Total soluble phenolics of each fraction (chloroform, ethyl
acetate, 1-butanol, aqueous) were determined with FolinCiocalteu
reagent according to the method of Singleton et
al. (1999) using gallic acid as a standard phenolic
compound. A volume of 40 µL of each fraction and
standard was transferred into separate test tubes and to
each added 3.16 mL water and 200 µL of FolinCiocalteu's
reagent. The mixture was mixed well, waited
for 8 min and then added 600 µL of sodium carbonate
solution with continuous stirring. The solution was
allowed to stand for 1 hour at room temperature in a dark
place and the absorption was measured at 750 nm using
a spectrophotometer. The concentration of total phenolic
compounds of all fractions of A.unedo fruit was
determined as milligrams of gallic acid equivalent (GAE).