Elsevier

Environmental Pollution

Volume 159, Issue 10, October 2011, Pages 2713-2719
Environmental Pollution

Chronic toxicity of ZnO nanoparticles, non-nano ZnO and ZnCl2 to Folsomia candida (Collembola) in relation to bioavailability in soil

https://doi.org/10.1016/j.envpol.2011.05.021Get rights and content

Abstract

The chronic toxicity of zinc oxide nanoparticles (ZnO-NP) to Folsomia candida was determined in natural soil. To unravel the contribution of particle size and free zinc to NP toxicity, non-nano ZnO and ZnCl2 were also tested. Zinc concentrations in pore water increased with increasing soil concentrations, with Freundlich sorption constants Kf of 61.7, 106 and 96.4 l/kg (n = 1.50, 1.34 and 0.42) for ZnO-NP, non-nano ZnO and ZnCl2 respectively. Survival of F. candida was not affected by ZnO-NP and non-nano ZnO at concentrations up to 6400 mg Zn/kg d.w. Reproduction was dose-dependently reduced with 28-d EC50s of 1964, 1591 and 298 mg Zn/kg d.w. for ZnO-NP, non-nano ZnO and ZnCl2, respectively. The difference in EC50s based on measured pore water concentrations was small (7.94–16.8 mg Zn/l). We conclude that zinc ions released from NP determine the observed toxic effects rather than ZnO particle size.

Highlights

► ZnO nanoparticles and non-nano ZnO were equally toxic to Folsomia candida in soil. ► Pore water from soil spiked with ZnO nanoparticles showed saturation with zinc suggesting aggregation. ► Pore water based EC50 values for ZnO nanoparticles and ZnCl2 were similar. ► ZnO nanoparticle toxicity in soil was most probably due to Zn dissolution from the nanoparticles.

Introduction

In recent years, there is increasing interest in the environmental risks posed by engineered nanoparticles (NP), particles with at least one dimension of less than 100 nm and differing from non-nano material in physico–chemical properties, such as surface area and charge density (Handy et al., 2008). Zinc oxide, one of the most commonly used types of metal-based NP, is primarily used in electronics, personal care products and other applications requiring UV protection like sunscreens. Zinc oxide nanoparticles (ZnO-NP) can enter the environment via waste water at industrial sites or through domestic sewage from showering or swimming. NP can be transported to soil via sewage sludge, which is used for land application (e.g. fertilizer). Gottschalk et al. (2009) reported this to be the main route of soil exposure and estimated a predicted environmental concentration (PEC) of 0.093 μg/kg/y for European soils based on an annual production volume of 9845 tonnes; in case of sludge application on land, PEC is estimated at 3.25 μg/kg/y.

Consequently organisms living in the soil, such as earthworms, springtails, nematodes and isopods, may be harmed. Ecotoxicological studies with NP have mainly been performed with aquatic organisms (Heinlaan et al., 2008, Zhu et al., 2008, Wiench et al., 2009). A small but growing number of studies on the toxicity of NP to soil organisms has been published in the last two years (Peralta-Videa et al., 2011), assessing effects of short-term NP exposure to the isopod Porcellio scaber (Drobne et al., 2009, Jemec et al., 2008), the nematode Caenorhabditis elegans (Ma et al., 2009, Wang et al., 2009), and the earthworms Eisenia fetida (Hu et al., 2010, Unrine et al., 2010) and Lumbricus rubellus (Lapied et al., 2010, Lapied et al., 2011, Van der Ploeg et al., 2011). In addition, more research is needed to provide insight into the ecotoxicological effects of chronic exposure to NP on organisms living in soil.

In case of metal-based NP like ZnO, TiO2, Ag and CeO2, toxicity is at least partly due to the specific properties related to the small size and consequent high surface activity of NP, while effects may be further enhanced by the release of free metal ions (Auffan et al., 2009). If the free ions are more toxic than the original particles, this process of dissolution is likely to lead to an increase of the overall toxicity. Franklin et al. (2007) showed that the toxicity of ZnO-NP to the micro-algae Pseudokirchneriella subcapitata was attributable to dissolved zinc, while others could not explain the toxicity for the nematode C. elegans adequately by dissolution of the particles alone (Wang et al., 2009). Speciation of metal NP in soils is not yet understood. Given the well-known toxicity of the ionic forms of some metals, the solubility of metal-based NP may require particular attention. Dissolution of ZnO-NP has been observed in moderately hard reconstituted water to yield Zn concentrations of approx. 0.4 mg/l (Poynton et al., 2011). In soil, sorption is also an important factor that needs to be taken into account when performing toxicity or bioaccumulation tests. This depends strongly on soil characteristics such as organic matter content, cation exchange capacity and pH.

Due to the complex behaviour of NP in soil, realising realistic exposure in ecotoxicity testing poses major challenges. Soil by definition is heterogeneous, which requires conscientious introduction of any test compound and frequently sub-samples are analyzed to ensure that spiked portions of soil are homogeneous. In general, manufactured NP are considered to be insoluble in water and tend to aggregate, making it a difficult task to obtain homogeneity of NP distribution in spiked soil. Depending on their physical and chemical properties as well as soil properties, NP tend to form aggregates and are likely to settle within a relatively short time (Klaine et al., 2008). Currently used spiking procedures may need to be adapted with respect to preparation of the soil to be spiked, introduction of NP to the prepared soil and mixing to evenly distribute NP throughout the soil.

Springtails (Collembola) are common and widespread arthropods that are abundant in soils throughout the world. They represent ecologically and relevant test species, because they are an integral part of soil ecosystems and are vulnerable to the effects of soil contamination. The cuticle and ventral tube (diameter approx. 5 μm) are important exposure routes for chemicals dissolved in soil pore water (Fountain and Hopkin, 2005). Folsomia candida, a parthenogenetic species, was chosen because of its relatively short generation time and the ease of culturing. The fact that there is only one limit-test with ZnO-NP and springtails yet published in the literature (Manzo et al., 2011) indicates the need for further research on these organisms.

The purpose of the present study was to determine the chronic toxicity of ZnO-NP to F. candida, by studying survival and reproduction of the springtail as effect parameters and to unravel the contribution of zinc oxide particle size and free zinc to NP toxicity. To help elucidate toxic effects of ZnO-NP in soil, two reference compounds, non-nano ZnO and ZnCl2 were studied for comparison. We hypothesized that ZnO-NP is more toxic than non-nano ZnO due to its larger specific surface area and that the release of free zinc from ZnO-NP, rather than the NP itself, is responsible for toxic effects.

Section snippets

Test compounds

ZnO-NP powder, with a reported diameter size of <200 nm, was purchased from BASF (Z-COTE®). Powders were coated with carbon and photos of the particles were taken using a (field emission) scanning electron microscope (JEOL JSM-6301F). Non-nano ZnO (Merck, pro analysi, >99%) and ZnCl2 (Merck, zinc chloride pure) were used for comparison. Fig. 1 shows that the diameter size of ZnO-NP powder is at the nano-scale (i.e. <100 nm), although the fraction <100 nm has not been established. The diameter

Zinc distribution in the spiking solution

Fig. 2 shows transmission electron micrographs of Lufa 2.2 soil solution without ZnO (A), with non-nano ZnO (B) and ZnO-NP (C, D). Fig. 2C visualizes the binding of ZnO-NP to organic matter and shows a homogenous zinc distribution in the sample. Free ZnO nanoparticles, as shown on the SEM pictures (Fig. 1), were not observed. Non-nano ZnO was present as bigger particles and the binding to organic matter was less evident than for ZnO-NP (Fig. 2B).

Zinc concentrations and pH

The results of the zinc measurements in the soil

Characterization of NP in the spiking solution

To date, very little work has been conducted on the identification of engineered NP in soil. The accurate detection of NP in soils requires their separation from natural soil solids (Lead and Wilkinson, 2006). In this study, transmission electron microscopy (TEM) was applied to visualize ZnO-NP in the soil solution (approx. 1000 mg Zn/l) used to spike the soils. Introduction of NP to soil in a filtered soil extract was chosen to prevent agglomeration or flocculation as particles bind to natural

Acknowledgement

The authors thank Saskia Kars and Rien Dekker for performing the electron microscopy.

The work reported here was conducted in the context of NanoFATE, Collaborative Project CP-FP 247739 (2010-2014) under the 7th Framework Programme of the European Commission (FP7-NMP-ENV-2009, Theme 4), coordinated by C. Svendsen and D. Spurgeon of NERC – Centre for Ecology and Hydrology, UK-Wallingford; www.nanofate.eu.

BASF SE (Ludwigshafen, Germany) kindly provided Z-COTE® (nano-scale zinc oxide) as test

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