Aquatic Mesocosm Strategies for the Environmental Fate and Risk Assessment of Engineered Nanomaterials.

In the past decade, mesocosms have emerged as a useful tool for the environmental study of engineered nanomaterials (ENMs) as they can mimic the relevant exposure scenario of contamination. Herein, we analyzed the scientific outcomes of aquatic mesocosm experiments, with regard to their designs, the ENMs tested, and the end points investigated. Several mesocosm designs were consistently applied in the past decade to virtually mimic various contamination scenarios with regard to ecosystem setting as well as ENMs class, dose, and dosing. Statistical analyses were carried out with the literature data to identify the main parameters driving ENM distribution in the mesocosms and the potential risk posed to benthic and planktonic communities as well as global ecosystem responses. These analyses showed that at the end of the exposure, mesocosm size (water volume), experiment duration, and location indoor/outdoor had major roles in defining the ENMs/metal partitioning. Moreover, a higher exposure of the benthic communities is often observed but did not necessarily translate to a higher risk due to the lower hazard posed by transformed ENMs in the sediments (e.g., aggregated, sulfidized). However, planktonic organisms were generally exposed to lower concentrations of potentially more reactive and toxic ENM species. Hence, mesocosms can be complementary tools to existing standard operational procedures for regulatory purposes and environmental fate and risk assessment of ENMs. To date, the research was markedly unbalanced toward the investigation of metal-based ENMs compared to metalloid- and carbon-based ENMs but also nanoenabled products. Future studies are expected to fill this gap, with special regard to high production volume and potentially hazardous ENMs. Finally, to take full advantage of mesocosms, future studies must be carefully planned to incorporate interdisciplinary approaches and ensure that the large data sets produced are fully exploited.

Nanoparticle Uptake in Plants: Gold Nanomaterial Localized in Roots of Arabidopsis thaliana by X-ray Computed Nanotomography and Hyperspectral Imaging.

Terrestrial plants can internalize and translocate nanoparticles (NPs). However, direct evidence for the processes driving the NP uptake and distribution in plants is scarce at the cellular level. Here, NP-root interactions were investigated after 10 days of exposure of Arabidopsis thaliana to 10 mg·L-1 of negatively or positively charged gold NPs (∼12 nm) in gels. Two complementary imaging tools were used: X-ray computed nanotomography (nano-CT) and enhanced dark-field microscopy combined with hyperspectral imaging (DF-HSI). The use of these emerging techniques improved our ability to detect and visualize NP in plant tissue: by spectral confirmation via DF-HSI, and in three dimensions via nano-CT. The resulting imaging provides direct evidence that detaching border-like cells (i.e., sheets of border cells detaching from the root) and associated mucilage can accumulate and trap NPs irrespective of particle charge. On the contrary, border cells on the root cap behaved in a charge-specific fashion: positively charged NPs induced a higher mucilage production and adsorbed to it, which prevented translocation into the root tissue. Negatively charged NPs did not adsorb to the mucilage and were able to translocate into the apoplast. These observations provide direct mechanistic insight into NP-plant interactions, and reveal the important function of border cells and mucilage in interactions of plants with charged NPs.

Remote Biodegradation of Ge-Imogolite Nanotubes Controlled by the Iron Homeostasis of Pseudomonas brassicacearum.

The toxicity of high-aspect-ratio nanomaterials (HARNs) is often associated with oxidative stress. The essential nutrient Fe may also be responsible of oxidative stress through the production of reactive oxygen species. In the present study, it has been examined to what extent adding Fenton reaction promoting Fe impacted the toxicity of an alumino-germanate model HARN. Structural addition of only 0.95% wt Fe to Ge-imogolite not only alleviated the toxicity observed in the case of Fe-free nanotubes but also stimulated bacterial growth. This was attributed to the metabolization of siderophore-mobilized Fe from the nanotube structure. This was evidenced by the regulation of the homeostasis-monitoring intracellular Fe levels. This was accompanied by a biodegradation of the nanotubes approaching 40%, whereas the Fe-free nanomaterial remained nearly untouched.

Transformation of pristine and citrate-functionalized CeO2 nanoparticles in a laboratory-scale activated sludge reactor.

Engineered nanomaterials (ENMs) are used to enhance the properties of many manufactured products and technologies. Increased use of ENMs will inevitably lead to their release into the environment. An important route of exposure is through the waste stream, where ENMs will enter wastewater treatment plants (WWTPs), undergo transformations, and be discharged with treated effluent or biosolids. To better understand the fate of a common ENM in WWTPs, experiments with laboratory-scale activated sludge reactors and pristine and citrate-functionalized CeO2 nanoparticles (NPs) were conducted. Greater than 90% of the CeO2 introduced was observed to associate with biosolids. This association was accompanied by reduction of the Ce(IV) NPs to Ce(III). After 5 weeks in the reactor, 44 ± 4% reduction was observed for the pristine NPs and 31 ± 3% for the citrate-functionalized NPs, illustrating surface functionality dependence. Thermodynamic arguments suggest that the likely Ce(III) phase generated would be Ce2S3. This study indicates that the majority of CeO2 NPs (>90% by mass) entering WWTPs will be associated with the solid phase, and a significant portion will be present as Ce(III). At maximum, 10% of the CeO2 will remain in the effluent and be discharged as a Ce(IV) phase, governed by cerianite (CeO2).

Influence of the length of imogolite-like nanotubes on their cytotoxicity and genotoxicity toward human dermal cells.

Physical-chemical parameters such as purity, structure, chemistry, length, and aspect ratio of nanoparticles (NPs) are linked to their toxicity. Here, synthetic imogolite-like nanotubes with a set chemical composition but various sizes and shapes were used as models to investigate the influence of these physical parameters on the cyto- and genotoxicity and cellular uptake of NPs. The NPs were characterized using X-ray diffraction (XRD), small angle X-ray scattering (SAXS), and atomic force microscopy (AFM). Imogolite precursors (PR, ca. 5 nm curved platelets), as well as short tubes (ST, ca. 6 nm) and long tubes (LT, ca. 50 nm), remained stable in the cell culture medium. Internalization into human fibroblasts was observed only for the small particles PR and ST. None of the tested particles induced a significant cytotoxicity up to a concentration of 10(-1) mg·mL(-1). However, small sized NPs (PR and ST) were found to be genotoxic at very low concentration 10(-6) mg·mL(-1), while LT particles exhibited a weak genotoxicity. Our results indicate that small size NPs (PR, ST) were able to induce primary lesions of DNA at very low concentrations and that this DNA damage was exclusively induced by oxidative stress. The higher aspect ratio LT particles exhibited a weaker genotoxicity, where oxidative stress is a minor factor, and the likely involvement of other mechanisms. Moreover, a relationship among cell uptake, particle aspect ratio, and DNA damage of NPs was observed.

Is there a Trojan-horse effect during magnetic nanoparticles and metalloid cocontamination of human dermal fibroblasts?

This study investigates the issue of nanoparticles/pollutants cocontamination. By combining viability assays, physicochemical and structural analysis (to probe the As speciation and valence), we assessed how γFe(2)O(3) nanoparticles can affect the cytotoxicity, the intra- and extracellular speciation of As(III). Human dermal fibroblasts were contaminated with γFe(2)O(3) nanoparticles and As(III) considering two scenarios: (i) a simultaneous coinjection of the nanoparticles and As, and (ii) an injection of the nanoparticles after 24 h of As adsorption in water. In both scenarios, we did not notice significant changes on the nanoparticles surface charge (zeta potential ∼ -10 mV) nor hydrodynamic diameters (∼950 nm) after 24 h. We demonstrated that the coinjection of γFe(2)O(3) nanoparticles and As in the cellular media strongly affects the complexation of the intracellular As with thiol groups. This significantly increases at low doses the cytotoxicity of the As nonadsorbed at the surface of the nanoparticles. However, once As is adsorbed at the surface the desorption is very weak in the culture medium. This fraction of As strongly adsorbed at the surface is significantly less cytotoxic than As itself. On the basis of our data and the thermodynamics, we demonstrated that any disturbance of the biotransformation mechanisms by the nanoparticles (i.e., surface complexation of thiol groups with the iron atoms) is likely to be responsible for the increase of the As adverse effects at low doses.

Growth kinetic of single and double-walled aluminogermanate imogolite-like nanotubes: an experimental and modeling approach.

Atomic Force Microscopy (AFM) and in situ Small Angle X-ray Scattering (SAXS) were used to investigate the evolution of the aluminogermanate imogolite-like nanotubes concentration and morphology during their synthesis. In particular, in situ SAXS allowed quantifying the transformation of protoimogolite into nanotubes. The size distribution of the final nanotubes was also assessed after growth by AFM. A particular attention was focused on the determination of the single and double walled nanotube length distributions. We observed that the two nanotube types do not grow with the same kinetic and that their final length distribution was different. A model of protoimogolites oriented aggregation was constructed to account for the experimental growth kinetic and the length distribution differences.

Evidence of double-walled Al-Ge imogolite-like nanotubes. a cryo-TEM and SAXS investigation.

It has been recently discovered that the synthesis of Al-Ge imogolite-like nanotubes is possible at high concentration. Despite this initial success, the structure of these Al-Ge imogolite-like nanotubes remains not completely understood. Using high resolution cryo-TEM and Small Angle X-ray Scattering, we unravel their mesoscale structure in two contrasted situations. On the one hand, Al-Ge imogolite nanotubes synthesized at 0.25 M are double-walled nanotubes of 4.0 +/- 0.1 nm with an inner tube of 2.4 +/- 0.1 nm. Moreover, SAXS data also suggest that the two concentric tubes have an equal length and identical wall structure. On the other hand, at higher concentration (0.5M), both SAXS and cryo-TEM data confirm the formation of single-walled nanotubes of 3.5 +/- 0.15 nm. Infrared spectroscopy confirms the imogolite structure of the tubes. This is the first evidence of any double-walled imogolite or imogolite-like nanotubes likely to renew interest in these materials and associated potential applications.

Investigation of copper speciation in pig slurry by a multitechnique approach.

It is now well-known that copper (Cu) can accumulate on the surface of soils upon which pig slurry has been applied. This is due to the high quantity of Cu in pig slurry resulting from its use as a growth promoter in animal feeds. The mobility and bioavailability of Cu from pig slurry spreading can be better predicted by determining the speciation of this element in addition to its total concentration. The aim of this study was to present a multitechnique approach to investigate Cu speciation in pig slurry. First, size fractionation and chemical characterization of each size fraction were performed to complement results obtained in raw samples. Micro X-ray fluorescence spectroscopy (µXRF) highlighted the colocalization of Cu and sulfur (S). Finally, X-ray absorption near-edge structure spectroscopy (XANES) showed that Cu speciation in raw pig slurry and size fractions could be described by Cu(2)S and that its oxidation state is Cu(I). In addition, geochemical calculation demonstrated that chalcocite (Cu(2)S) was the major Cu species present under pig slurry lagoon physical-chemical conditions. This Cu speciation in pig slurry may be the main reason for the observed Cu accumulation at the soil surface.

Aqueous aging of a silica coated TiO2 UV filter used in sunscreens: investigations at the molecular scale with dynamic nuclear polarization NMR.

Short-term, aqueous aging of a commercial nanocomposite TiO2 UV filter with a protective SiO2 shell was examined in abiotic simulated fresh- and seawater. Under these conditions, the SiO2 layer was quantitatively removed (∼88-98%) within 96 hours, as determined using inductively coupled plasma-atomic emission spectroscopy (ICP-AES). While these bulk ICP-AES analyses suggested almost identical SiO2 shell degradation after aging in fresh- and seawater, surface sensitive 29Si dynamic nuclear polarization (DNP) solid-state nuclear magnetic resonance (SSNMR), with signal enhancements of 5-10× compared to standard SSNMR, was able to distinguish differences in the aged nanocomposites at the molecular level. DNP-SSNMR revealed that the attachment of the silica layer to the underlying TiO2 core rested on substantial Si-O-Ti bond formation, bonds which were preserved after freshwater aging, yet barely present after aging in seawater. The removal of the protective SiO2 layer is due to ionic strength accelerated dissolution, which could present significant consequences to aqueous environments when the photoactive TiO2 core becomes exposed. This work demonstrates the importance of characterizing aged nanocomposites not only on the bulk scale, but also on the molecular level by employing surface sensitive techniques, such as DNP-NMR. Molecular level details on surface transformation and elemental speciation will be crucial for improving the environmental safety of nanocomposites.

Comparison of Nanomaterials for Delivery of Double-Stranded RNA in Caenorhabditis elegans.

RNA interference is a promising crop protection technology that has seen rapid development in the past several years. Here, we investigated polyamino acid biopolymers, inorganic nanomaterials, and hybrid organic-inorganic nanomaterials for delivery of dsRNA and efficacy of gene knockdown using the model nematode Caenorhabditis elegans. Using an oral route of delivery, we are able to approximate how nanomaterials will be delivered in the environment. Of the materials investigated, only Mg-Al layered double-hydroxide nanoparticles were effective at gene knockdown in C. elegans, reducing marker gene expression to 66.8% of that of the control at the lowest tested concentration. In addition, we identified previously unreported injuries to the mouthparts of C. elegans associated with the use of a common cell-penetrating peptide, poly-l-arginine. Our results will allow the pursuit of further research into promising materials for dsRNA delivery and also allow for the exclusion of those with little efficacy or deleterious effects.

Synthesis of imogolite fibers from decimolar concentration at low temperature and ambient pressure: a promising route for inexpensive nanotubes.

To date, the successful low-temperature synthesis of the aluminosilicate imogolite nanotubes always involved initial concentrations of the reagents in the millimolar range, higher concentrations being reported to lead to the formation of the less well characterized allophane phase. The present work shows that reaction kinetics and not initial concentration control the formation of the nanotubes: substantial amounts of well formed imogolite were obtained from a decimolar initial concentration, i.e. 100 times higher than the "standard" protocol. The allophane-like spheroids expected from the high reagent concentration were not observed in this work, and proto-imogolite was obtained instead.

Spectroscopic characterization of organic matter of a soil and vinasse mixture during aerobic or anaerobic incubation.

Mineralization potentials are often used to classify organic wastes. These methods involve measuring CO(2) production during batch experiments, so variations in chemical compounds are not addressed. Moreover, the physicochemical conditions are not monitored during the reactions. The present study was designed to address these deficiencies. Incubations of a mixture of soil and waste (vinasse at 20% dry matter from a fermentation industry) were conducted in aerobic and anaerobic conditions, and liquid samples obtained by centrifugation were collected at 2h, 1d and 28 d. Dissolved organic carbon (DOC) patterns highlighted that: there was a "soil effect" which increased organic matter (OM) degradation in all conditions compared to vinasse incubated alone; and OM degradation was faster under aerobic conditions since 500 mgkg(-1) of C remained after aerobic incubation, as compared to 4000 mgkg(-1) at the end of the anaerobic incubation period. No changes were detected by Fourier transform infrared spectroscopy (FTIR) between 2h and 1d incubation. At 28 days incubation, the FTIR signal of the aerobic samples was deeply modified, thus confirming the high OM degradation. Under anaerobic conditions, the main polysaccharide contributions (nu(C-O)) disappeared at 1000 and 1200 cm(-1), as also confirmed by the (13)C NMR findings. Under aerobic incubation, a 50% decrease in the polysaccharide proportion was observed. Under anaerobic conditions, significant chemical modifications of the organic fraction were detected, namely formation of low molecular weight organic acids.

Monitoring the Environmental Aging of Nanomaterials: An Opportunity for Mesocosm Testing?

Traditional aging protocols typically examine only the effects of a limited number of stresses, and relatively harsh conditions may trigger degradation mechanisms that are not observed in actual situations. Environmental aging is, in essence, the complex interaction of multiple mechanical, physicochemical and biological stresses. As yet, there is no (pre)standardized procedure that addresses this issue in a satisfactory manner. Mesocosm experiments can be designed to specifically cover the aging of nanomaterials while characterizing the associated exposure and hazard. The scenario of exposure and the life time of the nanomaterial appear as the predominant factors in the design of the experiment, and appropriate precautions need to be taken. This should the subject of guidance that may be divided into product/application categories.

Synthesis of large quantities of single-walled aluminogermanate nanotube.

A simple aqueous synthesis yielded about 100 times more structurally well-organized single-walled aluminogermanate nanotubes than previously reported "standard" procedures. The structure analyses using XRD, IRTF, TEM, and XAS were greatly facilitated by the high concentrations available, and they ascertained the imogolite-like structure of the nanotubes. Simplicity and yield of the synthesis protocol are likely to favor commercial applications of theses materials as well as simplified syntheses of other nanophases.

Non-linear release dynamics for a CeO2 nanomaterial embedded in a protective wood stain, due to matrix photo-degradation.

The release of CeO2-bearing residues during the weathering of an acrylic stain enriched with CeO2 nanomaterial designed for wood protection (Nanobyk brand additive) was studied under two different scenarios: (i) a standard 12-weeks weathering protocol in climate chamber, that combined condensation, water spraying and UV-visible irradiation and (ii) an alternative accelerated 2-weeks leaching batch assay relying on the same weathering factors (water and UV), but with a higher intensity of radiation and immersion phases. Similar Ce released amounts were evidenced for both scenarios following two phases: one related to the removal of loosely bound material with a relatively limited release, and the other resulting from the degradation of the stain, where major release occurred. A non-linear evolution of the release with the UV dose was evidenced for the second phase. No stabilization of Ce emissions was reached at the end of the experiments. The two weathering tests led to different estimates of long-term Ce releases, and different degradations of the stain. Finally, the photo-degradations of the nanocomposite, the pure acrylic stains and the Nanobyk additive were compared. The incorporation of Nanobyk into the acrylic matrix significantly modified the response of the acrylic stain to weathering.

Dynamic Nuclear Polarization NMR as a new tool to investigate the nature of organic compounds occluded in plant silica particles.

The determination of the chemical nature of the organic matter associated with phytoliths remains a challenge. This difficulty mainly stems from amounts of organic carbon (C) that are often well below the detection limit of traditional spectroscopic tools. Conventional solid-state 13C Nuclear Magnetic Resonance (NMR) is widely used to examine the nature and structure of organic molecules, but its inherent low sensitivity prohibits the observation of diluted samples. The recent advent of commercial microwave source in the terahertz range triggered a renewed interest in the Dynamic Nuclear Polarization (DNP) technique to improve the signal to noise ratio of solid-state NMR experiments. With this technique, the 13C spectrum of a phytolith sample containing 0.1% w/w C was obtained overnight with sufficient quality to permit a semi-quantitative analysis of the organic matter, showing the presence of peptides and carbohydrates as predominant compounds. Considering the natural abundance of the 13C isotope, this experiment demonstrates that DNP NMR is sufficiently sensitive to observe spin systems present in amounts as low as a few tens of ppm.

Stealth Biocompatible Si-Based Nanoparticles for Biomedical Applications.

A challenge regarding the design of nanocarriers for drug delivery is to prevent their recognition by the immune system. To improve the blood residence time and prevent their capture by organs, nanoparticles can be designed with stealth properties using polymeric coating. In this study, we focused on the influence of surface modification with polyethylene glycol and/or mannose on the stealth behavior of porous silicon nanoparticles (pSiNP, ~200 nm). In vivo biodistribution of pSiNPs formulations were evaluated in mice 5 h after intravenous injection. Results indicated that the distribution in the organs was surface functionalization-dependent. Pristine pSiNPs and PEGylated pSiNPs were distributed mainly in the liver and spleen, while mannose-functionalized pSiNPs escaped capture by the spleen, and had higher blood retention. The most efficient stealth behavior was observed with PEGylated pSiNPs anchored with mannose that were the most excreted in urine at 5 h. The biodegradation kinetics evaluated in vitro were in agreement with these in vivo observations. The biocompatibility of the pristine and functionalized pSiNPs was confirmed in vitro on human cell lines and in vivo by cytotoxic and systemic inflammation investigations, respectively. With their biocompatibility, biodegradability, and stealth properties, the pSiNPs functionalized with mannose and PEG show promising potential for biomedical applications.

Involvement of nitrogen functional groups in high-affinity copper binding in tomato and wheat root apoplasts: spectroscopic and thermodynamic evidence.

Carboxylic groups located in plant cell walls (CW) are generally considered to be the main copper binding sites in plant roots, despite the presence of other functional groups. The aim of this study was to investigate sites responsible for copper binding in root apoplasts, i.e. CW and outer surface of the plasma membrane (PM) continuum. Binding sites in root apoplasts were investigated by comparing isolated CW of a monocotyledon (Triticum aestivum L.) and dicotyledon (Solanum lycopersicum L.) crop with their respective whole roots. Copper speciation was examined by X-ray absorption (XAS) and (13)C-nuclear magnetic resonance spectroscopies while the affinity of ligands involved in copper binding was investigated by modeling copper sorption isotherms. Homogeneous speciation and binding of copper was found in wheat and tomato root apoplasts. Only Cu-N and Cu-O bonds were detected in wheat and tomato root apoplasts. Nitrogen/oxygen ligands were identified in slightly higher proportions (40-70%) than single oxygen ligands. Furthermore, low- and high-affinity binding sites contributed in an almost equivalent proportion to copper binding in root apoplasts. The high-affinity N functional groups embedded in root apoplasts participated in copper binding in the same magnitude than the low-affinity carboxylic groups.

Inhibition of sulfate reducing bacteria in aquifer sediment by iron nanoparticles.

Batch microcosms were setup to determine the impact of different sized zero valent iron (Fe(0)) particles on microbial sulfate reduction during the in situ bio-precipitation of metals. The microcosms were constructed with aquifer sediment and groundwater from a low pH (3.1), heavy-metal contaminated aquifer. Nano (nFe(0)), micro (mFe(0)) and granular (gFe(0)) sized Fe(0) particles were added to separate microcosms. Additionally, selected microcosms were also amended with glycerol as a C-source for sulfate-reducing bacteria. In addition to metal removal, Fe(0) in microcosms also raised the pH from 3.1 to 6.5, and decreased the oxidation redox potential from initial values of 249 to -226 mV, providing more favorable conditions for microbial sulfate reduction. mFe(0) and gFe(0) in combination with glycerol were found to enhance microbial sulfate reduction. However, no sulfate reduction occurred in the controls without Fe(0) or in the microcosm amended with nFe(0). A separate dose test confirmed the inhibition for sulfate reduction in presence of nFe(0). Hydrogen produced by Fe(0) was not capable of supporting microbial sulfate reduction as a lone electron donor in this study. Microbial analysis revealed that the addition of Fe(0) and glycerol shifted the microbial community towards Desulfosporosinus sp. from a population initially dominated by low pH and metal-resisting Acidithiobacillus ferrooxidans.