Additive manufacturing of bio-inspired functional granular hydrogels: from soft to mineral-based composites

The fabrication of bioinspired soft materials that are lightweight, robust, biocompatible, and processable under mild conditions remains a key challenge. These properties are vital for sustainable materials and tissue engineering scaffolds for complex environments. Developing injectable or 3D printable hydrogel and hybrid composites using natural components faces trade-offs between mechanics, biofunctionality, and sustainability. Conventional strategies rely on synthetics or energy-intensive treatments, limiting bio-integration. This Thesis addresses these by exploring how granular hydrogels, composed of microgel building blocks, can be turned into inks for 3D printing soft and mineralized materials under mild, in vivo-compatible conditions. Chapter 4 introduces a granular hydrogel ink based on κ-carrageenan, an algae-derived polysaccharide used in food applications. To enable 3D printing at room temperature, κ-carrageenan is processed into jammed microgels and reinforced by polymerizing a second, covalently crosslinked hydrogel network within each microgel, forming double-network granular hydrogels (DNGHs). This chapter first uses a second network, which greatly improves stiffness and toughness. Reinforcement strategies using metal coordination (e.g., zirconium) and glucose further boost mechanical performance while ensuring processability and scalability for food and tissue engineering. Chapter 5 builds on this platform by introducing enzyme-functionalized granular inks for localized mineralization. These inks use natural biopolymer microfragments with urease. After 3D printing, the inks are incubated in calcium- and urea-containing media, where urease triggers calcium carbonate formation. The resulting composites contain up to 92 wt% mineral and 56 vol% porosity, achieving strengths similar to trabecular bone despite weaker calcium carbonate. This strategy avoids thermal processing or bacterial systems, ensuring cell culture and in vivo compatibility. In Chapter 6, this enzyme-mediated approach is extended to hydroxyapatite (HA) composites using ALP for phosphate mineralization. As with calcium carbonate systems, microfragments are printed at room temperature and incubated in mineralizing media. By adjusting enzyme-containing fragments, porosity and HA distribution are tuned, achieving up to 68 wt% mineral and 52 vol% porosity. This sintering-free approach preserves biopolymer activity and supports osteoblast viability, promising for bone substitutes and regeneration. Overall, this research shows how natural polymers, enzymes, and 3D printing yield defined hydrogels and mineral-organic composites. The modular granular approach enables tuning of rheology, porosity, and mechanics under mild, cell-friendly fabrication. By tackling scalability, biocompatibility, and complexity, the materials and methods introduced here offer a sustainable approach with potential for in vivo bone regeneration. Mimicking biological principles like compartmentaliza