Biochemical Characterization of Peroxidase from Horseradish Armoracia rusticana

Peroxidases are used in a number of industrial and analytical applications. There is increased interest to find alternative/additional peroxidases with novel properties. Therefore, the present study focused on the establishment, purification and characterization of peroxidase from Saudi horseradish roots. For establishment the enzyme, the rate of peroxidase activity was linearly increased with increasing the peroxidase concentration and incubation time. The purification procedure of horseradish peroxidase was carried out using ammonium sulphate precipitation, chromatography on DEAE-Sepharose column followed by Sephacryl S-200 column. The molecular weight of horseradish POIII was 56 kDa. The oxidation of some phenolic compounds by horseradish POIII has been examined. The activity with the guaiacol is regarded as 100% activity. o-Phenylenediamine and guaiacol had the same peroxidase activity (100%), while o-dianisidine had moderate peroxidase activity. Pyrogallol and p-aminoantipyrine had low affinity toward POIII. The apparent Km value of POIII for H2O2 and guaiacol were 3.3 and 16.4 mM/ml, respectively. The catalytic efficiency Vmax was 0.69 and 0.71 units/ml for H2O2 and guaiacol, respectively. POIII was found to have temperature optimum at 40˚C and the enzyme activity was remained stable up to 40˚C. POIII exhibited pH optimum at pH 5.5. All the examined metal cations had partially inhibitory effects toward POIII, except of Fe3+ enhanced the activity by 160% at 5 mM. All the metal chelators caused partially inhibitory effects toward POIII, except of EDTA at 1 mM which had no effect on the enzyme. In conclusion, the study reported that horseradish POIII had the ability for oxidation of some of phenolic compounds which considered pollutant compounds for environment.