For nearly a decade, researchers have debated the mechanisms by which AgNPs exert toxicity to bacteria and other organisms. The most elusive question has been whether the AgNPs exert direct "particle-specific" effects beyond the known antimicrobial activity of released silver ions (Ag(+)). Here, we infer that Ag(+) is the definitive molecular toxicant. We rule out direct particle-specific biological effects by showing the lack of toxicity of AgNPs when synthesized and tested under strictly anaerobic conditions that preclude Ag(0) oxidation and Ag(+) release. Furthermore, we demonstrate that the toxicity of various AgNPs (PEG- or PVP- coated, of three different sizes each) accurately follows the dose-response pattern of E. coli exposed to Ag(+) (added as AgNO(3)). Surprisingly, E. coli survival was stimulated by relatively low (sublethal) concentration of all tested AgNPs and AgNO(3) (at 3-8 μg/L Ag(+), or 12-31% of the minimum lethal concentration (MLC)), suggesting a hormetic response that would be counterproductive to antimicrobial applications. Overall, this work suggests that AgNP morphological properties known to affect antimicrobial activity are indirect effectors that primarily influence Ag(+) release. Accordingly, antibacterial activity could be controlled (and environmental impacts could be mitigated) by modulating Ag(+) release, possibly through manipulation of oxygen availability, particle size, shape, and/or type of coating.

The antibacterial activity of silver nanoparticles (AgNPs) is partially due to the release of Ag(+), although discerning the contribution of AgNPs vs Ag(+) is challenging due to their common co-occurrence. We discerned the toxicity of Ag(+) versus a commercially available AgNP (35.4 ± 5.1 nm, coated with amorphous carbon) by conducting antibacterial assays under anaerobic conditions that preclude Ag₀ oxidation, which is a prerequisite for Ag(+) release. These AgNPs were 20× less toxic to E. coli than Ag(+) (EC₅₀: 2.04 ± 0.07 vs 0.10 ± 0.01 mg/L), and their toxicity increased 2.3-fold after exposure to air for 0.5 h (EC₅₀: 0.87 ± 0.03 mg/L) which promoted Ag(+) release. No significant difference in Ag(+) toxicity was observed between anaerobic and aerobic conditions, which rules out oxidative stress by ROS as an important antibacterial mechanism for Ag(+). The toxicity of Ag(+) (2.94 μmol/L) was eliminated by equivalent cysteine or sulfide; the latter exceeded the solubility product equilibrium constant (K(sp)), which is conducive to silver precipitation. Equivalent chloride and phosphate concentrations also reduced Ag(+) toxicity without exceeding K(sp). Thus, some common ligands can hinder the bioavailability and mitigate the toxicity of Ag(+) at relatively low concentrations that do not induce silver precipitation. Furthermore, low concentrations of chloride (0.1 mg/L) mitigated the toxicity of Ag(+) but not that of AgNPs, suggesting that previous reports of higher AgNPs toxicity than their equivalent Ag(+) concentration might be due to the presence of common ligands that preferentially decrease the bioavailability and toxicity of Ag(+). Overall, these results show that the presence of O₂ or common ligands can differentially affect the toxicity of AgNPs vs Ag(+), and underscore the importance of water chemistry in the mode of action of AgNPs.

No abstract

No abstract