**4. Conclusions**

Low-intensity ultrasonic processing stimulated the activation of PPO but exposure to high-intensity ultrasonic processing exhibited an inactivation effect on the PPO enzyme. The increasing power under ultrasonic processing induced the polydispersity of the protein, as well as the change of the secondary and tertiary structures of the protein. The loss of α-helical contents after ultrasonic processing led to the reorganization of the secondary structure of the protein. High-intensity ultrasonic processing caused the exposure of Trp residues to a more polar environment, thereby leading to the quenching of fluorescence intensity and subsequently decreasing the suitability of the catalytic center of the enzyme for reaction with substrates.

**Supplementary Materials:** The following are available online. Figure S1: Elution profile of protein extraction, Figure S2: Lineweaver–Burk equation of purified enzyme using catechol and pyrogallol as substrates. Table S1: Purification of Polyphenol Oxidase (PPO) from orange (*Citrus sinensis* Osbeck).

**Author Contributions:** Data curation, A.M.; formal analysis, Y.L.; project administration, X.X. and S.P.; software, S.L.; supervision, W.H.; visualization, J.L. and A.I.; writing—original draft, L.Z. (Lijuan Zhu); writing—review and editing, L.Z. (Linhu Zhu).

**Funding:** This study was gratefully supported by the Fundamental Research Funds for the Central Universities (No. 2662018JC018), National Natural Science Foundation of China (No. 31401507), the project "Research and Demonstration on Key Technologies of New Citrus Juice Processing" (Project No. 2017 yfd0400701) and Hubei Province Technical Innovation Special Major Projects (No. 2018ABA072).

**Conflicts of Interest:** No conflict of interest exits in the submission of this manuscript, and manuscript is approved by all authors for publication. The submitted work was not carried out in the presence of any personal, professional or financial relationships that could potentially be construed as a conflict of interest.
