Effects of nano-scale TiO2, ZnO and their bulk counterparts on zebrafish: Acute toxicity, oxidative stress and oxidative damage
Research Highlights
► Toxicity and oxidative effects of nano and bulk ZnO and TiO2 on zebrafish. ► Toxicity mechanism of ZnO and TiO2 NPs differed. ► Activities of diverse NPs in various tissues differed. ► Metal ions released by nano and bulk ZnO contributed partly to toxicity. ► Extracellular *OH generated by nano-TiO2 hurts gill cell membranes directly.
Introduction
Nano-scale (diameters between 100 and 1 nm) titanium dioxide and zinc oxide, which have broad UV spectrum attenuation properties, are used in a variety of applications, including sunscreen (Maier and Korting, 2005), wastewater treatment (Chen et al., 2004), and environmental remediation (Aitken et al., 2006). Direct and indirect release of these nanoparticles (NPs) into aquatic environments via bathing, sewage effluent (Daughton and Ternes, 1999, Handy and Shaw, 2007, Ternes et al., 2004) and other engineering applications (Chen et al., 2004, Nagaveni et al., 2004) have increased the exposure chances of humans and ecosystems to NPs (Nowack and Bucheli, 2007). Consequently, the potential impacts of TiO2 and ZnO NPs on aquatic ecosystems have attracted special attention due to the unique properties of NPs (Handy and Shaw, 2007, Handy et al., 2008, Oberdörster et al., 2005, Wiench et al., 2009).
Generally, the toxicity of NPs is associated with their small size and high specific surface area (Lovern and Klaper, 2006). Nanomaterials are theoretically expected to be more toxic than their bulk counterparts due to their greater surface reactivity and the ability to penetrate into and accumulate within cells and organisms (Carlson et al., 2008, Ispas et al., 2009, Mironava et al., 2010). However, investigations of aggregations of NPs with size distributions similar to those of their bulk particles in suspension have revealed that the toxicity mechanisms of NPs are more complex (Warheit et al., 2006, Zhang et al., 2008). In suspensions, NPs tend to form large particles and most of the aggregates will settle out of the suspensions in a few hours, which may reduce their toxicities (Adams et al., 2006). On the other hand, reactive oxidative species (ROS) generated through various mechanisms, such as illumination of NPs (Jiang et al., 2008) and disruption of intra-cellular metabolic activities (Long et al., 2006) may disturb the anti-oxidant system (Brown et al., 2004), leading to damage to lipids, carbohydrates, proteins and DNA (Kelly et al., 1998). Previous studies have shown that TiO2 NPs induced oxidative stress in the brains of rainbow trout (Oncorhynchus mykiss) (Federici et al., 2007). ZnO NPs participated in damaging the membrane of Escherichia coli through ROS mechanisms, thus exhibiting an antibacterial action (L. Zhang et al., 2007). Another crucial factor that may affect the toxicity of metal oxide nanoparticles is the release of metal ions. It has been shown previously that the toxicity of nZnO was mainly attributed to the dissolved Zn ions (Aruoja et al., 2009, Franklin et al., 2007). However, contradictory results have been noted in recent studies, suggesting the possibility for a size dependence of toxicity, distinct from adverse effects associated with the presence of dissolved ions (Nair et al., 2009, Wong et al., 2010).
Although there have been a number of studies on the potential hazards of TiO2 and ZnO NPs to aquatic ecosystems, their environmental impacts on aquatic organisms and toxicity mechanisms still have not been fully elucidated. Particularly, the toxicity tests on adult fish have mainly focused on carbon-based NPs (Oberdörster, 2004, Smith et al., 2007, Zhu et al., 2006). Moreover, those investigations of the toxicity of metal oxide NPs (TiO2, ZnO NPs) to fish have concentrated on early developmental stages (Zhu et al., 2008, Zhu et al., 2009). Few researchers have touched the toxicity of TiO2 and ZnO NPs to adult fish. Based on the fact that zebrafish represents a bridge between in vitro cell culture models and in vivo mammalian models (Fako and Furgeson, 2009), the objective of this study was to evaluate the acute toxicity and oxidative effects of ZnO and TiO2 NPs on adult zebrafish, especially the effects of particle formulations (nano or bulk), ·OH radicals generated by NPs and metal ions released from metal oxide particles.
Section snippets
Preparation and characterization of particle suspensions
TiO2 and ZnO NPs were purchased from the Nano Applied Research Center of Nanjing University of Technology. The surfaces of TiO2 and ZnO NPs were not modified. Bulk TiO2, ZnO and ZnSO4·7 H2O were purchased from Tianjin Guangcheng Chemical Reagent Co., Ltd. The purities, diameters, crystal structures and zeta potentials of the NPs and bulk particles were listed in Table 1.
Suspensions of nano-scale ZnO and TiO2 and their counterparts were prepared with aerated single-distilled water and dispersed
Characteristics of NPs and bulk particles in suspension
The TEM images and size distributions of NPs and bulk particles are shown in Fig. 1 and Table 2. In suspensions of TiO2 and ZnO NPs and of bulk TiO2 and ZnO, particles were collected into aggregates of irregular shape (Fig. 1). The mean size of the NP aggregates increased with increasing concentrations (Table 2). As shown in Fig. 1, the initial diameter of individual NPs (about 30 nm as provided by the suppliers) was much smaller than the corresponding bulk particles. However, the measured
Acute toxicity of TiO2 NPs, ZnO NPs and bulk particles to zebrafish
Although the particle size analysis indicated that the average diameters of TiO2 were similar to their bulk particles in suspension, as noted in other studies (Adams et al., 2006, Long et al., 2006), the acute toxicity of TiO2 NPs (96-h LC50 of 124.5 mg/L) was far higher than that of bulk TiO2, which produced 0% mortality in zebrafish even at a concentration of 300 mg/L. Similarly, the toxicities of TiO2 NPs to the nematode Caenorhabditis elegans (Wang et al., 2009) and to D. magna (Zhu et al.,
Conclusions
From the results of this study, it can be concluded that: (1) the acute toxicity of TiO2 NPs to zebrafish was significantly higher than that of bulk TiO2 while zinc oxides were equally toxic in NP and bulk formulations; (2) NPs could cause toxic effects despite that they formed aggregates in suspensions; (3) Metal ions released by ZnO contributed to toxicities but were not the main lethal mechanism of the ZnO NPs and bulk ZnO suspensions tested; (4) TiO2 NPs were able to cause toxicity effects
Acknowledgment
This research was supported by the Key Project of the National Science and Technology Program (2008ZX07103001)and the National Natural Science Foundation of China (No. 21037004).
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