| Summary: | A full factorial design (ascorbic acid/<span style="font-variant: small-caps;">l</span>-cysteine inhibitors, temperature, and time as factors) study was conducted to enhance inhibition of polyphenol oxidase (PPO) activity without decreasing cocoa polyphenol concentrations. The data obtained were modelled through a new equation, represented by Γ, which correlates both high polyphenol content with reduced specific PPO activity. At optimized values (70 mM inhibitory solution at 96 °C for 6.4 min, Γ = 11.6), 93.3% PPO inhibition and total polyphenol of 94.9 mg GAE/g were obtained. In addition, microscopy images confirmed the cell morphological changes measured as the fractal dimension and explained the possible cell lysis and denaturation as a result of heat treatment and chemical inhibitors. Results also showed that PPO enzyme was most suitable (higher <i>v</i><sub>max</sub>/<i>K<sub>m</sub></i> ratio) for catechol, with a reduction in its affinity of 13.7-fold after the inhibition heat treatment. Overall, this work proposed a suitable and food-safe procedure for obtaining enriched polyphenol extract with low enzyme activity.
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