I.U.B.: 1.11.1.7
Peroxidase (HRP) is a hemoprotein catalyzing the oxidation by hydrogen peroxide of a number of substrates such as ascorbate, ferrocyanide, cytochrome c and the leuco form of many dyes. HRP has a molecular weight of 40 kDa and an optimum pH of 7.0. Stability: HPOFF is stable for 9-12 months at 2-8°C. HPOD is stable 2 to 3 years at 2-8°C.
Unit Definition: One Worthington Unit decomposes 1µmole of H2O2 per minute at 25°C, pH 7.0 using aminoantipyrine and phenol.
Technical Note: The RZ (Reinheitzahl) which is the absorbance ration, A403/A275, has been used as an indication of purity. However, Shannon et al., JBC, 241, 266 (1966) report that this ratio for the isozymes varies from 2.50 to 4.19. This, together with the influence exerted by buffer and pH, would seem to render questionable the preciseness of this ratio as a criterion of purity.
Numerous different methodologies are utilized for the determination of peroxidase activity. Listed below are some approximate conversions as determined by Worthington. • 1 Worthington unit = 4.6 o-dianisidine units previously used by Worthington • 1 Worthington unit = 0.62 ABTS units (µmole of dye oxidized per minute, pH 6.0, 25°C, 1.7 mM dye) • 1 Worthington unit = 2 ABST units (µmole of dye oxidized per minute, pH 5.0, 25°C, 8.7 mM dye) • 1 Worthington unit = 0.5 guiacol units (µmole of guiacol oxidized per minute, pH 7.0, 25°C) • 1 Worthington unit = 0.5 pyrogallol to purpogallin unit (mg of product per 20 seconds, pH 6.0, 20°C) • 1 Worthington unit = 5 pyrogallol to purpogallin units (µmole of product per minute at pH 6.0, 30°C)
