Effect of glutathione and Maillard reaction products prepared from glucose or fructose with glutathione on polyphenoloxidase from apple—II. Kinetic study and mechanism of inhibition

Maillard reaction products (MRPs) prepared from aqueous equimolar glucose or fructose with glutathione (GSH) model solutions (0.25 M), when heated at 90 °C for 15–39 h, were previously recognized as strong apple PPO inhibitors. This paper reports the inhibition mode of the purified o-diphenoloxidase activity from apple, using 4-methylcatechol as the substrate. Assuming a reversible inhibition, Lineweaver-Burk plotting showed that both MRPs were mixed-type inhibitors, glucose/GSH being the most efficient model system, with Ki values ranging between 11.6 and 1.1 μl MRPs, according to the heating treatment and the sugar tested. Enzyme inactivation took place during pre-incubation (0–180 min) of PPO with various MRPs at 0 °C, and was only partly reversed by exhaustive dialysis or gel filtration. Activity initially lost was also partly restored when cupric ions (CuSO4) were added to the reaction medium, suggesting a chelating effect of copper ion at the active site of PPO, as already observed with cysteine-derived MRPs. MRPs derived from the tripeptide GSH were more potent inhibitors of apple PPO than those prepared from cysteine.