Kinetic studies and partial purification of peroxidase in wild pear

Peroxidase, extracted from wild pears was isolated by ammonium sulfate precipitation technique and purified by ion exchange chromatography. The crude enzyme having 19.5 U/mL activity and 1.3 U/mg specific activity was subjected to ammonium sulfate precipitation technique for partial purification and the resulted activity and specific activity were 16.5 U/mL and 3.17 U/mg respectively. After ion exchange chromatography through DEAE-cellulose, fraction between 35-42 exhibited maximum activity of 15 U/mL and specific activity of 7.14 U/mg. The enzyme under discussion was found to be quite active with optimum temperature of 45oC. Optimum pH for the enzyme was 6. Thermal treatment of crude extract of wild pears peroxidase was more stable at pH 6. It was found that enzyme followed the Michealis-Menton mechanism and 21 units/mg.protein and 70 mM were the calculated values for Vmax and Km in precence of various concentrations of guaiacol and constant concentration of H2O2. The results showed that wild pear peroxidase was a thermostable enzyme. After 30 min at 60°C, the remaining activity was 35%.