Size dependent bioaccumulation and ecotoxicity of gold nanoparticles in an endobenthic invertebrate: The Tellinid clam Scrobicularia plana
Highlights
► AuNPs were aggregated to a maximum distribution peak at 0.6 μm in seawater. ► Scrobicularia plana accumulated Au with higher accumulation for bigger NPs. ► Induction of MT, CAT, SOD and GST showed oxidative stress in clams. ► TBARS levels advocated no significant oxidative damage in clams. ► Gold is an inert metal, ecotoxic effects are probably due to the nanoparticle form.
Introduction
Nanotechnology is a rapidly emerging discipline with a variety of applications. In nanomaterials, the exponential increase in the ratio of surface-to-bulk atoms with decreasing particle size results in greater activity per unit mass compared to identical bulk materials (Oberdörster et al., 2005). AuNPs are of special interest because of their functions in medicine and therapeutics (reviewed in Kunzmann et al., 2011), in electronics, catalysis, cosmetic, and food industries (Daniel and Astruc, 2003). Widespread use of NPs will likely lead to their increased release into the environment (Klaine et al., 2008), especially in coastal environments that are close to human populations, and potentially impact organisms through the release of their component (e.g. ions for metal-containing NPs) or through one of the most common effect of oxidative stress caused by their inherent mechanical, catalytic, optical properties, and electrical conductivity (Klaine et al., 2008, Johnston et al., 2011, Moore, 2006, Nel et al., 2006). AuNPs have been shown to induce reactive oxygen species (ROS) production with (Pan et al., 2007, Pan et al., 2009, Tedesco et al., 2010b) or without (Farkas et al., 2010) consequent toxicity. The large surface area and high surface reactivity of NPs is likely to induce oxidative stress (Tedesco et al., 2010b), thus even particles made of low toxicity material like polystyrene can be deleterious in nano scales (Brown et al., 2001), and this toxicity may be size-dependent (Johnston et al., 2011, Pan et al., 2007).
Catalase (CAT), glutathione S-transferase (GST) and superoxide dismutase (SOD) have been proposed as markers involved in antioxidant systems of defense in various aquatic species such as bivalves (Van der Oost et al., 2005). Metallothioneins are a useful tool to reveal the presence of bioavailable metals (reviewed in Amiard et al., 2006), and may be involved in both metal detoxification and antioxidant defense (Viarengo et al., 1999). Defense against the stress caused by contaminants is energy consuming, and lactate dehydrogenase (LDH) activity is particularly important when a considerable amount of energy is rapidly required (Diamantino et al., 2001) under adverse conditions. When the defense mechanisms in organisms are not sufficient to cope with oxidative stress, changes in the concentrations of thiobarbituric acid reactive substances (TBARS) are able to reflect the state of lipid peroxidation of the membranes (Knight et al., 1988). In addition, behavioral biomarkers (burrowing, feeding rate) are sensitive tools to assess the impact of contaminants at concentrations far below lethal levels (Amiard-Triquet, 2009), as well-documented in the bivalve mollusk Scrobicularia plana (Bonnard et al., 2009, Solé et al., 2009). To date the exact mechanisms concerning the in vivo toxicity of NPs (including AuNPs), and the link between biomarkers at different levels of biological organization (e.g. molecular vs. individual) remain poorly understood.
Sediments are the final sink for most contaminants, including NPs. Ferry et al. (2009) showed recently that Au nanorods passed from the water column to sediment (concentration factor was 33.1 at 12 d) and the marine food web (e.g. in clam concentration factor was 2.28 × 104 at 12 d) using laboratory-constructed estuarine mesocosms. The aquatic benthic bivalve S. plana is a key species in the structure and functioning of estuarine ecosystems and a major link in estuarine food webs, playing an important role in biogeochemical cycles of both nutrients and contaminants as a consequence of its bioturbation activity. It has been shown to be a relevant model organism for the determination of biomarkers (Bonnard et al., 2009, Solé et al., 2009) and was successfully adopted in our recent nanotoxicity studies on CuO NPs (Buffet et al., 2011). In the present study, we carried out in vivo evaluation of biological responses of S. plana to AuNPs of three sizes (5, 15, and 40 nm). The fate of AuNPs of the three sizes tested was characterized in stock solution and experimental media (seawater with or without the presence of bivalves). In this aim, complementary methods were used to assess size distribution, zeta potential and purity. Animal metabolic condition (LDH activities) and the bioaccumulation of Au from NPs, together with tiered biomarker sets of defense (MT, CAT, GST, and SOD) and damage (TBARS and AChE activity) as well as behavioral biomarkers (postexposure feeding and burrowing) were adopted.
Section snippets
Gold nanoparticle characterization
AuNPs of three different sizes (5, 15 and 40 nm) were prepared at JRC (Joint Research Centre, Ispra, Italy) as a suspension of 98.5 mg L−1 in citrate buffer, stored at 20 °C for experiments for one week at the maximum. The particle size distribution was initially examine by dynamic light scattering at high concentration of Au NPs (nearing 100 mg L−1) See supplementary material (Fig. S1). At present, Disc Centrifuge Photosedimentation (CPS) was used allowing to test lower concentration (model
AuNPs aggregation and distribution in exposure medium
The nanosize distribution determined by TEM is depicted in Fig 1(a–c). Particle sizes are in agreement with nominal sizes (5, 15, 40 nm AuNP) as indicated by the scale bar of 50 nm. Fig. 1(d) shows the presence of Au as indicated by the three peaks corresponding to the gold M shell (2.2 keV, L shell (9.7 keV) and X shell (10.8 keV).
The results obtained by CPS at the two concentrations tested (1 and 10 mg L−1), both in the presence and absence of bivalves, were qualitatively similar and for clarity the
Aggregation and bioaccumulation of AuNPs
The nominal size of AuNPs and their purity were verified by using concomitantly TEM, EDS as well CPS in pure water. For a better understanding of the toxicity effect of NPs, it is important to know the particle behavior in exposure medium (Card and Magnuson, 2010). In seawater NPs lose their electrostatic stabilization due to the charge screening by the large amount of NaCl (Chinnapongse et al., 2011). In agreement with these findings, our results indicate that in seawater AuNPs (5, 15, 40 nm)
Conclusion
In summary, our findings may give a better insight into the process of nanometal bioaccumulation and toxic effects in the invertebrates from estuarine environments. There are urgent needs to better understand the nanotoxicity considering the extremely expanding use of nanoscale materials. Au NPs are generally considered non toxic like bulk gold, which is inert and biocompatible. However, gold that is physiologically inert at microscale has been shown to have biological activity at the nanoscale
Acknowledgements
This work was supported by grants no. [214478] from the European Community’s Seventh Framework Programme (FP7/2007-2013), and by a post-doctoral scholarship from the Fondation Franco-Chinoise pour la Science et ses Applications (FFCSA), China Scholarship Council (CSC) and the Région Pays de la Loire, France, while the first author visiting the University of Nantes.
References (49)
- et al.
Metallothioneins in aquatic invertebrates: their role in metal detoxification and their use as biomarkers
Aquatic Toxicology
(2006) - et al.
Honeybee Apis mellifera acetylcholinesterase – a biomarker to detect deltamethrin exposure
Ecotoxicology and Environmental Safety
(2008) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding
Analytical Biochemistry
(1976)- et al.
Size-dependent proinflammatory effects of ultrafine polystyrene particles: a role for surface area and oxidative stress in the enhanced activity of ultrafines
Toxicology and Applied Pharmacology
(2001) - et al.
Microsomal lipid peroxidation
Methods in Enzymology
(1978) - et al.
Behavioural and biochemical responses of two marine invertebrates Scrobicularia plana and Hediste diversicolor to copper oxide nanoparticles
Chemosphere
(2011) - et al.
A. Persistence of singly dispersed silver nanoparticles in natural freshwaters, synthetic seawater, and simulated estuarine waters
Science of Total Environment
(2011) - et al.
Evaluation of sublethal biomarkers in Litopenaeus vannamei on foodborne exposure to methyl parathion
Ecotoxicology and Environmental Safety
(2005) - et al.
Effects of copper and cadmium on cholinesterase and glutathione S-transferase activities of two marine gastropods (Monodonta lineata and Nucella lapillus)
Comparative Biochemistry and Physiology – Part C: Toxicology
(2007) - et al.
Lactate dehydrogenase activity as an effect criterion in toxicity tests with Daphnia magna straus
Chemosphere
(2001)
Effects of silver and gold nanoparticles on rainbow trout (Oncorhynchus mykiss) hepatocytes
Aquatic Toxicology
Semi-automated colorimetric and enzymatic assays for aquatic organisms using microplate readers
Water Research
Glutathione s-transferases
Journal of Biological Chemistry
Toxicology of engineered nanomaterials: focus on biocompatibility, biodistribution and biodegradation
Biochimica and Biophysica Acta
An enzymatic function for erythrocuprein
Journal of Biological Chemistry
Do nanoparticles present ecotoxicological risks for the health of the aquatic environment?
Environment International
Occurrence, behavior and effects of nanoparticles in the environment
Environmental Pollution
Evaluation of dietary microalgae for culture of the calanoid copepod Pseudodiaptomus pelagicus
Aquaculture
Carbohydrate digestion in the bivalve Scrobicularia plana (da costa)
Comparative Biochemistry and Physiology – Part B: Biochemistry
Increased acetylcholinesterase activities in specimens of Sparus auratus exposed to sublethal copper concentrations
Chemico-Biological Interactions
Pollution biomarkers in two estuarine invertebrates, Nereis diversicolor and Scrobicularia plana, from a marsh ecosystem in sw spain
Environment International
Gold nanoparticles and oxidative stress in Mytilus edulis
Marine Environmental Research
Exposure of the blue mussel, Mytilus edulis, to gold nanoparticles and the pro-oxidant menadione
Comparative Biochemistry and Physiology – Part C: Toxicology & Pharmacology
Oxidative stress and toxicity of gold nanoparticles in Mytilus edulis
Aquatic Toxicology
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