In the ovarian ascorbic acid depletion (OAAD) assay of Parlow (1958, 1961) i.v. injection of luteinizing hormone (LH) or human chorionic gonadotrophin (HCG) into young, pseudopregnant female rats, causes a dose-related depletion of ovarian ascorbic acid. However, the specificity of the assay is directly related to the method of measuring the ovarian ascorbic acid, and a variety of procedures have been described. Most workers use the method of Mindlin & Butler (1938) based on the oxidation of ascorbic acid with 2,6-dichlorophenol-indophenol, but the disadvantages are that the colour reaction is unstable and non-specific, depending only on the reducing properties of ascorbic acid.
Schmall, Pifer & Wollish (1953) reacted diazotized 2-nitro-4-methoxyaniline with ascorbic acid, and the resulting yellow solution in the presence of alkali developed an intense blue colouration. Freed (1966) modified this method and used it for measuring the ascorbic acid content of food and drugs, obtaining comparable results to
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