Development and Mechanical Characterization of Novel Gelatin-Fish Oil Composite Hydrogel for 3D Bioprinting

Gelatin-based hydrogels are attractive bioinks for extrusion bioprinting because they combine biocompatibility, tunable rheology, and the ability to incorporate bioactive compounds. In this study, a novel multicomponent gelatin hydrogel containing fish oil, Tween 80, lecithin, glycerin, psyllium husk, and ascorbic acid was initially designed using AI-supported formulation screening and subsequently optimized experimentally for extrusion-based 3D printing. Nine acellular hydrogel constructs were fabricated using printing speeds of 1, 2, and 5 mm/s and infill densities of 5, 15, and 30%. Optical imaging, storage stability, experimentally measured density, water-contact-angle measurements, ATR-FTIR spectroscopy, and compression testing using flat cylindrical and spherical indenters were employed to characterize the printed constructs. Mass losses exceeded 60% after 7 days and reached approximately 91% after 110 days. Measured densities ranged from 0.22 to 1.24 g/cm³ across samples. All samples were hydrophilic, with contact angles of 30.07-50.63°, and higher infill increased contact angle. ATR-FTIR showed similar chemical profiles, with no clear infill-related chemical changes, suggesting that mechanical differences mainly reflected physical architecture and heterogeneity. Flat indentation generally produced higher maximum loads, except for sample S6, which reached 396.24 N under spherical loading and had the highest density (1.24 g/cm³), suggesting that its exceptional mechanical response may be associated with local material compactness and structural heterogeneity. Stress-ratio analysis yielded a mean σr/σtrue value of 4.43 with a coefficient of variation of 97.41%, while Bland-Altman analysis showed a mean bias of 0.15 MPa and limits of agreement from -0.70 to 1.01 MPa, demonstrating that the two testing methods are not directly interchangeable. Overall, hydrogel composition, internal network structure, printing infill, and local contact mechanics influenced mechanical behavior, highlighting the need for reproducible, method-specific hydrogel characterization.