Elsevier

Science of The Total Environment

Volume 565, 15 September 2016, Pages 951-960
Science of The Total Environment

Effect of chromium oxide (III) nanoparticles on the production of reactive oxygen species and photosystem II activity in the green alga Chlamydomonas reinhardtii

https://doi.org/10.1016/j.scitotenv.2016.01.028Get rights and content

Highlights

  • Cr2O3 nanoparticles are unstable and form large aggregates in the medium.

  • EC50 for growth inhibition of C. reinhardtii is 1.35 g L 1 at 72 h.

  • Cr2O3 nanoparticles increase ROS levels at 10 g L 1.

  • Cr2O3 nanoparticles affect photosynthetic electron transport.

Abstract

With the growth of nanotechnology and widespread use of nanomaterials, there is an increasing risk of environmental contamination by nanomaterials. However, the potential implications of such environmental contamination are hard to evaluate since the toxicity of nanomaterials if often not well characterized. The objective of this study was to evaluate the toxicity of a chromium-based nanoparticle, Cr2O3-NP, used in a wide diversity of industrial processes and commercial products, on the unicellular green alga Chlamydomonas reinhardtii. The deleterious impacts of Cr2O3-NP were characterized using cell density measurements, production of reactive oxygen species (ROS), esterase enzymes activity, and photosystem II electron transport as indicators of toxicity. Cr2O3-NP exposure inhibited culture growth and significantly lowered cellular Chlorophyll a content. From cell density measurements, EC50 values of 2.05 ± 0.20 and 1.35 ± 0.06 g L 1 Cr2O3-NP were obtained after 24 and 72 h of exposure, respectively. In addition, ROS levels were increased to 160.24 ± 2.47% and 59.91 ± 0.15% of the control value after 24 and 72 h of exposition to 10 g L 1 Cr2O3-NP. At 24 h of exposure, the esterase activity increased to 160.24% of control value, revealing a modification of the short-term metabolic response of algae to Cr2O3-NP exposure. In conclusion, the metabolism of C. reinhardtii was the most sensitive to Cr2O3-NP after 24 h of treatment.

Introduction

Due to their high surface area and reactivity, chromium oxide materials, such as chromium(III) oxide (Cr2O3), are used in several applications of commercial or industrial interests. These range from refractory and ceramic applications to oxidation reaction catalysts, sintering agents, pigments, coatings for thermal protection, and food supplements (Wang et al., 2000, Munoz et al., 2004, Kocon et al., 2005, Zha et al., 2009). Large particles with sizes near 200 nm are preferred to produce pigments due to their higher opacity and coating power, while smaller nanoscale particles (NP) are used due to their magnetic properties or to improve sintering (Balachandran et al., 1995). The reactive properties of Cr2O3-NP can also be used in the catalytic reactions of hydrogen sulfide (H2S) (Uhm et al., 1999) and sulfur dioxide (SO2), the dehydration and dehydrogenation of alcohol, for methanol synthesis, to break double bonds in olefins, and in ethylene polymerization or chlorofluorocarbon synthesis (Uhm et al., 1999, Santulli et al., 2011).

With the growth of nanotechnology and the widespread use of nanomaterials in consumer products, the risks of environmental contamination by chromium-based nanomaterials are increasing. Once present in the ecosystems, these materials can interact with aquatic life to induce toxic effects. The toxicity of nanomaterials is known to be dependent on their physico-chemical properties, which influence their environmental biotransformation, biological transport, and cellular interactions (Dhawan et al., 2009, Perreault et al., 2010). However, the toxicological risks of nanomaterials remain difficult to evaluate because the mechanisms causing their toxicity are still debated. The toxicity of some metallic NPs is enhanced at the nanoscale because a reduced particle size leads to the enhanced solubilization of NPs into toxic metal ions (Perreault et al., 2014a, Perreault et al., 2014b, Navarro et al., 2008, Thit et al., 2013). On the other hand, particle specific effects were also observed due to the reactive nature of nanoscale particles (Bondarenko et al., 2013, Perreault et al., 2014a, Perreault et al., 2014b). However, for Cr2O3-NPs, the different mechanisms involved are mainly unknown due to the limited amount of information on the toxicity of nanoscale chromium-based nanomaterials.

For chromium nanomaterials, toxicity will be dependent on the oxidation state of the chromium ions released from the particles. The most toxic form of chromium is the hexavalent form, a strong oxidant that can lead to cytotoxicity, genotoxicity and oxidative stress (World Health Organization (WHO), 1988, De Flora and Wetterhahn, 1989, De Flora et al., 1990, Syracuse Research Corporation, 1993, Losi et al., 1994, Costa, 1997, USEPA: Toxicological Review of hexavalent chromium, 1998). The higher toxicity of hexavalent chromium is also due to its tetrahedral configuration, analogous to sulfate and phosphates ions. Because of this similarity, the cell membrane does not differentiate Cr(VI) from essential ions, which results in cellular internalization. When Cr(VI) enters the cytoplasm, it is reduced to Cr(III) in reaction with glutathione peptides (GSH). The Cr-GSH complex diffuses into the cytoplasm and connects to DNA, causing damages that lead to genetic changes (Labra et al., 2007, Benite et al., 2007, Swaminathan et al., 2009). In photosynthetic organisms, chromium-induced oxidative stress causes structural damage in chloroplasts, alters photosynthetic enzymatic functions, and inhibits photosynthetic processes (Dayan and Paine, 2001, Panda and Khan, 2003, Choudhury and Panda, 2005).

In this report, the toxicity potential of Cr2O3-NPs in aquatic environments was characterized using the green microalga Chlamydomonas reinhardtii. This algal species is an excellent ecotoxicological model for aquatic ecosystems since microalgae are very sensitive to contaminants, respond rapidly to environmental stresses, and represent the main source of biomass production in aquatic ecosystems (Merchant et al., 2007, Perreault et al., 2010, Melegari et al., 2013). The effects of Cr2O3-NP exposure on cell viability, ROS production, and cell morphology of C. reinhardtii were evaluated over the course of 72 h of exposition. Chlorophyll a fluorescence was used to determine the change in photosynthetic electron transport processed induced by Cr2O3-NP in microalgae. These different indicators of Cr2O3-NP toxicity were used to characterize the level of sensitivity of photosynthetic organisms in the environment in case of environmental contamination by chromium-based nanomaterials.

Section snippets

Algal culture

The C. reinhardtii culture (wild type, CC-125) used in this study was obtained from the Chlamydomonas Genetic Center (Duke University, Durham, NC, USA). Cells were cultivated in batch cultures containing 1 L of High Salt Medium — HSM (Harris, 2009). All chemicals employed were obtained from Sigma-Aldrich (St. Louis, MO, USA). Cells were grown under continuous illumination (100 μmol of photons m 2 s 1) using white fluorescent lamps (Sylvania Grolux F 36 W, Havells Sylvania Europe Ltd., London, UK) at

Characterization of Cr2O3-NPs

The Cr2O3-NP suspension was characterized in HSM medium and ultrapure water (UW). The zeta potential and hydrodynamic diameter were measured at a concentration of 0.1 g L 1. The hydrodynamic diameter and potential zeta results are shown in Table 1.

When dispersed in the HSM medium used for assays, the hydrodynamic diameter and zeta potential values of Cr2O3-NPs was significantly changed compared to UW dispersions (p < 0.05). Higher absolute values of zeta potential (> 30.0) may indicate that the

Discussion

Previous studies have demonstrated that Cr2O3-NP exposure results in a decrease in cell viability and induces oxidative stress and cell apoptosis in bacteria and mammalian cell cultures (Horie et al., 2011, Singh et al., 2011, Khatoon et al., 2011). In addition, ionic Cr(III) toxicity was characterized in different algal species. Vignati et al. (2010) evaluated the toxicity of (Cr(NO3)3·9H2O) in the green alga Pseudokirchneriella subcapitata (now Raphidocelis subcapitata) and found an average

Conclusions

The interactions between Cr2O3-NP and the algal culture medium significantly affected the outcomes of the toxicity tests. The results of this study demonstrated that the viability of C. reinhardtii was affected by an increase in the Cr2O3-NP concentration after 24 h of exposure. These differences no longer occurred after 48 h, which indicate that this species may adjust to Cr2O3-NP exposure. Inhibition of photosynthetic activity was highest at the highest Cr2O3-NP concentrations tested. Our

Conflict of interest

The authors declare that they have no financial relationships or conflicts of interest with the commercial identities or companies cited on this manuscript.

Acknowledgments

This work was supported by research grants awarded by the CAPES (Proc. no. 017/2010) and CNPq (Proc. no. 552112/2011-9) (Brazil). The authors thank the technical staff at the Faculty for Electron Microscopy Research at McGill University.

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