INHIBITORY AND KINETIC STUDY OF PARTIALLY PURIFIED TYROSINASE FROM IRAQI QUINCE FRUIT

Enzymatic browning by tyrosinase was indeed responsible for developing new compounds that make a significant contribution to the unfavorable aroma, taste, and color of food when tissue is damaged during the storage and processing of fresh fruits and vegetables. Tyrosinase is one of these oxidizing enzymes containing copper and is widespread in many plants and organisms, from bacteria to mammals. Tyrosinase enzyme oxidizes the hydroxyl group in monophenols to di-phenols, which are oxidized to quinones. Quinones are highly sensitive active molecules that can polymerize with each other or with other proteins and lead to the formation of brown pigments. Because of the importance of this enzyme from a nutritional point of view, this study aimed to delay or prevent the enzymatic browning through inhibit tyrosinase activity. Tyrosinase of quince (Cydonia oblonga Miller) fruit pulp was partially purified through (NH4)2SO4 precipitation, dialysis, and ion-exchange chromatography and was used for its characterization. The results indicated that crude enzyme-specific activity was (342.07) U/mg. After applying precipitation by ammonium sulfate and dialysis with a specific activity of (217.86) and (258.09) U/mg and purification fold of (0.63) and (0.75) respectively. After ion-exchange chromatography using DEAE- Cellulose, the final result revealed one peak of tyrosinase with the specific activity of (11406.77) U/mg and purification fold of (33.34). Characterization of the partially purified enzyme showed that optimum pH and temperature were 6.8 and 40 respectively. However, pH stability proved in a pH range from 6 to 7.5, while thermal stability was determined at (40°C). The kinetic and thermal parameters of partially purified tyrosinase were assessed: Km (1.96 mM), Vmax (228.13 U.ml-1.min-1), Ea (6.742 Jole/mol), ΔH (4.14 Jole/mol), Q10 (22.391), Kcat (5.00 min−1) and Vo (2.551 mM−1.min−1).