Polyphenol oxidase and its relationship with oleuropein concentration in fruits and leaves of olive (Olea europaea) cv. 'Picual' trees during fruit ripening

Abstract

Oleuropein, the main phenolic compound of olive fruit, has important antioxidant properties that are responsible for some of the nutritional properties of fruits and the defence mechanism of leaves. Polyphenol oxidase (PPO) activity changes during fruit ripening in many plants. We studied the kinetics and molecular properties of PPO in fruits and leaves of olive (Olea europaea L.) cv. 'Picual' trees and the relationship between PPO and oleuropein concentration during fruit ripening. Polyphenol oxidase showed hyperbolic kinetics in fruits and leaves. Significant increases in PPO specific activity, V(max), K(m )and catalytic efficiency occurred during fruit ripening. Based on SDS-PAGE under partially denaturing conditions and in-gel staining with DL-3,4-dihydroxyphenylalanine, PPO activity was found in one major protein of 55 and 50 kDA in fruits and leaves, respectively. During the last stages of fruit maturation, a second 36 kDa protein was observed in fruits but not in leaves, indicating that this protein could serve as a marker of the final phase of fruit maturation. Under fully denaturing conditions, only one 27.7 kDa immunoreactive band was detected in fruits. Both the amount of PPO activity and the amount of PPO protein increased significantly during fruit maturation. Immunohistochemical studies indicated that PPO is located in the epidermis, parenchyma and companion vascular cells of leaves as well as in the epidermis of fruit. During fruit maturation, oleuropein concentration measured by HPLC significantly decreased in fruits and increased in leaves.