Rheological properties of gelatine hydrogels affected by flow- and horizontally-induced cooling rates during 3D cryo-printing

Relatively low viscosity with poor resolution and shape fidelity make soft gelatine hydrogel still challenging to use as an ink or bio-ink for 3D printing, despite its good shear-thinning behaviour and high potential in tissue engineering due to its biocompatibility and similarities to biological tissues. A horizontally-induced cooling platform has thus been proposed to accelerate the physical stabilisation of each printed deposit during such a multilayer printing by fastening its phase-change (gelation) transition, which would affect its flow (shear-thinning) and gel strength (viscoelasticity), and thus enable scaffolds to be created with more precise porosities and geometries. In order to verify the physical properties (sol-gel transitions, gelation point, gel strength) and the kinetics of gelatine solutions (5 and 10 wt%) during such printing conditions, the viscosity and oscillatory rheology as a function of kinetically different cooling / heating rates (5−48 °C/min) have been performed in the range from 37 to 0 vs. -30 °C. The gelation of 10 wt% gelatine was shown to decrease by 4 °C (to 18 °C) when the plate was cooled to 0 °C, and the cooling speed rate was changed from 5 to 42 °C/min, which increased its complex viscosity (η*) for 485 Pa (to 1135 Pa) by forming tight and in a flow direction-oriented clusters of multi/triple-helices, being confirmed with FTIR spectroscopy. The lower plate temperature (-30 °C) didn't change solutions' gelation transition significantly, while it only increased the η* (from 43 to 470 Pa) of 5 wt% hydrogels at lower cooling speed (5−12 °C/min) by forming larger and more disordered aggregates, getting immobilised by fast crystallisation of the aqueous medium. It can be concluded that, besides the gelatine concentration, the cooling speed has a major impact on its physical stabilisation by yielding macromolecularly differently structured gels, which shall affect the printability, as well as give different porous structures.