Therapeutic Window of Ligand‐Free Silver Nanoparticles in Agar‐Embedded and Colloidal State: In Vitro Bactericidal Effects and Cytotoxicity

Abstract

The inhibition of bacterial growth through effective non‐toxic antimicrobial substances is of great importance for the prevention and therapy of implant infections in various medical disciplines. For the evaluation of a therapeutic window of silver nanoparticles (AgNPs), their bactericidal properties were tested in agar composites and colloids on four medical relevant bacteria. Therefore, we produced AgNPs using high‐power nanosecond laser ablation in water showing a log‐normal particle diameter distribution centered at 17 nm. Bacteria were incubated with AgNP concentrations ranging from 5 to 70 µg · mL−1 and the growth rate was recorded. Additionally, cytotoxic effects of AgNPs on human gingival fibroblasts were examined. The experiments demonstrated that laser‐synthesized AgNPs resulted in a significant bacterial growth inhibition of more than 80% at the indicated concentrations in a solid agar model (Pseudomonas aeruginosa 10 µg · mL−1, Streptococcus salivarius 10 µg · mL−1, Escherichia coli 20 µg · mL−1, Staphylococcus aureus 70 µg · mL−1). In a planktonic bacteria model, the growth of the tested bacteria was significantly delayed by the addition of AgNPs at a concentration of 35 µg · mL−1. The cytotoxic assays showed limited adverse effects on human fibroblasts at concentrations of less than 20 µg · mL−1. The present study illustrates the strong antibacterial effects of ligand‐free, laser‐generated AgNPs that exhibit moderate cytotoxic effects, resulting in a therapeutically applicable concentration of AgNPs for medical purposes between 10 and 20 µg · mL−1.

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“…So, taking into account the higher water swelling behaviour of Lyocell and bacteria cellulose we expect that larger silver nanoparticles are released into the elution medium, which will increase the overall silver concentration but not the cytotoxic or antibacterial effects. Investigations by Grade et al [8] using laser generated silver nanoparticles with sizes of about 17 nm showed much lower cytotoxic effects compared to our results and had no significant effect at concentration values below 35 mg/l on human gingival fibroblasts. Other groups like Hidalgo et al, which exposed human dermal fibroblasts at different concentrations of silver nitrate, found a cytotoxic potential for concentration values greater than 1.37 mg/l of silver nitrate (with 10 % of FCS) [7].…”

“…The concentration that was found for pure laser generated silver nanoparticles show an antibacterial effect against S. aureus in planktonic and solid agar models for silver concentrations above 35 mg/l [8]. Another group around Lkhahavajav et al found lower minimum inhibitory concentrations (MIC) in order of 4 mg/l for K. pneumoniae and S. aureus with silver nanoparticles sizes from 20 to 45 nm, generated by a sol-gel method [19].…”

“…Nevertheless, a therapeutic range was described by some reviewers for different kind of silver salts and silver nanoparticles [6,7]. So, the cytotoxic effect depends on the shape, dose and size of particles and the type of cells which were used for the investigation [7][8][9]. Other factors like the concentration of albumin in solid agar may reduce the positive and negative effects of AgNP [10].…”

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“…Note that the nanoparticles are immobilized in the agar, so the mass concentration is more relevant than the number concentration. The mass concentration range in the agar gel was selected based on earlier experiences [11]. After 24 h incubation, colony forming units were counted on four plates.…”

“…Silver nanoparticles used in antibacterial implant coatings and surfaces are often embedded in composites or hydrogels, which release the silver ions over time to the adjacent area [8,9]. Unfortunately, high concentrations of antibacterial silver ions also exhibit cytotoxic effects on eukaryotic cells, making it difficult to establish a possible therapeutic window [10][11][12]. The cytotoxic effects of silver nanoparticles on eukaryotic cells are described in many studies and the particle concentrations, which cause at least moderate cytotoxicity depend very much on the type of cells used [13][14][15][16].…”