Assay of total anthocyanins in Roselle extracts by
the pH differential method(24)
A stock solution (5 mg/mL) of Roselle extract
was freshly prepared in water. Two diluted solutions
(both 1 mg/mL) were prepared from the stock solution.
The first one was diluted with 0.025 M potassium
chloride buffer pH 1.0 and the other was diluted with
0.4 M sodium acetate buffer pH 4.5. After equilibrium
for 15 minutes at room temperature, 200 mL of each
diluted sample were transferred to a 96-well plate and
the UV absorbance was measured at 520 and 700 nm.
The absorbance of the sample was calculated according
to Eq (2):
A = (A520nm-A700nm)
pH 1.0-(A520nm-A700nm)pH4.5(2)
Where; A is absorbance of diluted sample,
A520nm is absorbance of diluted sample at 520 nm and
A700nm is absorbance of diluted sample at 700 nm.
Total monomeric anthocyanins in dried Roselle
extract was calculated as cyaniding-3-glucoside
according to Eq (3)