Kinetic characterisation and thermal inactivation study of polyphenol oxidase and peroxidase from table grape (Crimson Seedless)

Polyphenol oxidase (PPO) and peroxidase (POD) were extracted from a table grape (Crimson Seedless) using Triton X-I 14 and characterized using spectrophotometric methods. Both PPO and POD were activated by acid shock. However, in the presence of the anionic detergent sodium dodecil sulphate (SDS), PPO was activated whereas POD was inactivated. The enzymes were kinetically characterized and both followed Michaelis-Menten kinetics, although with different values of their kinetic parameters. The V-m/K-m ratio showed that Crimson Seedless grape PPO presents a similar affinity for 4-tert-butyl-catechol (TBC) whether activated by acid shock (0.018 min(-1)) or SDS (0.023 min(-1)). With regards to POD, the K and V-m values for 2,2'-azinobis(3-ethylbenzothiazolinesulphonic acid) (ABTS) were 0.79 mM and 1.20 mu M/min, respectively. In the case of H2O2, the K-m and V-m value were 0.4 mM and 0.93 mu M/min, respectively. PPO and POD showed similar thermostability, losing >90% of relative activity after only 5 min of incubation at 78 degrees C and 75 degrees C, respectively. In addition, PPOs activation energy was similar to that obtained for POD (295.5 kJ/mol and 271.9 kJ/mol, respectively). (C) 2008 Elsevier Ltd. All rights reserved.