Novel Antimicrobial Agents Based on Zinc-Doped Hydroxyapatite Loaded with Tetracycline

Abstract

In this paper, we present for the first time the development of zinc-doped hydroxyapatite enriched with tetracycline (ZnHApTe) powders and provide a comprehensive evaluation of their physico-chemical and biological properties. Various techniques such as X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), and Fourier transform infrared spectroscopy (FTIR) were used for the sample's complex evaluation. Moreover, the biocompatibility of zinc-doped hydroxyapatite (ZnHAp) and ZnHApTe nanoparticles was evaluated with the aid of human fetal osteoblastic cells (hFOB 1.19 cell line). The results of the biological assays suggested that these nanoparticles hold great promise as potential candidates for the future development of novel biocompatible and antimicrobial agents for biomedical applications. The antimicrobial properties of the ZnHAp and ZnHApTe nanoparticles were assessed using the standard reference microbial strains Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Candida albicans ATCC 10231. The results of the in vitro antimicrobial assay demonstrated that both tested materials exhibited good antimicrobial activity. Additionally, these data also indicated that the antimicrobial effects of the ZnHAp nanoparticles were intensified by the presence of tetracycline (Te). Furthermore, the results also suggested that the antimicrobial activity of the samples increased with the incubation time.

Keywords: biomedical applications; composition; hydroxyapatite; in vitro biological studies; tetracycline; zinc.

Figure 1

XRD patterns of the ZnHAp (a), ZnHApTe (b), and tetracycline (c) samples and standard database JCPDS #39-1987 (d) and JCPDS #09-0432 (e).

Figure 2

FTIR general spectra obtained for ZnHAp, ZnHApTe, and Te samples.

Figure 3

(a–c) FTIR spectra in the 900–1200 cm⁻¹ spectral domain, (d–f) second derivative spectra, and (g–i) deconvoluted FTIR spectra of the ZnHAp, ZnHApTe, and Te samples.

Figure 4

General XPS spectra of the ZnHAp (a), Te (b), and ZnHApTe (c) samples.

Figure 5

High-resolution XPS spectra and curve-fitting results of C1s (a), O1s (b), Ca2p (c), P2p (d), Zn2p (e) and N1s (f) for ZnHApTe sample.

Figure 6

TEM images of ZnHAp (a) and ZnHApTe (b).

Figure 7

(a,b) SEM images and (c) particle size distribution obtained for the ZnHAp sample.

Figure 8

(a,b) SEM images and (c) particle size distribution obtained for the ZnHApTe sample.

Figure 9

EDS spectra of the ZnHAp (a), Te (b), and ZnHApTe (c) samples.

Figure 10

Percentage hemolysis of sheep red blood cells (RBCs) exposed to different concentrations of ZnHAp, ZnHApTe, and Te.

Figure 11

Cell viability of hFOB 1.19 cells incubated with 50 µg/mL (a) and 200 µg/mL (b) of ZnHAp, ZnHApTe, and Te for 24, 48 and 72 h. The results are represented as mean ± standard deviation (SD) and are expressed as percentages of control (100% viability). The statistical differences between untreated and treated groups were determined by ANOVA, and the results are significant at p < 0.05 (*); p < 0.01 (**); p < 0.001 (***).

Figure 12

Lactate dehydrogenase (LDH) activity released in the culture medium of hFOB 1.19 cells after the treatment with 50 µg/mL (a) and 200 µg/mL (b) of ZnHAp, ZnHApTe, and Te for 24, 48 and 72 h. The results are represented as mean ± standard deviation (SD).

Figure 13

Graphical representation of the logarithmic values of colony forming units (CFU)/mL of S. aureus ATCC 25923, E. coli ATCC 25922, and C. albicans ATCC 10231 microbial strains after 24, 48, and 72 h of exposure to HAp, Te, ZnHAp and ZnHApTe. The results are presented as mean ± standard error. The statistical analysis was performed by ordinary one-way ANOVA. The p-values indicated are * p ≤ 0.002, ** p ≤ 0.001, *** p ≤ 0.0001. The red stars highlight the bactericidal effects of the samples.