Abstract: To find out whether the tilapia (Oreochromis) skin collagen peptides can effectively inhibit melanin production, we used enzymatic method to prepare the tilapia skin tyrosinase (TYR) inhibitory peptides (TSTIP) and studied the relevance between TYR inhibitory activity and Cu²⁺ binding activity. The results show that the product of tilapia skin hydrolyzed by alcalase for 4 h exhibited both the highest TYR inhibitory activity and Cu²⁺ binding activity, which were significantly positively correlated (R=0.856). When TSTIP was bound to Cu²⁺ or tyrosinase, the intrinsic fluorescence absorption had a consistent decreasing trend, but the UV absorption increased and the maximum absorption wavelength had a red shift. For FTIR results, TYR and Cu²⁺ bound to TSTIP mainly by carbonyl and amino groups. Circular dichroism shows that the β-turn and random curl contents of the two conjugates decreased relatively, while the β-fold content increased relatively, which was more obvious for TSTIP-Cu²⁺ conjugate. In conclusion, the structural change of TSTIP-TYR is similar with that of TSTIP-Cu²⁺ conjugate, which indicates that TSTIP can inhibit TYR's activity by binding to its Cu²⁺ active site.