## 4 Control of Polyphenol Oxidase (PPO) Using Mild Technologies  
PPO is a copper-containing oxidoreductase enzyme that catalyzes the oxidation of o-diphenylols in the presence of oxygen and changes them into o-quinones, which generate dark pigments causing browning in foodstuffs [48; 89; 131]. Generally, in the plant cell, PPO is positioned in cytoplasmic organelles (e.g., thylakoid membrane of chloroplasts, mitochondria, and peroxisomes), while polyphenols as their substrates are localized in the vacuole and apoplast or cell wall compartment. The enzymatic browning happens after the breakdown of cell structure, and the subsequent interaction between PPO and polyphenols causes color change and a reduction in the antioxidant activity, leading to the deterioration of food nutritional properties [132]. In addition, the reaction between PPO and polyphenols give rise to their oxidation resulting in a decrease in TPC, which has a negative impact in the extraction process of polyphenols from food by-products [21]. Therefore, it is crucial to find an appropriate solution to reduce the drawbacks derived from PPO and prevent the oxidation of polyphenols during the extraction process. To date, many strategies have been used to control the PPO activity in food matrices, including physical and chemical methods. These methods are utilized to eliminate the critical components for the enzymatic browning reaction, such as copper ion, oxygen, products, substrate (polyphenols), and even the enzyme itself (PPO) [132]. Nowadays, the research on sustainable food processing is concentrated on replacing these conventional anti-PPO treatments with nonthermal techniques [133]. Application of nonthermal technologies in the extraction of polyphenols may have different results for the enzymatic activity because they influence PPO by several factors, including the intensity, duration, mode of exposure, gas composition, electrical input, and degradation of enzymes or substrates [134]. Nonthermal techniques decrease the amount of PPO, resulting in the better maintenance of phenolic compounds [21]. Indeed, the reduction of PPO activity has a direct impact on the maintenance of polyphenols and could increase the recovery yield of polyphenols [28; 132; 135; 136]. [PERSON] et al. (2016) reported that raisins with a low PPO activity have a more quantity of polyphenols and accordingly have a richer nutritional value [137]. As an example, CP treatment may alter the composition of different enzymes [138]. In the CP process, the higher the frequency, the further the anti-PPO activity. The loss of this enzyme in the CP treatment may be due to the activity of free radicals produced in this process on protein bands. Another possible cause for the loss of enzymes is secondary structure modification and amino acid side chain modification of proteins, which is mediated by free radicals from CP [139]. [PERSON] et al. (2020) evaluated the effect of CP on phenolic content and PPO that existed in avocado pulp combined with lime extract, and they reported that it can significantly reduce the activity of PPO and increase the phenolic content. This could be explained by the small reduction in the pH of the pulp due to the addition of lime extract, making it slightly more acidic and lessening the activity of PPO by the complexation of the Cu\({}^{+2}\) group present in its active site. The combined impact of the pH decline and the extended treatment time and gas flow results in the generation of free radicals interacting with the released enzymes and causing structural changes in them. This leads to the loss of PPO activity. As a result of this reduction, the phenolic content of the food matrix is not used as the substrate of enzymatic browning and could be extracted with better quality [28]. To the best of our knowledge, there is no research evaluating the impact of nonthermal technologies on PPO during the extraction process of polyphenols from food by-products.