**5. Conclusions**

In conclusion, taken together, data presented in this study demonstrate that AphaMax® powder has pharmacologically promising positive activity on DNBS-mediated experimental colitis.

Although several mechanisms could be proposed to explain AphaMax ® beneficial e ffects, we highlight the reduction of inflammatory and oxidant mediators. However, further studies should be performed to resolve the exact molecular mechanisms. A limitation of this study is the lack of long-term observation following AphaMax ® treatment, since DNBS rats recovered within 10 days of the experiment, making it di fficult to observe and compare the long-term e ffects of treatment on IBD. In addition, hapten-administration shares many features with human inflammatory bowel diseases, but the "acute" inflammatory response evoked should be di fferent from the mechanism of chronic colitis in human IBD.

Moreover, the aim of the current study did not include the analysis of the e ffects of AphaMax ® treatment on the colon functions, we are aware that changes in colon motility and in intestinal permeability are described in DNBS-induced colitis in rats. Since it is well reported that antioxidant compounds are able to improve intestinal motility [56], additional studies could be performed to assess whether the mitigation of the inflammatory process by the AphaMax ® treatment, could help the colonic mucosa and the smooth muscle to recover their functionality. We hope that results of this study can prompt further investigations on AphaMax ® as a natural product in the managemen<sup>t</sup> of intestinal inflammatory diseases.

**Author Contributions:** Conceptualization, M.G.Z. and R.S.; Investigation, M.G.Z., A.B., G.C. and D.N.; Data curation, M.G.Z. and D.N.; Supervision S.S. and R.S.; Writing—original draft, M.G.Z.; Writing—review & editing, M.D.C., S.S. and R.S. All authors have read and agreed to the published version of the manuscript.

**Funding:** This work was supported by the company Nutrigea, Borgo Maggiore, Republic of San Marino.

**Conflicts of Interest:** S.S. is the R&D Director of the company that owns AphaMax ®'s patent.
