Objective To explore the effect of Toll-like receptor 4 (TLR4) activation on melanogenesis in melanocytes. Methods The primary melanocytes were isolated, cultured and then stimulated by TLR4 agonist lipopolysaccharide (LPS). Firstly, the melanin content changes were detected in melanocytes, and then quantitative real-time PCR and Western blotting were employed to determine the expression levels of pre-melanosomal protein (Pmel17), tyrosinase (TYR) and microphthalmia-associated transcription factor (MITF). Finally, the melanosomes were observed by electron microscopy. Results After stimulated by LPS, melanin formation in melanocytes was significantly reduced. The expressions of Pmel17 and TYR were markedly suppressed, which depended on the regulation of transcription factor MITF. The electron microscopy showed that melanosome synthesis was inhibited. Conclusion TLR4 activation reduces melanin synthesis in melanocytes, which indicates that the bacterial products may affect melanin synthesis through innate immunity.