We have developed a new method for determination of serum ceruloplasmin from its oxidase activity, in which o-dianisidine dihydrochloride is used as substrate. o-Dianisidine dihydrochloride is more stable than is the more widely used p-phenylenediamine substrate, and forms a stable product that is measured at 540 nm. Correlation was good between results of this method and those obtained with both a p-phenylenediamine substrate method (r = 0.98) and a radial immunodiffusion procedure (r = 0.97). There is little or no interference from reducing or colored components of serum. The coefficient of variation (day-to-day) for the method was 4.2%. A normal range of 62-140 U/ liter has been determined by the reported method.
We describe a method for calculating the absorptivity (in terms of substrate consumed) of the colored solution obtained when o-dianisidine dihydrochloride is oxidized by ceruloplasmin. By oxidizing o-dianisidine dihydrochloride with known amounts of hydrogen peroxide we could determine that the molar reacting ratio of o-dianisidine to hydrogen peroxide is 2:1. So calculated, absorptivity is 9.6 ml µmol-1 cm-1, the figure used to estimate ceruloplasmin oxidase activity in terms of International Units.
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