Laury Gauthier

Classification Framework for Graphene‐Based Materials

Graphing graphene: Because the naming of graphene-based materials (GBMs) has led to confusion and inconsistency, a classification approach is necessary. Three physical-chemical properties of GBMs have been defined by the GRAPHENE Flagship Project of the European Union for the unequivocal classification of these materials (see grid).

Characterisation and in vivo ecotoxicity evaluation of double-wall carbon nanotubes in larvae of the amphibian Xenopus laevis.

Because of their outstanding properties, carbon nanotubes (CNTs) are being assessed for inclusion in many manufactured products. Due to their massive production and growing number of potential applications, the impact of CNTs on the environment must be taken into consideration. The present investigation evaluates the ecotoxicological potential of double-walled carbon nanotubes (DWNTs) in the amphibian larvae Xenopus laevis at a large range of concentrations in water (from 10 to 500 mgL(-1)). Acute toxicity and genotoxicity were analysed after 12 days of static exposure in laboratory conditions. Acute toxicity was evaluated according to the mortality and the growth of larvae. The genotoxic effects were analysed by scoring the micronucleated erythrocytes of the circulating blood of larvae according to the International Standard micronucleus assay. Moreover, histological preparations of larval intestine were prepared after 12 days of exposure for observation using optical and transmission electron microscopy (TEM). Finally, the intestine of an exposed larva was prepared on a slide for analyse by Raman imaging. The results showed no genotoxicity in erythrocytes of larvae exposed to DWNTs in water, but acute toxicity at every concentration of DWNTs studied which was related to physical blockage of the gills and/or digestive tract. Indeed, black masses suggesting the presence of CNTs were observed inside the intestine using optical microscopy and TEM, and confirmed by Raman spectroscopy analysis. Assessing the risks of CNTs requires better understanding, especially including mechanistic and environmental investigations.

Environmentally relevant approaches to assess nanoparticles ecotoxicity: A review

Despite the increasing production and use of nanoparticles (NPs), there is a lack of knowledge about their environmental fate and ecotoxicity. Studies in environmentally relevant conditions are necessary to better assess these parameters, but such studies are rather rare. The present work represents first time that studies on engineered NPs using environmentally relevant exposure methods have been reviewed. These exposure methods differ from standardized protocols and can be classified into three groups: experimental trophic chains that allow study of the trophic route, multi-species exposures under laboratory conditions that allow for complex but controlled exposure and outdoor exposures that are more similar to environmentally realistic conditions. The majority of studies of micro- or mesocosms have focused on NP partitioning and bioaccumulation. The other major parameter that has been studied is NP ecotoxicity, which has been assessed in single species, in single species via the trophic route, and at the community level. The induction of biochemical defense systems, immunomodulation, effects on growth and reproduction, behavioral alterations and mortality have been used as indicators of major toxicity, depending on the species studied. The major effects of NPs on both microbial and algal communities include modifications of community compositions and diversities, decreased biomass and changes in community activities.

Mutagenic impact on fish of runoff events in agricultural areas in south-west France.

When heavy rainfall follows herbicide application, the intense surface runoff causes stream water contamination. Aquatic organisms are then briefly exposed to a complex mixture of contaminants. The aim of the present study is to investigate the genotoxic impact of such events on fish. A model fish, the Crucian carp (Carassius carassius) was exposed in controlled conditions, for 4 days, to water sampled daily in the Save River (France). The watershed of this stream is representative of agricultural areas in south-west France. Three hydrological conditions were compared: basal flow, winter flood, and spring flood. Chemical analysis of the water samples confirmed the higher contamination of the spring flood water, mainly explained by a peak of metolachlor. Genotoxicity was evaluated by micronucleus (MN) test and comet assay in peripheral erythrocytes. A significant increase in DNA breakdowns compared to controls was detected by the comet assay for all conditions. Exposure to spring flood water resulted in the highest damage induction. Moreover, induced chromosomal damage was only detected in this condition. In addition, fish were exposed, for 4 days, to an experimental mixture of 5 herbicides representative of the spring flood water contamination. Fish exhibited moderate DNA damage induction and no significant chromosomal damage. The mutagenicity induced by field-collected water is then suspected to be the result of numerous interactions between contaminants themselves and environmental factors, stressing the use of realistic exposure conditions. The results revealed a mutagenic impact of water contamination during the spring flood, emphasizing the need to consider these transient events in water quality monitoring programs.

Study of the genotoxic activity of six halogenated acetonitriles, using the SOS chromotest, the Ames-fluctuation test and the newt micronucleus test

Three short-term assays (the SOS chromotest, the Ames-fluctuation test and the newt micronucleus test) were carried out to evaluate the genotoxicity of six halogenated acetonitriles identified in chlorinated waters (monochloro-, dichloro-, trichloro-, monobromo-, dibromo- and bromochloroacetonitrile). With the SOS chromotest, three of the chemicals studied (dichloro-, dibromo- and bromochloroacetonitrile) were found to induce primary DNA damage in Escherichia coli PQ37. In the Ames-fluctuation test, all the compounds except dibromoacetonitrile showed mutagenic activity on Salmonella typhimurium strain TA100. The newt micronucleus assay detected a clastogenic effect on the peripheral blood erythrocytes of Pleurodeles waltl larvae for all the six haloacetonitriles studied. Moreover, two structure-activity relationships were noted: (1) the genotoxic activity of haloacetonitriles containing bromine substituents appeared higher than the corresponding chlorinated acetonitriles and (2) the clastogenic activity of the chlorinated acetonitriles increased with the number of chlorine substituents.

Carbon nanotube ecotoxicity in amphibians: assessment of multiwalled carbon nanotubes and comparison with double-walled carbon nanotubes

The potential impact of industrial multiwalled carbon nanotubes (MWNTs) was investigated under normalized laboratory conditions according to the International Standard micronucleus assay ISO 21427-1 for 12 days of half-static exposure to 0.1, 1, 10 and 50 mg/l of MWNTs in water. Three different end points were carried out for 12 days of exposure: mortality, growth inhibition and micronuclei induction in erythrocytes of the circulating blood of larvae. Raman spectroscopy analysis was used to study the presence of carbon nanotubes in the biological samples. Considering the high diversity of carbon nanotubes according to their different characteristics, MWNTs were analyzed in Xenopus larvae, comparatively to double-walled carbon nanotubes used in a previous study in similar conditions. Growth inhibition in larvae exposed to 50 mg/l of MWNTs was evidenced; however, no genetoxicity (micronucleus assay) was noticed, at any concentration. Carbon nanotube localization in the larvae leads to different possible hypothesis of mechanisms explaining toxicity in Xenopus.

In vivo detection of waste water and industrial effluent genotoxicity: use of the Newt Micronucleus test (Jaylet test)

The genotoxic potential of various waste waters has been evaluated in a micronucleus test using amphibian larvae. Genotoxicity was detected after dilution, in waste water from tanneries and from various petrochemical industries. Further studies have shown that sample treatment used for in vitro testing may affect the genotoxic response. Sterilization by gamma irradiation lowered genotoxic activity. Furthermore, microfiltration of effluent and extraction of organic micropollutants on XAD-4 resins, lead to the preparation of extracts which are not fully representative of the initial water sample. Testing of concentrates, as required for in vitro studies, will limit the scope of a survey to that part of the organic matter that can be recovered by concentration techniques. Many of the problems encountered in in vitro genotoxicity studies of waters, may be circumvented with direct testing on aquatic organisms. Thus, there is no need to concentrate or sterilise a sample. The tests can be carried out with intact animals, thus taking into account uptake and elimination, internal transport and metabolism. Finally, in vivo test-systems, such as the Newt Micronucleus Test, are more relevant to eukaryotes than bacterial assays and are suitable to assess the real impact of genotoxins discharged in the aquatic environment.

Evaluation of the genotoxic and teratogenic potential of a municipal sludge and sludge-amended soil using the amphibian Xenopus laevis and the tobacco: Nicotiana tabacum L. var. xanthi Dulieu

The toxic, genotoxic and teratogenic potential of a municipal sewage sludge was assessed using the micronucleus assay on the larvae of the amphibian Xenopus laevis and with the tobacco somatic mutation test using the yellow-green xanthi Dulieu mutant a(1)(+)/a(1) a(2)(+)/a(2). The teratogenic potential was assessed by means of the Frog Embryo Teratogenesis Assay-Xenopus (FETAX). Various doses of the pasty sludge added to a crop soil were tested using the three bioassays. The test systems were performed either directly with sludge or sludge-amended soil samples (plant model) or with aqueous extracts (aquatic animal model). Using the tobacco model, we found no mutagenic impact of the soil amended with the sludge, perhaps because the clay-like nature of the soil, with its high adsorption capacity, may have prevented the contaminants from reaching the target. All leachates of amended soils produced a significant size reduction in Xenopus embryos. Depending on the soil/sludge ratio, some leachates were found to be genotoxic but were never teratogenic. This battery of in vivo test systems enabled us to estimate the global long-term effects under agricultural conditions with various genetic endpoints on ecologically relevant organisms characteristic of the aquatic and terrestrial compartments.

International amphibian micronucleus standardized procedure (ISO 21427-1) for in vivo evaluation of double-walled carbon nanotubes toxicity and genotoxicity in water

Considering the important production of carbon nanotubes (CNTs), it is likely that some of them will contaminate the environment during each step of their life cycle. Nevertheless, there is little known about their potential ecotoxicity. Consequently, the impact of CNTs on the environment must be taken into consideration. This work evaluates the potential impact of well characterized double‐walled carbon nanotubes (DWNTs) in the amphibian larvae Xenopus laevis under normalized laboratory conditions according to the International Standard micronucleus assay ISO 21427‐1:2006 for 12 days of half‐static exposure to 0.1–1–10 and 50 mg L−1 of DWNTs in water. Two different endpoints were carried out: (i) toxicity (mortality and growth of larvae) and (ii) genotoxicity (induction of micronucleated erythrocytes). Moreover, intestine of larvae were analyzed using Raman spectroscopy. The DWNTs synthetized by catalytic chemical vapor deposition (CCVD) were used as produce (experiment I) and the addition of Gum Arabic (GA) was investigated to improve the stability of the aqueous suspensions (experiment II). The results show growth inhibition in larvae exposed to 10 and 50 mg L−1 of DWNTs with or without GA. No genotoxicity was evidenced in erythrocytes of larvae exposed to DWNTs, except to 1 mg L−1 of DWNTs with GA suggesting its potential effect in association with DWNTs at the first nonacutely toxic concentration. The Raman analysis confirmed the presence of DWNTs into the lumen of intestine but not in intestinal tissues and cells, nor in the circulating blood of exposed larvae.

Toxicity of CeO2 nanoparticles at different trophic levels--effects on diatoms, chironomids and amphibians.

The aim of the present work is to provide wider information on the toxicity of cerium dioxide nanoparticles (CeO2 NPs) in aquatic environments, by studying the toxicity of two types of CeO2 NPs for four species (diatoms Nitzschia palea, the sediment-dwelling invertebrate Chironomus riparius, and the amphibian larvae Xenopus laevis and Pleurodeles waltl.). The two types of CeO2 NPs have different intrinsic properties: some of them are small citrate-coated spheres (2-5 nm), and the others are larger uncoated plates (20-60 nm). Acute toxicity (mortality at 48 or 96 h, depending on the test-organism) was assessed for the four species, from 0.1 to 100 mg L(-1) of NPs. Sub-lethal effects were assessed on chironomids exposed between 0.01 and 1 mg L(-1) of NPs. Mortality, growth inhibition and genotoxic effects were evaluated on amphibian larvae from 0.1 to 10 mg L(-1). Results reveal that no acute toxicity occurs on any species after short exposures, even at the highest concentrations. Mortality (35%) is observed on Xenopus larvae after 12d of exposure at the highest concentration of one type of NPs. No significant effects were observed on chironomids during chronic exposure. Xenopus larvae growth was inhibited from 1 mg L(-1) of both NPs while growth inhibition is observed on Pleurodeles only at the highest concentration of one type of NPs. No genotoxicity was observed on Xenopus but Pleurodeles exhibited dose-dependent genotoxic effects when exposed to one type of NPs. Observed differences in toxicity are discussed focusing on the studied compartment, routes of exposure, species and NPs.