Influence of gastrointestinal environment on free radical generation of silver nanoparticles and implications for their cytotoxicity

With the increasing number of applications of silver nanoparticles (Ag NPs) in consumer products, including food contact applications, it is important to understand potential health effects of ingestion of Ag NPs. The biosafety analysis of Ag NPs in various mammalian cells has been widely studied; however, the influence of the gastrointestinal environment on the physicochemical properties and toxicity of Ag NPs has not been fully addressed. In the present study, we investigated the impact of simulated human gastrointestinal tract (GIT) fluids on the physicochemical properties of Ag NPs and their impact on the in vitro cytotoxicity of Ag NPs in intestinal epithelial (Caco-2) cells. Polyvinylpyrrolidone (PVP)-coated Ag NPs of different sizes (30, 50, 100 nm) were incubated in three GIT fluids that differed in composition and pH (1.6, 5.0, 6.5) and were designed to mimic human gastric fluid in a fasted state (FaSSGF), human intestinal fluid in a fasted state (FaSSIF), and human intestinal fluid in a fed state (FeSSIF). Time-dependent decreases in UV-Vis absorption and increases in dynamic light scattering (DLS) were observed during the three-hour incubation of Ag NPs in GIT fluids. The results of transmission electron microscopy (TEM) coupled with energy dispersive X-ray spectroscopy (EDS) suggested that the Ag NPs aggregated and precipitated as AgCl salts in the presence of FaSSGF and FeSSIF. In the presence of FaSSGF and hydrogen peroxide (H2O2), a particle size- and concentration-dependent generation of hydroxyl radicals (%OH) was detected by electron spin resonance (ESR) spectroscopy. When compared to the effects observed with the incubation of Ag NPs in H2O, incubation of Ag NPs in FaSSGF showed decreased cellular uptake in Caco-2 cells. In contrast, the incubation of Ag NPs in FeSSIF demonstrated increased cellular uptake and cytotoxicity. The results of this study demonstrate that GIT fluids have a significant influence on the physicochemical properties and cytotoxicity of Ag NPs and they provide valuable information that may be used in the safety evaluation of Ag NPs for consumer products.