Melanin is known as one of the main pigment components of ink in pharaoh cuttlefish, which has a strong bond with the hydroxyl group of TiO2. The aim of this study was to determine the appropriate method for extracting melanin from the ink of pharaoh cuttlefish fish (Sepia pharaonis). Melanin was extracted using four methods i.e., NaOH and HCL, extraction using squid ink dilution, extraction using a drying ink bag, and extraction by centrifugal washing. The Radius Power method was also used to measure the regenerative power of the sample. Data analysis showed a significant difference among different extraction methods so the method of washing with deionized water and centrifuge with 66.12% and the method of washing with deionized and homogenized water with 22.42% were determined as the optimal methods. This is despite the fact that the best antioxidant performance of melanin in the washing method was observed using sodium hydroxide and hydrochloric acid and the washing method with deionized water and centrifugation. The best antioxidant performance was observed in all four methods at 1 mg/ml. According to the obtained results, among the studied methods, the method of washing with deionized water and centrifugation was determined as the best method of melanin extraction.