The extraction was performed as described previously by
Nardi et al. (2003).
Air-dried bark (154 g) was chopped into
small pieces and extracted three times with 250mL of 80%
aqueous EtOH at room temperature.
The combined extracts were
filtered and the solventwas evaporated in a vacuum to give 42.9 g
of the ethanolic extract.
The aqueous suspension of this extract
was successively partitioned with ether, ethyl acetate (EtOAc)
and n-butanol (n-BuOH) furnishing an ether (2.0 g), ethyl acetate
(EAF) (17.6 g), n-butanol (FBuOH) (22.2 g) and aqueous (FAq)
(6.9 g) fractions.
The EAF (9.0 g) was separated by chromatography over a
column of silica gel water (20%) inactivated and eluted with
hexane/EtOAc (4:1) and the polarity was increased by gradual
addition of EtOAc and methanol (MeOH).
After thin layer chromatography
(TLC) analysis, four sub-fractions were obtained,
named 11SF, 19SF, 35SF and 63SF.
Sub-fraction 11SF was further
purified by flash chromatography using 40% hexane/59%
EtOAc/1% AcOH as solvent to give catechin and gallocatechin.