A full factorial design (ascorbic acid/l-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 v_max/K_m 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.

中文翻译：

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对可可豆中多酚氧化酶酶抑制作用和总多酚回收率的见解。

进行了充分的因子设计（抗坏血酸/ l-半胱氨酸抑制剂，温度和时间作为因素）研究，以增强对多酚氧化酶（PPO）活性的抑制，而不会降低可可多酚的浓度。通过以Γ表示的新方程对获得的数据进行建模，该方程将高多酚含量与降低的比PPO活性相关。在最佳值下（在96°C下70 mM抑制溶液持续6.4 min，Γ= 11.6），PPO抑制率为93.3％，总多酚含量为94.9 mg GAE / g。此外，显微镜图像证实了以分形维数测量的细胞形态变化，并解释了热处理和化学抑制剂可能导致的细胞裂解和变性。结果还表明，PPO酶最合适（较高邻苯二酚的v _max / K _m比），抑制热处理后其亲和力降低13.7倍。总的来说，这项工作提出了一种合适的，食品安全的方法，以获得酶活性低的浓缩多酚提取物。