Revisiting Physicochemical and Biological Properties of Zn2+ - Enriched Hydroxyapatite

Łukasz Pajchel,1 Monika Budnicka,1 Milena Wawryniuk,2 Roman Grygoruk,3 Ewa Bednarczyk,3 Joanna Kolmas1 1Department of Pharmaceutical Chemistry and Biomaterials, Faculty of Pharmacy, Medical University of Warsaw, Warsaw, Poland; 2Department of Toxicology and Bromatology, Faculty of Pharmacy, Medical University of Warsaw, Warsaw, Poland; 3Department of Construction Engineering and Biomedical Engineering, Faculty of Mechanical and Industrial Engineering, Warsaw University of Technology, Warsaw, PolandCorrespondence: Joanna Kolmas, Department of Pharmaceutical Chemistry and Biomaterials, Faculty of Pharmacy, Medical University of Warsaw, Banacha 1 str, Warsaw, 02-097, Poland, Email [email protected]: This nanotechnology-oriented study provides insights into the nanoscale structural and compositional modulation of hydroxyapatite. This study investigated the effect of zinc ions (Zn2+) content (0– 1.8 mol%) in nanocrystalline hydroxyapatite on its physicochemical and biological properties, focusing on biomedical applications.Materials and Methods: A series of zinc-enriched nanocrystalline hydroxyapatites was synthesized via aqueous precipitation. Their ultrastructure and crystallinity were characterized by transmission electron microscopy (TEM) and powder X-ray diffraction (PXRD), including unit cell analysis. Chemical composition—specifically OH−, HPO42-, and CO32- groups—was examined using Fourier transform infrared spectroscopy (FT-IR), Raman spectroscopy, and solid-state nuclear magnetic resonance (ssNMR). Zn2+ content and release over seven weeks were quantified via flame atomic absorption spectrometry (F-AAS). Cytotoxicity was evaluated using MTT and NRU assays.Results: Increasing Zn2+ concentration led to reduced crystal size and crystallinity. Zinc ions were incorporated both into the crystalline core and the hydrated surface layer of hydroxyapatite. At concentrations ≥ 1.0 mol%, an amorphous zinc phosphate phase appeared. Higher Zn2+ levels also correlated with decreased hydroxyl groups and carbonate impurities, accompanied by increased water content and acidic phosphate groups. Zinc ion release remained minimal across all samples, independent of the initial zinc concentration. Cytotoxicity assays revealed that samples containing 0– 0.6 mol% Zn2+ were non-toxic, while those with 1.0 mol% and 1.8 mol% Zn2+ exhibited cytotoxic effects.Conclusion: Zn-doped hydroxyapatite containing up to 0.6 mol% Zn2+ exhibits enhanced structural stability and cytocompatibility, establishing 0.6 mol% as the optimal threshold for biomedical applications.Keywords: zinc, nanocrystalline calcium phosphates, physicochemical analysis, ion release, chemical structure