Bone tissue substitutes are increasing in importance. Hydroxyapatite (HA) and β-tricalcium phosphate (β-TCP) act as a cell matrix and improve its mechanical properties. One of their raw materials is marine-origin by-products. Objectives: To synthesize, characterize, and evaluate the cellular cytotoxicity of a 3D biomaterial based on HA and β-TCP from mussel shells. Methods: We prepared pellets with 150, 200, and 250 mg and evaluated them through sintering, XRD, FTIR, ICP-OES, Scanning Electron Microscopic (SEM), and immunocytochemical tests. The Alamar Blue method was applied to the Balb-T3T cell line within 72 h to evaluate cytotoxicity. Results: Our biomaterials presented a smooth surface with slight irregularity and porosities presenting different diameters and morphologies and showed chemical, morphological, and ultrastructural similarity to bone hydroxyapatite, mainly the 150 and 200 mg pellets. Significance: We produced promising HA/β-TCP bioceramics with characteristics that allowed cell culture, promoting adhesion, spreading, and proliferation.