Copper-zinc/chitosan complex hydrogels: Rheological, degradation and biological properties

Polymer hydrogels crosslinked by therapeutic metal ions have attracted increased interest in recent years due to their unique and versatile properties. Chitosan hydrogels are widely investigated for various biomedical applications such as tissue engineering and drug delivery. Copper and zinc ions are considered as therapeutic metal ions, that have important roles in bone regeneration. The aim of this study was to investigate the physicochemical and biological properties of bimetallic-chitosan complex hydrogels with different cupric and zinc ions content. Scanning electron microscopy (SEM) revealed changes in the morphology from the microstructure with larger, tubular pores for aerogels with higher Zn content, to the sheets-like structure with long pores for samples with higher Cu content. FTIR analysis indicated the formation of bimetallic-chitosan aerogels. The obtained X-ray diffraction patterns showed a broadening of chitosan's characteristic diffraction maximum, while characterization of physical properties showed decreased swelling ability and increased shear modulus with higher Cu content. ICP-MS results showed a negligible amount of copper and zinc ions released under physiological conditions during 24 h indicating a strong physical crosslink between metal ions and chitosan chains. Furthermore, accelerated in vitro degradation showed that hydrogels maintained good stability during four weeks of lysozyme activity. The MTT assay indicated that the cytotoxicity of Cu²⁺-Zn²⁺/chitosan complexes could be adjusted by the amount of cupric ions. All results imply that Cu²⁺ and Zn²⁺ ions act as physical crosslinkers of the polymer network. Also, results are in agreement with the prediction of density functional theory (DFT) which indicated stronger chitosan-Cu tetrahedral aqua complex interactions in comparison to the chitosan-[Zn(H₂O)₄]²⁺ interactions.