Abstract
Application of the α,α-dipyridyl method for determination of ascorbic acid in urine is described. The urine sample was acidified with trichloroacetic acid and shaken with activated carbon to remove interfering substances. The acid filtrate was first neutralized (pH 7.0) by adding Na 2HPO 4. The dehydroascorbic acid was then reduced back to ascorbic acid by incubation with dithiothreitol. After removal of the excess dithiothreitol with N-ethylmaleimide, ascorbic acid was determined by measuring the reduction of ferric ion. The ferrous ion produced was coupled to α,α-dipyridyl in the presence of H 3PO 4. Ferrous ion in urine samples, which theoretically interferes with the method, was removed by a combination of Na 2HPO 4 and H 3PO 4.
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