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A Sensitive Method for the Estimation of Hydrogen Peroxide in Biological Materials

Abstract

DURING recent years a number of methods for the estimation of hydrogen peroxide which utilize the principle of coupled oxidation have been published. Catalase or peroxidase combines with hydrogen peroxide as it is liberated, and the complex thus formed then brings about the oxidation of such substances as nitrite1, ethanol2, cytochrome c 3, or manganous ions in presence of p-cresol4. The extent of these oxidations may be measured by manometric, colorimetric or spectrophotometric means. The method of detecting hydrogen peroxide to be described is based on the same principle but has the advantage of greater sensitivity. In this case the disappearance of the fluorescent peroxidase substrate scopoletin (6-methyl-7-hydroxy-1 : 2-benzopyrone) is observed with the aid of a suitable fluorometer. The Beckman spectrophotometer model DU used as for the estimation of fluorescence in solution (Beckman Bull. 149-G) was found satisfactory, a concentration of 2 × 10−10 mole per ml. giving a reading of 100 divisions on the intensity scale with the instrument adjusted to its highest sensitivity. Up to 2.5 × 10−9 mole per ml., the intensity of fluorescence is proportional to the concentration of scopoletin (Fig. 1).

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References

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ANDREAE, W. A Sensitive Method for the Estimation of Hydrogen Peroxide in Biological Materials. Nature 175, 859–860 (1955). https://doi.org/10.1038/175859a0

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