Objective To test the silver ion release kinetics of three kinds of silver nanoparticles under different conditions (such as mass concentration, time, and solution) and assess the cytotoxicity of released silver ions.
Methods Uncoated 20nm, uncoated 50nm, and polyvinylpyrrolidone coated 20nm silver nanoparticles (nano-Ag20, nano-Ag50, and nano-Ag20PVP) were dispersed in different pH buffers such as pH3.6 acetate buffer, pH6.0 acetate buffer, and pH7.0 deionized water to prepare 0.2, 0.5, and 1.0 g/L suspensions. After incubation at 37℃, silver ions were separated by ultracentrifugation and ultrafiltration for detection. Nano-Ag20PVP were dissolved in cell culture to prepare solutions of 0, 20, 40, 80, and 160mg/L concentrations, then incubated at 37℃, and followed by silver ion detection after 24 and 48h, respectively. A549 and HepG2 cells were exposed to nanoAg20PVP suspensions, with silver nitrate solution used as positive control, to determine cell viability by MTT assay.
Results Higher degree of dispersion was found in nano-Ag20PVP than in nano-Ag20 or nano-Ag50. In all designed pH conditions, the silver ion content of three kinds of silver nanoparticles was positively correlated with mass concentration and time. Under same pH and mass concentration conditions, the silver ion ratio of nano-Ag20PVP was always higher than that of nano-Ag20 (P < 0.05). At pH7.0, the silver ion ratio of nano-Ag50 was higher than that of nano-Ag20 (P < 0.05) in the 1.0 g/L group. The silver ion ratios of the 20 mg/L and the 160 mg/L nano-Ag20PVP solutions were 0.02% and 0.09%, respectively. After incubation for 24 h and 48h, the silver ion ratios of the 160mg/L nano-Ag20PVP were 0.01% and 0.02%, respectively. After treatment for 24h and 48h with nanoAg20PVP, both silver content and silver ion ratio in A549 and HepG2 cells were positively associated with the mass concentration of nano-Ag20PVP. After 24 h with nano-Ag20PVP treatment, the content of silver in HepG2 cells was higher than that in A549 cells (P < 0.05), but it was lower after 48 h (P < 0.05). After treatment for 24 h and 48 h with nano-Ag20PVP, the contents of silver ion in HepG2 cells were lower than those in A549 cells (P < 0.05). After treatment for 24 h and 48 h with 10-160 mg/L nano-Ag20PVP, the cell viabilities of HepG2 cells were higher than those of A549 cells (P < 0.05). After treatment for 24 h and 48 h with 5 and 10 mg/L silver nitrate, the cell viabilities of HepG2 cells were higher than those of A549 cells (P < 0.05). At the same mass concentration (5 and 10 mg/L), the cell viability of the nano-Ag20PVP group was higher than that of the silver nitrate group (P < 0.05).
Conclusion Coated silver nanoparticles release more silver ions than uncoated silver nanoparticles. Nano-Ag20PVP could still release silver ions in cells, which plays an important role in the cytotoxicity induced by silver ions in selected cell lines. Silver nanoparticles and released silver ions jointly induce cytotoxicity.
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