The comprehensive control of hydroxyapatite (HAp), involving morphological and structural variations, particle sizes, and defect formations, has garnered considerable attention for its versatile functionalities, rendering it applicable in diverse contexts. This work examined the shape, structure and optical characteristics, and defect formation in hydroxyapatite (HAp) extracted from Nile tilapia (Oreochromis niloticus) scales with various pre-treatments through experiments and density functional theory (DFT) calculations. Utilizing scanning electron microscopy, our findings revealed that dried fish scales (FS-D) exhibited a layered pattern of collagen fibers, while boiled fish scales (FS-B) had smoother surfaces and significantly reduced collagen content. After calcination, the FS-D sample produced nanorods with an average length of 150 ± 44 nm, whereas the FS-B samples yielded agglomerated spherical particles whose size increased with the rising calcining temperature. In-depth analysis through X-ray diffraction and Fourier-transform infrared spectroscopy confirmed the presence of biphasic calcium phosphates in the FS-B samples, while the FS-D sample presented a pure HAp phase. The boiled fish scale calcined at 800 °C (FS-B800) exhibited an optical band gap (Eg) of 5.50 eV, whereas the dried fish scale calcined at 800 °C (FS-D800) showed two Eg values of 2.87 and 3.97 eV, as determined by UV-visible spectroscopy. DFT calculations revealed that the band gap of 3.97 eV correlated with OH- vacancies, while that of 2.87 eV indicated Mn-substituted HAp, explaining the blue powder. The Eg value for the white powder resembled pure HAp, S- and Cl- substituted OH- vacancies, and various cations substituting Ca sites of HAp. Different pre-treatment procedures influence the characteristics of HAp, offering opportunities for applications in bone replacement and scaffolds for bone tissue engineering.
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