Capping Agent-Dependent Toxicity and Antimicrobial Activity of Silver Nanoparticles: An In Vitro Study. Concerns about Potential Application in Dental Practice

Abstract

Objectives: In dentistry, silver nanoparticles (AgNPs) have drawn particular attention because of their wide antimicrobial activity spectrum. However, controversial information on AgNPs toxicity limited their use in oral infections. Therefore, the aim of the present study was to evaluate the antibacterial activities against a panel of oral pathogenic bacteria and bacterial biofilms together with potential cytotoxic effects on human gingival fibroblasts of 10 nm AgNPs: non-functionalized - uncapped (AgNPs-UC) as well as surface-functionalized with capping agent: lipoic acid (AgNPs-LA), polyethylene glycol (AgNPs-PEG) or tannic acid (AgNPs-TA) using silver nitrate (AgNO₃) as control. Methods: The interaction of AgNPs with human gingival fibroblast cells (HGF-1) was evaluated using the mitochondrial metabolic potential assay (MTT). Antimicrobial activity of AgNPs was tested against anaerobic Gram-positive and Gram-negative bacteria isolated from patients with oral cavity and respiratory tract infections, and selected aerobic Staphylococci strains. Minimal inhibitory concentration (MIC) values were determined by the agar dilution method for anaerobic bacteria or broth microdilution method for reference Staphylococci strains and Streptococcus mutans. These strains were also used for antibiofilm activity of AgNPs. Results: The highest antimicrobial activities at nontoxic concentrations were observed for the uncapped AgNPs and the AgNPs capped with LA. It was found that AgNPs-LA and AgNPs-PEG demonstrated lower cytotoxicity as compared with the AgNPs-TA or AgNPs-UC in the gingival fibroblast model. All of the tested nanoparticles proved less toxic and demonstrated wider spectrum of antimicrobial activities than AgNO₃ solution. Additionally, AgNPs-LA eradicated Staphylococcus epidermidis and Streptococcus mutans 1-day biofilm at concentration nontoxic to oral cells. Conclusions: Our results proved that a capping agent had significant influence on the antibacterial, antibiofilm activity and cytotoxicity of AgNPs. Clinical significance: This study highlighted potential usefulness of AgNPs against oral anaerobic Gram-positive and Gram-negative bacterial infections and aerobic Staphylococci strains provided that pharmacological activity and risk assessment are carefully performed.

Keywords: antibacterial activity; antibiofilm activity; capping agent; cytotoxicity; human gingival fibroblasts; silver nanoparticles.

Figure 1

Characterization of AgNPs using transmission electron microscopy (TEM). The representative microscopy images show shape of AgNPs; the histograms illustrate the range of particle size distribution obtained from TEM measurements of more than 300 particles: (A) AgNPs capped with lipoic acid, (B) AgNPs capped with polyethylene glycol, (C) AgNPs capped with tannic acid and (D) uncapped AgNPs.

Figure 1

Characterization of AgNPs using transmission electron microscopy (TEM). The representative microscopy images show shape of AgNPs; the histograms illustrate the range of particle size distribution obtained from TEM measurements of more than 300 particles: (A) AgNPs capped with lipoic acid, (B) AgNPs capped with polyethylene glycol, (C) AgNPs capped with tannic acid and (D) uncapped AgNPs.

Figure 2

AgNPs-induced decrease in cell viability. The 24 h treatments of cells with AgNPs decreased HGF1 cell viability. Data are mean ± SD of 3-4 separate determinations. ***p < 0.001 as compared with control.

Figure 3

Susceptibility of 1-day biofilm (MBEC) formed by reference strains bacteria to AgNPs (µg/mL)