**4. Conclusions**

In this study, *U. intestinalis* proteins were enzymatic hydrolyzed using five different proteases, with the trypsin-digested hydrolysates exhibiting the highest ACE inhibition rate. Based on single-factor analysis and the RSM method, the optimum conditions were as follows: pH 8.4, temperature 28.5 ◦C, E/S 4.0%, substrate concentration 15 mg/mL, enzymolysis time 5.0 h. After a series of chromatographic separation and purification steps, two novel ACE inhibitory peptides were identified: Phe-Gly-Met-Pro-Leu-Asp-Arg (FGMPLDR; MW 834.41 Da) and Met-Glu-Leu-Val-Leu-Arg (MELVLR; MW 759.43 Da). The purified peptides displayed potent ACE inhibitory activity, with IC50 values of 219.35 and 236.85 μM, respectively. Based on in vitro digestion results, FGMPLDR and MELVLR demonstrated good stability for pepsin and trypsin digestion. Furthermore, we investigated interactions between the peptides and ACE by molecular docking, and the results indicated that hydrogen bonds and interaction with the Zn2+ of ACE contribute to the ACE inhibition activities of the two peptides. Overall, these peptides derived from trypsin hydrolysates of *U. intestinalis* may be considered for use in the industrial production of functional foods. However, further research such as antihypertensive activity experiments in mice should be performed.

**Author Contributions:** N.X. and X.C. conceived and designed the experiments; S.S. and X.X. performed the experiments; X.X., X.Z. and X.S. conducted the data analysis; S.S., X.Z. and N.X. wrote and revised the manuscript.

**Funding:** This work was supported by the Key Program of Natural Science Foundation of Zhejiang Province, China (LZ17D060001) and the Ningbo International Cooperation Program (2017D10019), and the program of Ningbo Science and Technology Bureau (2016C10034). This research was also sponsored by the KC Wong Magna Fund at Ningbo University.

**Conflicts of Interest:** The authors declare no conflict of interest.
