May. 13, 2024
Chemicals
We employed a yeast water extract to reduce silver ions and form silver nanoparticles (AgNPs), validated through UV–visible spectra. This process is marked by surface plasmon resonance (SPR) during nanoparticle formation, indicating specific electromagnetic absorption. Both samples exhibited a peak at 420 nm (Fig. 1A), consistent with previous findings for cell-free fungal extracts of Trichoderma and Aspergillus fumigatus. According to Mie's theory, a single SPR band signifies spherical metal nanoparticles. Transmission electron microscopy (TEM) confirmed their spherical morphology (Fig. 1B).
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Figure 1(A) UV–Vis absorption spectra of biosynthesized AgNPs_L (circle) and AgNPs_H (diamond). (B) Example of TEM image of AgNPs.
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Important physicochemical properties determine the cell uptake and fate of nanomaterials in bioapplications. We evaluated size, stability, and surface chemistry of biomanufactured AgNPs using dynamic light scattering (DLS). Synthesized AgNPs showed median sizes of 20.1 nm and 17.5 nm for AgNPs_L and AgNPs_H, respectively (Fig. 2). Stability was gauged using the polydispersity index (PdI), where higher values indicate less monosized particles. PdI values initially were 0.107 and 0.397, later 0.327 and 0.319 after eight months, proving excellent stability and minimal aggregation.
Figure 2Size of the AgNPs_L (A) and AgNPs_H (B) evaluated by hydrodynamic light scattering analysis. PdI, polydispersity index.
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TEM and energy dispersive X-ray spectroscopy (EDS) were deployed for morphology and elemental composition analysis. High-resolution TEM (HRTEM) micrographs revealed clear lattice fringes with interplanar spacings of 0.235 nm, confirming the silver phase. STEM-HAADF and EDS indicated sulfur presence, suggesting sulfur-rich yeast extract compounds act as stabilizers. Many studies align, noting that biologically synthesized nanoparticles often utilize sulfur or protein stabilizers over conventional synthetic methods that can pose health risks.
Figure 3(A) HRTEM images of a single silver nanoparticle with lattice fringes. (B, C) HAADF STEM image of AgNPs. (D–F) EDS elemental mapping images, with silver (red), sulfur (blue), and an overlay.
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Silver nanoparticles exhibit multifunctional antimicrobial action. We tested their efficacy against bacteria strains Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and the fungus Candida albicans using zone of inhibition assays. After 24 hours, significant bacterial growth inhibition was observed, but not for yeast strains. Previous studies have shown high antibacterial properties for AgNPs, but our findings contradict some reports on their antifungal efficacy against Candida albicans, possibly due to particle size and sulfur coatings.
Figure 4Antimicrobial activity of the AgNPs after 24 h of incubation against Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, and Candida albicans.
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Bacterial biofilms pose a challenge as they resist antimicrobial agents. Due to their minute size, AgNPs can disrupt these biofilms. We tested their efficacy with crystal violet staining on E. coli and P. aeruginosa. Results showed concentration-dependent biofilm inhibition, with significant reductions for higher AgNPs concentrations. Literature points to AgNPs’ ability to inhibit biofilm production and combat multidrug-resistant strains, resonating with our findings.
Figure 5Biofilm inhibition after treatment of AgNPs in E. coli (A) and P. aeruginosa (B). Surface-associated biofilm OD measurements are presented.
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The challenge of biofilm eradication in hospital infections calls for potent agents like AgNPs. We evaluated their efficiency against E. coli and P. aeruginosa biofilms. We observed significant biofilm biomass reduction, especially at 1 mg/mL and 2 mg/mL concentrations, displaying a dose-dependent eradication effect. Previous studies corroborate the importance of concentration in biofilm management using AgNPs.
Figure 6Biofilm eradication after treatment of AgNPs in E. coli (A) and P. aeruginosa (B). The degradation percentage of biofilm was calculated relative to untreated controls.
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MTT assays were used to evaluate AgNPs' effect on cell lines, focusing on mitochondrial activity. Various concentrations were tested on mouse embryonic fibroblasts (NIH 3T3), human keratinocytes (HaCaT), human osteosarcoma (U-2OS), and human non-small cell lung carcinoma (NCI-1299). Data revealed dose-dependent decreases in metabolic activity, particularly significant at 2 mg/mL. Notably, cancer cells exhibited higher resistance to oxidative stress induced by AgNPs, suggesting different cellular responses among normal and cancer cells.
Figure 7Cell metabolic activity of the NIH 3T3 (A), HaCaT (B), U-2OS (C), and NCI-1299 (D) after 24 hours exposure to various AgNPs concentrations. Results indicate notable differences in metabolic activity reduction.
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Typically, one method is used to report nanomaterial cytotoxicity70-74, potentially skewing results as cells show activity even during apoptosis75. We used AO/EB staining and MTT assays to assess live/dead cell ratios and metabolic activities. At 1 mg/mL AgNPs, dead cells significantly increased across cell lines (Fig. 8A-D). In contrast, lower concentrations had minimal toxicity, confirming the safety range for further applications. Our results align with literature emphasizing nanomaterial type and stabilizers regarding cytotoxic effects.
Figure 8Cell viability of the NIH3T3 (A), HaCaT (B), U2OS (C), and NCI-1299 (D) after exposure to AgNPs. Staining and imaging confirm live (green) and dead (red) cells. Graphs represent percentage of dead cells.
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Cancer cell migration and invasion critically affect disease progression. We conducted scratch assays on human keratinocytes and osteosarcoma cells to see if AgNPs influence these processes. Untreated control cells closed scratches within 48 hours, while those exposed to AgNPs showed inhibited migration. Higher impact was seen in cancer cells, highlighting AgNPs' potential in metastasis inhibition.
Figure 9Cell migration ability in response to AgNPs. Human keratinocytes (A) and osteosarcoma cells (B) at different intervals (0, 24, 48 hours) showed varying migratory capacities post-exposure.
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Photomicrographs show silver nanoparticles covering the root canal walls (Fig. 10A) and penetrating dentin around 20 µm deep (Fig. 10B), crucial for deep bactericidal action. Nanoparticle agglomerates can obstruct bacterial return from dentin to the macrocanal, enhancing bactericidal effects.
Figure 10The wall of the macrocanal covered with silver nanoparticles (A) and the micrograp of the penetration of silver nanoparticles into the dentinal canal (B) with agglomerates of nanoparticles (inset).
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Utilizing post-culturing water for silver NP synthesis offers an efficient, cost-effective method relevant for industrial-scale production. These AgNPs are beneficial for various applications, from antimicrobial coatings to targeted drug delivery, combating cancer cells more effectively. This method provides a productive approach to sustainable and versatile nanomaterial manufacturing.
Antimicrobial packaging is being developed to extend the shelf-life and safety of foods and beverages. There are concerns about transferring potentially harmful materials, such as silver nanoparticles, into consumables. Researchers reporting in ACS Applied Materials & Interfaces have found that silver embedded in antimicrobial plastics can exit the material and form nanoparticles in foods and beverages, particularly in sweetened ones.
Polymers containing nanoparticles or nanocomposites can inhibit microorganism growth responsible for spoilage and illness. Although not yet approved for packaging in the U.S., researchers are exploring nanoparticle-embedded polymers for future use. Previous studies showed leaching into water-based food simulants, but little is known about interactions with real foods. Sugars can convert silver ions into nanoparticles upon ingestion, prompting Timothy Duncan and colleagues to study how these ingredients form nanoparticles, during direct exposure or storage in silver-laced packaging.
To explore silver aggregation in edible mixtures, researchers spiked liquid foods, including sugary solutions, soda, milk, juices, yogurt, and starch-based slurry with silver, incubating them at 104 F for over ten days to simulate prolonged storage. Nanostructures were detected in various concentrations. Sugary liquids with citrates, starches, and fats had most nanoparticles. In contrast, acidic liquids initially formed aggregates that later dissolved. Further experiments with small packets of silver-laced polyethylene polymer showed similar findings, with sugars maintaining leaching and nanoparticle creation during prolonged storage. Thus, dietary exposure to silver nanoparticles from sweetened foods packaged in antimicrobial materials is feasible under typical long-term storage conditions.
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