Abstract

This study aimed to valorize needlefish (Tylosurus acus melanotus) skin as a marine by-product for producing collagen hydrolysates using alkaline protease (NCA) and papain (NCP), comparing their structure, antioxidant activity, and in vitro wound-healing potential with a commercial fish collagen hydrolysate (CCH). Hydrolysates were prepared from needlefish skin using alkaline protease and papain. Structure was evaluated by SDS–PAGE and FTIR. Antioxidant activity was assessed via DPPH, ABTS, and reducing power assays, while biocompatibility and wound healing were examined using cell viability and scratch assays with BALB/3T3 clone A31 fibroblasts at 24 and 48 h. SDS–PAGE showed faint, diffuse patterns with no bands above 37 kDa, confirming extensive degradation into low-molecular-weight peptides, especially in NCA. FTIR revealed typical collagen amide bands, with broader, less intense bands in NCA and NCP indicating structural modification. NCA showed the strongest antioxidant activity, with the lowest IC₅₀ for DPPH (1.83 ± 0.06 mg/mL) and ABTS (1.34 ± 0.02 mg/mL) and highest reducing power (90.00 ± 5.27 mg TE/g), followed by NCP and CCH. All hydrolysates were non-cytotoxic and enhanced fibroblast viability, with NCA and NCP outperforming CCH (peak at 50 μg/mL after 48 h) and promoting fibroblast migration and wound closure. Alkaline protease hydrolysis was most effective for producing bioactive needlefish skin collagen peptides with superior antioxidant capacity and fibroblast-supporting activity, highlighting potential applications in wound-healing biomaterials, nutraceuticals, and marine-based functional formulations.