Biological Evaluation of a Melatonin-Infused Hyaluronic Acid–Hydroxyapatite Biopolymer Sheet for Bone Regeneration

The current research compares a multifunctional hyaluronic acid (HA)-based biopolymer sheet loaded with hydroxyapatite and melatonin (HHIM) to a plain HA biopolymer sheet for possible bone regeneration usage. Hemocompatibility testing showed that both HA and HHIM scaffolds resulted in very low hemolysis at all concentrations tested; thus, they are compatible with blood. Antibacterial evaluation demonstrated that the HA scaffold and HHIM biopolymer sheet had almost no antimicrobial effects against Streptococcus mutans, Staphylococcus aureus, Pseudomonas aeruginosa and Klebsiella pneumoniae. The antioxidant capacity measured by the DPPH assay revealed a notably higher radical scavenging activity of HHIM with respect to HA, which was concentration-dependent and almost reached the activity of ascorbic acid at the highest concentration. Likewise, the HHIM scaffold has shown higher anti-inflammatory activity, with inhibition values similar to that of diclofenac sodium, while HA had only minor effects. In vitro cytocompatibility tests confirmed that both scaffolds were conducive to cell viability, however, HHIM resulted in better cell adhesion, proliferation, and spreading at 24 and 48 hours. The enhanced biological performance of HHIM makes it a promising candidate as a bioactive scaffold for bone regeneration and maxillofacial tissue engineering.