Abstract:
Species of the family Combretaceae are used extensively in traditional medicine against
inflammation and infections, and although antibacterial activity has been reported in non-polar
extracts, further rationale for the widespread use of the Combretaceae is expected to exist.
Methanol extracts of leaves of ten different Combretum species were evaluated for antioxidant
activity by spraying TLC chromatograms of each leaf extract with 2, 2-diphenyl-1-
picrylhydrazyl (DPPH). Compounds with antioxidant activity were detected by bleaching of the
purple DPPH colour. Leaf extracts of Combretum apiculatum subsp. apiculatum had the most
antioxidant compounds. This species was consequently selected for phytochemical
investigation. A DPPH assay-directed fractionation of the leaf extracts of C. apiculatum led to
the isolation of four antioxidant compounds from the ethyl acetate and butanol soluble fractions.
The structures of the compounds were determined by spectroscopic analyses (1H-NMR, 13CNMR
and MS) and identified as: cardamonin (1), pinocembrin (2), quercetrin (3) and kaempferol (4). In a quantitative antioxidant assay, the more polar fractions (ethyl acetate and
butanol) obtained by solvent-solvent fractionation had the highest antioxidant activity among the
solvent fractions obtained from C. apiculatum, with EC50 values of 3.91 ± 0.02 and 2.44 ± 0.02
μg/mL respectively. Of the four isolated compounds, quercetrin (4) and kaempferol (3) had the
strongest antioxidant activity, with EC50 values of 11.81 ± 85 and 47.36 ± 0.03 μM respectively.
Cardamonin (1) and pinocembrin (2) did not demonstrate strong activity. L-ascorbic acid was
used as standard antioxidant agent (EC50 = 13.37 ± 0.20 μM or 2.35 μg/mL).
The cytotoxicity of cardamonin and pinocembrin was evaluated on Vero kidney cells using the
MTT (3-(4, 5-dimethylthiazol)-2,5-diphenyl tetrazolium bromide) assay with berberine as
positive control. At concentrations higher than 50 μg/ml of cardamonin or pinocembrin, the cells
were not viable. Cardamonin was more toxic (LC50 = 1.97 μg/ml) than pinocembrin (LC50 =
29.47 μg/ml) and even the positive control, berberine (LC50 = 12.35 μg/ml).
