3D printing of alginate scaffolds for low-pH environments through slow alginic-acid hydrogel formation

3D printing additive-free alginate hydrogel scaffolds for acidic environments, including gastric-like acidic media, remains challenging because bioink flow, shape fidelity, and final crosslinking must be controlled at low pH. This balance is already difficult for alginate systems without additional rheology modifiers and becomes more restrictive under acidic conditions. A central issue is to decouple printed-architecture formation from the crosslinking step that controls the final mechanical properties of the scaffold. This challenge is addressed through a two-step reactive-printing strategy in which an alginate bioink is first converted into a transient alginic-acid pregel and then mixed with calcium carbonate for post-deposition ionic consolidation. Slow acidic aging in the presence of glucono-δ-lactone transforms the initial viscous alginate solution into a shear-thinning, yield-stress-bearing bioink. Rheological characterization showed that the storage modulus, loss modulus, and apparent yield stress evolve strongly with GDL concentration and aging time, providing a tunable processing window for extrusion. The aged alginic-acid bioink was mixed during extrusion with an alginate/CaCO₃ formulation, so that calcium release and ionic crosslinking occurred mainly after deposition. Printing fidelity was governed by pregel maturation: once a sufficient yield stress was reached, the printed structures showed limited spreading and high shape retention. Calcium-mediated crosslinking reinforced the preformed structure, with final storage moduli tunable from about 1 to more than 20 kPa. Overall, slow acidic aging decouples alginate shaping from calcium-mediated mechanical reinforcement, producing self-supporting alginate scaffolds that remain macroscopically stable under acidic conditions and may guide alginate scaffold development in low-pH environments.