with tap water and distilled water, respectively. Then,
peel was dried in hot air oven at 60๐C for 24 hours. Dried
peel was then taken for grinding by blender and powdered
form of plant sample was soaked in 95% ethanol, 40%
ethanol (rice whisky)or acetone in a ratio of 1:8 in the
dark, at room temperature for 3 days separately. The mixtures
were filtered through a sterile filter cloth and a sterile filter
paper (Whatman No. 1), respectively. The filtrates
obtained were subsequently concentrated to a small
volume under vacuum on a rotary evaporator (Laborata
4000; Heidolph Instruments GmbH & Co. KG, Germany).
All the concentrated extracts were stored at -20๐C under
dark condition prior to analysis. The final weight of the
crude extracts were weighted and calculated for the %yield
per original materials.
2.2 Determination of Total Phenolic content
Total phenolics of peel extracts were determined
using the method of Singleton and Rossi(20).100μl
portions of diluted extracts were introduced into test tubes
followed by addition of 750μl of freshFolin-Ciocalteu
reagent (1:10). After standing at room temperature for 5 min,
750μl of 6% (w/v) sodium carbonate (Na2CO3) was added
and allowed to completely reacted for 90 min at the room
temperature in the dark condition. And then, the absorbance
ofthe reaction mixtures were recorded at 725 nm against
blank. Total phenolics were calculated from standard
gallic acid solutions (0.02 - 0.1 mg/ml) used under the same
conditions, and concentrations were expressed as mg gallic
acid equivalent per gramdry weight (mg GAE/g DW).
2.3 Total Flavonoids of Extracts
The content of flavonoids in the peel extracts were
determined by aluminium chloride colorimetric assay (21).
Briefly, 200ml of sample extracts and 2.3 ml 30% of methanol
were added in tested tubes. Then, the mixture was added
100 μl of 0.5 M NaNO2 and 100 μl 0.3 M AlCl3, respectively.
Next, the sample solution was thoroughly mixed
with vortex and kept in the dark. And then, the absorbance
was measured 5 min later at 506 nm. Total flavonoid were