Multiscale Characterization and Bioactivity of Freshwater Unionid Mussel Shells as Sustainable Natural Biomaterials

The complex biological material of the freshwater mussel's shell serves various functions, contributing to the support of ecosystem services. Beyond offering structural integrity and reinforcement, the shell emerges as a significant natural biomaterial candidate thanks to its distinctive composition and structure. The aim of this study was to investigate the potential use of freshwater mussel shells from two unionid species Anodonta anatina (Linnaeus, 1758) and Unio delicatus (Lea, 1863) as a natural biomaterial. The shells were examined for their chemical composition, physical characteristics, and mechanical strength, in addition to undergoing a thorough analysis of their biointerface structure and surface morphology. The major mineral composition of CaCO₃ and the presence of other elements in their ionic forms in both mussel species demonstrated a favorable association with the shell's composition, microstructure, biointerface, biomineralization, and biological responses. The mineralogical/biointerfacial changes observed in various parts of the mussels were clearly depicted in the FE-SEM micrographs. Additionally, studies involving XRD, XRF, FTIR, EDXS, and Raman spectroscopy were used to confirm the presence of CaCO₃ crystals (aragonite and calcite forms) in the mussel shells along with the detection of organic components with various functional groups. Furthermore, in vitro bioactivity and hemocompatibility assays were conducted to investigate the effect of shell composition on the biological properties of samples of freshwater mussel shells. The results demonstrated that shell samples with different contents facilitated fibroblast attachment, migration, and promoted cell proliferation. These findings also verified the hemocompatibility of the shell samples and the favorable impact on the hemostatic behavior of the shells mineralogical structure. The findings indicated that the inherent shells of freshwater mussels Anodonta anatina and Unio delicatus are distinctive biological materials that hold promise for various applications benefiting human well-being and contributing to the preservation of environmental quality.