Copper peptide, a low molecular weight peptide composed of glycyl-L-histidyl-l-lysine-copper, possesses anti-aging, anti-inflammatory, and wound healing properties. In this study, we investigated the effects of a combination agent CP-AcT, composed of palmitoyl copper peptide (pal-GHK-Cu) and acetyl tyrosine (N-Acetyl-l-tyrosine), on melanin production in the human malignant melanoma cell line A375 and the mouse melanoma cell line B16. Firstly, the cytotoxicity of CP-AcT at various concentrations (0-8 μg/mL) on HaCat, HFF, A375, and B16 cells was evaluated. Subsequently, the effects of the CP-AcT on tyrosinase activity both extracellular and intracellularly, as well as on melanin production in two melanoma cell lines, were evaluated under conditions that did not compromise cell viability. Additionally, quantitative gene plex (QGP) combined with branched DNA (bDNA) technology was used to analyze the effects of CP-AcT on the expression of melanin-related genes in A375 cells, with a focus on five specific genes. Finally, the effects of the CP-AcT on the expression of three proteins involved in the biosynthesis pathway of melanin: tyrosinase (TYR), dopachrome tautomerase (DCT), and endothelin 3 (EDN3) were analyzed. The results indicate that the complex CP-AcT effectively promotes melanin production in both types of melanoma cells.