Liquid Chromatography Quadrupole Time-of-Flight Tandem Mass Spectrometry Characterization of Ethyl Acetate Fraction from and Its Anti-Melanogenesis Effect in B16F10 Melanoma Cells and Zebrafish Model.

Melanin overproduction causes various skin diseases, such as spots, freckles, and wrinkles, resulting in the requirement of melanin synthesis inhibitors like 1-phenyl-2-thiourea (PTU) and kojic acid, which have been commonly used in the pharmaceutical industry. However, these inhibitors can cause side effects such as skin irritation and allergies. Therefore, it is necessary to develop safe and effective melanin inhibitors from natural resources. The purpose of this study was to investigate a whitening agent from natural substances using B16F10 melanoma cells and zebrafish model. We investigated the melanogenesis-inhibiting activities of the fractions from extract. The ethyl acetate fraction from extract (SPEF) significantly decreased tyrosinase activity. SPEF also significantly reduced α-melanocyte stimulating hormone (MSH)-induced intracellular tyrosinase activity and melanin content in B16F10 cells. Moreover, SPEF inhibited the expression levels of key melanogenic proteins such as tyrosinase, TRP-1, TRP-2, and MITF by downregulating the phosphorylation levels of CREB and PKA in α-MSH-stimulated melanoma cells. Furthermore, SPEF significantly suppressed melanin synthesis in the zebrafish model with no developmental toxicity. LC-Q-TOF-MS/MS analysis identified that SPEF was composed of 12 phytochemical compounds, including diterpenes, which were the dominant metabolites. These results altogether show that SPEF effectively suppresses melanogenesis in B16F10 melanoma cells and in a zebrafish model, with potential for usage in pharmaceuticals and cosmeceuticals.