**5. Conclusions**

From the above data, we concluded that low concentrations of Ectoine (0.5–1.5 μM) could downregulate α-MSH and melanin production via the suppression of POMC and tyrosinase pathway in UVA irradiated HaCaT cells, indicating its anti-melanogenesis efficacy. Additionally, Ectoine was also involved in the suppression of intracellular ROS production in HaCaT cells. Unlike HaCaT cells, high concentrations of Ectoine (50–400 μM) were able to show the similar effect in B16F10 melanoma cells that have signified the fact that keratinocytes could play a key role in the Ectoine mediated anti-melanogenesis and skin-whitening effects in skin cells. Most importantly, Ectoine mediated beneficial effects via the activation of the Nrf2 pathway, that induces the expression of antioxidant proteins HO-1, NQO-1, and γ-GCLC. AKT was shown to be the first signaling pathway that initiates the activation of Nrf2 followed by the other pathways (p38, PKC, and CKII). Finally, silencing of Nrf2 directly provided the evidence that Nrf2 plays a key role in the regulation of intracellular ROS as well as the α-MSH production. We concluded that the main whitening mechanism of Ectoine should be reasoned by the inhibition of ROS-p53/POMC-α-MSH pathway in UVA-irradiated HaCaT cells. Therefore, Ectoine or its derivatives could be an active ingredient in the moisturizers and lotions that are used as potential and natural-based skin whitening agents in the cosmetic industry.

**Author Contributions:** Conceptualization—Y.-C.H. and H.-L.Y.; Methodology—X.-Z.C., Y.V.G., H.-R.Y. and J.-Y.C.; investigation—X.-Z.C., Y.V.G., H.-R.Y. and J.-Y.C.; data curation—X.-Z.C., Y.V.G., H.-R.Y. and J.-Y.C.; writing—original draft preparation—Y.-C.H. and Y.V.G.; writing—review and editing—Y.-C.H. and Y.V.G.; supervision—Y.-C.H. and H.-L.Y.; funding acquisition—Y.-C.H. and H.-L.Y. All authors have read and agreed to the published version of the manuscript.

**Funding:** This study was supported by the Ministry of Science and Technology, Asia University, and China Medical University, Taiwan (grants MOST-106-2320-B-039-054-MY3 and MOST-107-2320-B-039-013-MY3, CMU 107-ASIA-15, and CMU106-ASIA-19. This work was financially supported by the "Chinese Medicine Research Center, China Medical University" from The Featured Areas Research Center Program within the framework of the Higher Education Sprout Project by the Ministry of Education (MOE) in Taiwan (CMRC-CHM-8).

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