Marine-derived chitosan is a valuable biomaterial due to its biocompatibility and functional properties. In this work, chitosan extracted from the cuttlebone of Sepia pharaonis was used to prepare silverâ��chitosan nanoparticles (Ag-Chi-NPs) by a chemical reduction process. The synthesis was confirmed by comprehensive characterization: UVâ��Vis spectra displayed a plasmon resonance peak at 420 nm, FTIR identified interactions between amino and hydroxyl groups, XRD revealed crystalline silver phases, and FESEM showed spherical nanoparticles with a size range of 15â��40 nm. Following these analyses, the Ag-Chi-NPs exhibited significant bioactivity. Antioxidant activity was substantial, with hydrogen peroxide scavenging of 84.5 (ICâ� â�� 18.09 µg/mL), DPPH radical inhibition of 65.6 (ICâ� â�� 29.04 µg/mL), and a total antioxidant capacity of 55.5 at 100 µg/mL. Antimicrobial assays further demonstrated inhibition zones of 21 ± 0.6 mm for Streptococcus mutans, 19 ± 0.2 mm for Staphylococcus aureus, and 14 ± 0.1 mm for Escherichia coli, while Candida albicans showed no susceptibility. Cytotoxicity testing using KB-1 carcinoma cells indicated excellent biocompatibility, with cell viability above 95 at lower concentrations and concentration-dependent inhibition at 200 µg/mL. Overall, these results highlight Ag-Chi-NPs as sustainable bio-nanocomposites that combine antioxidant, antibacterial, and cytocompatibility properties, making them promising candidates for wound healing materials, antimicrobial coatings, and potential therapeutic applications. © 2025 Elsevier B.V., All rights reserved.