Purification and characterization of catalase enzyme from Thymus nummularius

Plants used in the natural treatment of diseases from the past to the present have become the focus of the pharmaceutical industry as well as traditional medicine thanks to the bioactive phenolic components they contain. Due to its antioxidant characteristics in biological and metabolic systems, the catalase enzyme is crucial in the removal of harmful and oxidizing hydrogen peroxide (H2O2) from cells. In this study; catalase enzyme was purified from Thymus nummularius (Thyme) plant, kinetic properties (Km, Vmax), thermal and storage stability were investigated, optimum temperature, optimum pH was determined and molecular weight was determined by SDS-PAGE electrophoresis. The total amount of protein in the raw catalase extract was found to be 6.150 mg/mL, the total specific activity value was found to be 161.50 U/mg protein. Maximum activity was found in precipitation with ammonium sulfate at a salt concentration of 70%. As a result of protein determination, the total amount of protein was found to be 2.350 mg/mL and the total specific activity value was found to be 236.73 U/mg protein. Protein determination was performed after PD-10 column chromatography and the total protein amount was found to be 0.156 mg/mL, the total specific activity value was 1260.96 U/mg protein. At the end of this process, the CAT enzyme was purified 7.81 times. As a result of optimum pH and temperature studies of the enzyme; optimum pH 8.0; the optimum temperature was determined to be 25°C. When their thermal stability was compared, it was determined that they maintained 18.99% of enzyme activity at the end of 24 hours at 40 °C and 40.51% of enzyme activity at 25 °C. The storage stability was examined at 4 °C and it was found that it retained 5.96% of the catalase enzyme activity at the end of 15 days. Km value was determined as 38.151 mM and the Vmax was determined as 303.03 U/mg protein. As a result of SDS-PAGE electrophoresis, the molecular weight of the enzyme was found to be between 42 and 52 kDa.