This study presents the green synthesis of zinc oxide-chitosan nanoparticles (Zin-Chi-NPs) using Sepia pharaonis cuttlebone as a marine biowaste source and chitosan as a natural stabilizing agent. X-ray diffraction (XRD) analysis confirmed a crystalline structure with 69.8% crystallinity, while Fourier-transform infrared spectroscopy (FTIR) revealed characteristic functional groups such as hydroxyl (3366 cm(-)1), amine (1566 cm(-)1), and carboxyl (1354 cm(-)1), indicating effective biogenic stabilization. Field emission scanning electron microscopy (FESEM) reveals densely packed, irregularly shaped nanoparticles dispersed across the chitosan matrix. Zin-Chi-NPs exhibited notable antibacterial activity, with inhibition zones of 15 +/- 0.12 mm (E. coli), 18 +/- 0.12 mm (Streptococcus mutans), and 19 +/- 0.14 mm (Staphylococcus aureus); no inhibition was observed against Candida albicans. Cytotoxicity was evaluated on KB1 oral cancer cells using the MTT assay. A dose-dependent response was observed, with 18% cell death at 10 mu g/mL and a marked increase to 97% at 200 mu g/mL. The IC50 was calculated as 38.9 mu g/mL, suggesting moderate cytotoxic efficiency and potential therapeutic relevance at higher concentrations. Overall, the synthesized Zin-Chi-NPs demonstrate promising structural stability, antimicrobial activity, and dose-dependent cytotoxicity, supporting their potential application in biomedical and pharmaceutical fields.