Isolation method 1 (Rakotomalala, 1

Isolation method 1 (Rakotomalala, 1992). To aqueous extract
obtained from each of the three extraction methods was added
ammonium sulphate to a final concentration of 20 g/L, in order to
precipitate proteins by increasing the ionic strength. To make CGA
more soluble in ethyl acetate, 4% phosphoric acid was added.
Extracts were then treated three times with methylene chloride
(300 mL) to eliminate caffeine in the organic phase. The aqueous
phase containing CGA was extracted 4 times using ethyl acetate
(300 mL). The four ethyl acetate phases were pooled, dried with
anhydrous sodium sulphate (10 g), filtered (N81 filter paper,
Whatman, Mexico City, Mexico), and dried at 40 8C and 120 rpm in
a rotary evaporator. The residue was analyzed by TLC and by HPLC.