Luminescent Citrate-Functionalized Terbium-Substituted Carbonated Apatite Nanomaterials: Structural Aspects, Sensitized Luminescence, Cytocompatibility, and Cell Uptake Imaging

This research was funded by Spanish Agencia Estatal de Investigación of the Ministerio de Ciencia e Innovación y Universidades (MCIU), Bioscaffold project, ref. PGC2018-102047-B-I00 (MCIU/AEI/FEDER, UE), and FEDER Operational Program 2020/Junta de Andalucía-Consejería de Economía y Conocimiento/Project (B-CTS-562-UGR20). JGM, RFP, CVE, IR, FJAM and DCL belong to the Thematic network “CrysFact, ref. RED2018-102574-T (MCIU/AEI).The authors acknowledge technical support from a technician of the CIC of the University of Granada.This work explores the preparation of luminescent and biomimetic Tb3+-doped citrate-functionalized carbonated apatite nanoparticles. These nanoparticles were synthesized employing a citrate-based thermal decomplexing precipitation method, testing a nominal Tb3+ doping concentration between 0.001 M to 0.020 M, and a maturation time from 4 h to 7 days. This approach allowed to prepare apatite nanoparticles as a single hydroxyapatite phase when the used Tb3+ concentrations were (i) ≤ 0.005 M at all maturation times or (ii) = 0.010 M with 4 h of maturation. At higher Tb3+ concentrations, amorphous TbPO4 ·nH2O formed at short maturation times, while materials consisting of a mixture of carbonated apatite prisms, TbPO4 ·H2O (rhabdophane) nanocrystals, and an amorphous phase formed at longer times. The Tb3+ content of the samples reached a maximum of 21.71 wt%. The relative luminescence intensity revealed an almost linear dependence with Tb3+ up to a maximum of 850 units. Neither pH, nor ionic strength, nor temperature significantly affected the luminescence properties. All precipitates were cytocompatible against A375, MCF7, and HeLa carcinogenic cells, and also against healthy fibroblast cells. Moreover, the luminescence properties of these nanoparticles allowed to visualize their intracellular cytoplasmic uptake at 12 h of treatment through flow cytometry and fluorescence confocal microscopy (green fluorescence) when incubated with A375 cells. This demonstrates for the first time the potential of these materials as nanophosphors for living cell imaging compatible with flow cytometry and fluorescence confocal microscopy without the need to introduce an additional fluorescence dye. Overall, our results demonstrated that Tb3+-doped citrate-functionalized apatite nanoparticles are excellent candidates for bioimaging applications.Andalucía-Consejería de Economía y Conocimiento/Project B-CTS-562-UGR20- RED2018-102574-TMinisterio de Ciencia e Innovación y Universidades PGC2018-102047-B-I00European Regional Development FundAgencia Estatal de Investigació