%A   Tham, Sock Ying
%A   Foo, C.H.
%J Malaysian Journal of Biochemistry and Molecular Biology
%T Ascorbic Acid Assay using Sequential Injection Analysis
%X A sequential injection analysis (SIA) method for ascorbic acid assay in pharmaceuticals utilizing crude zucchini (Cucurbita pepo) extract was compared with commercial horseradish peroxidase (HRP) assay. The peroxidase activity in the crude extract of zucchini was determined by the formation of a chromophore between guaiacol (0.05M) and hydrogen peroxide (10 mM) measured spectrophotometrically at 470 nm. The method adopted for assaying ascorbic acid is based on its inhibition of peroxidase catalyzing the formation of tetraguaiacol from guaiacol in the presence of hydrogen peroxide using a sequential injection analysis (SIA) manifold controlled by an in-house developed software. The percentage of inhibition of tetraguaiacol formation is proportional to the concentration of ascorbic acid. Km values for hydrogen peroxide using HRP and zucchini peroxidase were found to be 1.530mM and 0.868mM respectively. The conditions for the SIA system used to assay ascorbic acid in pharmaceutical and food samples were optimized at 0.9 ml/min flow rate, pH of 7 and hydrogen peroxide and guaiacol concentrations at 1.75 mM and 0.025M respectively as well as reaction time of 10 seconds. Linear range for the standards was found to be 0 to 120?g/ml (r = 0.9967 - 0.9998, n = 7). The recovery of ascorbic acid from samples ranged from 91.1 to 112.1% for the zucchini extract. Results were comparable with titrimetry using 2,6-dichlorophenolindophenol (USP method). The described SIA procedure presents a simple and rapid assay for ascorbic acid in pharmaceuticals where ingredients contained in the tablets do not interfere with the analytical method.
%N 1
%K Ascorbic acid, sequential injection analysis, peroxidase, inhibition
%P 8-13
%V 12
%D 2005
%I Malaysian Society for Biochemistry and Molecular Biology
%L myais2254