reaction mixture was incubated for 5 min, mixed with 50 ul of 1 M sodium hydroxide, and diluted with another 275 ul of distilled water. The absorbance of the mixture was measured immediately using a plate reader (VICTORTM X, Perkin Elmer, USA) at 510 nm. The results were expressed as catechin equivalent value calculated based on a standard curve of catechin solutions.