Magnoliae Flos is a traditional herbal medicine used to treat nasal congestion associated with
headache,
empyema, and
allergic rhinitis. In our preliminary screening of crude drugs used in Japanese
Kampo formulas for
melanin synthesis, the
methanol extract of Magnoliae Flos was found to exhibit strong
melanin synthesis activity. However, there have been no studies evaluating the effects of Magnoliae Flos or its constituents on melanogenesis. The present study aimed to isolate the active compounds from Magnoliae Flos that activate
melanin synthesis in
melanoma cells and three-dimensional human skin equivalent, and to investigate the molecular mechanism underlying
melanin induction. The
methanol extract of Magnoliae Flos induced an increase of
melanin content in both B16-F1 and HMV-II cells. A comparison of
melanin induction by three fractions prepared from the extract showed that the
ethyl acetate fraction markedly induced
melanin synthesis. Bioassay-guided separation of the
ethyl acetate fraction resulted in the isolation of seven
lignans (1: - 7: ). Among them, (+)-
magnolin (5: ) strongly induced
melanin synthesis and intracellular
tyrosinase activity. Furthermore, the
ethyl acetate fraction and 5: clearly induced
melanin content in a three-dimensional human skin equivalent. Molecular analysis revealed that 5: triggered the
protein expression of
tyrosinase,
tyrosinase-related protein-1, and
tyrosinase-related protein-2. Further analysis of transcriptional factors and signaling pathways demonstrated that 5: induces the
protein expression of
tyrosinase,
tyrosinase-related protein-1, and
tyrosinase-related protein-2 activated by the
protein kinase A- and
p38 mitogen-activated protein kinase-dependent pathways, leading to cAMP-responsive
element-
binding protein phosphorylation and
microphthalmia-associated transcription factor expression. These findings demonstrate the potential of 5: as a potent therapeutic agent for
hypopigmentation.