Bone is a well known natural self-healing composite material, which consists of an organic component in the form of a collagen fibril-based hydrogel and an inorganic component in the form of calcium phosphate (CaP) crystals which reinforce the hydrogel.

In this project, an enzymatic method inspired by the self-healing properties of bone was applied to generate hydrogel-CaP composite biomaterials intended for applications in bone regeneration. This biomimetic approach involved the enzyme alkaline phosphatase (ALP), which mineralizes bone tissue with CaP. ALP was incorporated into several hydrogel types to induce mineralization, after which physicochemical and cell biological characterization was performed. Physicochemical characterization involved determination of the amount and type of CaP formed and its distribution within the hydrogel, changes in mechanical strength resulting form CaP formation and morphology of the CaP deposits. Cell biological characterization involved assessment of the adhesion, viability and proliferation of bone-forming cells (osteoblasts). The approach developed in this project is applicable to any kind of hydrogel.

As expected, mineralization led to an improvement in compressive strength. The amount and type of CaP formed was dependent on hydrogel. Generally speaking, predominantly calcium-deficient hydroxyapatite CDHA formed in hydrogels containing a large number of calcium-binding chemical groups, e.g. catechol/poly(ethylene glycol) (cPEG), gellan gum (GG), GG enriched with polydopamine, while a mixture of CDHA and amorphous CaP formed in other hydrogels, e.g. platelet-rich fibrin (PRF) collagen, oligo(poly(ethylene glycol) fumarate) (OPF) and chitosan. CaP formation was predominantly in the outer region of hydrogels. In the case of chitosan hydrogels, ALP not only caused mineralization, but also acceleration of hydrogel formation. In the case ofGG, CaP formation also significantly promoted osteoblast adhesion and proliferation. The results obtained led to four journal publications and one patent and pave the way for the application of the hydrogel-CaP composites as implant materials for bone regeneration.