Preparation, characterization, and assessment of cytotoxicity, antioxidant, antibacterial, antifungal, and cutaneous wound healing properties of titanium nanoparticles using aqueous extract of Ziziphora clinopodioides Lam leaves

In recent decades, nanotechnology is growing rapidly owing to its widespread application in science and industry. The aim of the experiment was chemical characterization and evaluation of cytotoxicity, antioxidant, antibacterial, antifungal, and cutaneous wound healing activities of titanium nanoparticles using aqueous extract of Ziziphora clinopodioides Lam leaves (TiNPs@Ziziphora). These nanoparticles were characterized by fourier transformed infrared spectroscopy (FT-IR), field emission scanning electron microscopy (FE-SEM), energy dispersive X-ray spectroscopy (EDS), and UV–visible spectroscopy. The synthesized TiNPs@Ziziphora had great cell viability dose-dependently (Investigating the effect of the plant on human umbilical vein endothelial cells (HUVECs) cell line) and revealed this method was nontoxic. Then, 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging test was done to assess the antioxidant properties, which indicated similar antioxidant potentials for TiNPs@Ziziphora and butylated hydroxytoluene. Agar diffusion tests were applied to determine the antibacterial and antifungal characteristics. Minimum Inhibitory Concentration (MIC), Minimum Bactericidal Concentration (MBC), and Minimum Fungicidal Concentration (MFC) were specified by macro-broth dilution assay. The data were analyzed by SPSS 21 software (Duncan post-hoc test). TiNPs@Ziziphora indicated higher antibacterial and antifungal effects than all standard antibiotics (p ≤ 0.01). Also, TiNPs@Ziziphora inhibited the growth of all bacteria at 2-16 mg/ml concentrations and removed them at 2-32 mg/ml concentrations (p ≤ 0.01). In case of antifungal properties of TiNPs@Ziziphora, they prevented the growth of all fungi at 2-8 mg/ml concentrations and destroyed them at 2-16 mg/ml concentrations (p ≤ 0.01). In vivo experiment, after creating the cutaneous wound, the rats were randomly divided into six groups: untreated control, treatment with Eucerin basal ointment, treatment with 3% tetracycline ointment, treatment with 0.2% TiO₂ ointment, treatment with 0.2% Z. clinopodioides ointment, and treatment with 0.2% TiNPs@Ziziphora ointment. These groups were treated for 10 days. For histopathological and biochemical analysis of the healing trend, a 3 × 3 cm section was prepared from all dermal thicknesses at day 10. Use of TiNPs@Ziziphora ointment in the treatment groups substantially reduced (p ≤ 0.01) the wound area, total cells, neutrophil, and lymphocyte and remarkably raised (p ≤ 0.01) the wound contracture, hydroxyl proline, hexosamine, hexuronic acid, fibrocyte, and fibrocytes/fibroblast rate compared to other groups. In conclusion, the results revealed the useful non-cytotoxic, antioxidant, antibacterial, antifungal, and cutaneous wound healing effects of TiNPs@Ziziphora.