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Our knowledge of uptake, toxicity and detoxification mechanisms as related to nanoparticles' (NPs') characteristics remains incomplete. Here we combine the analytical power of three advanced techniques to study the cellular binding and uptake and the intracellular transformation of silver nanoparticles (AgNPs): single-particle inductively coupled mass spectrometry, mass cytometry and synchrotron X-ray absorption spectrometry. Our results show that although intracellular and extracellularly bound AgNPs undergo major transformation depending on their primary size and surface coating, intracellular Ag in 24 h AgNP-exposed human lymphocytes exists in nanoparticulate form. Biotransformation of AgNPs is dominated by sulfidation, which can be viewed as one of the cellular detoxification pathways for Ag. These results also show that the toxicity of AgNPs is primarily driven by internalized Ag. In fact, when toxicity thresholds are expressed as the intracellular mass of Ag per cell, differences in toxicity between NPs of different coatings and sizes are minimized. The analytical approach developed here has broad applicability in different systems where the aim is to understand and quantify cell-NP interactions and biotransformation., (© 2021. The Author(s), under exclusive licence to Springer Nature Limited.)

Nanoenabled foliar-applied agrochemicals can potentially be safer and more efficient than conventional products. However, limited understanding about how nanoparticle properties influence their interactions with plant leaves, uptake, translocation through the mesophyll to the vasculature, and transport to the rest of the plant prevents rational design. This study used a combination of Au quantification and spatial analysis to investigate how size (3, 10, or 50 nm) and coating chemistry (PVP versus citrate) of gold nanoparticles (AuNPs) influence these processes. Following wheat foliar exposure to AuNPs suspensions (∼280 ng per plant), adhesion on the leaf surface was increased for smaller sizes, and PVP-AuNPs compared to citrate-AuNPs. After 2 weeks, there was incomplete uptake of citrate-AuNPs with some AuNPs remaining on the outside of the cuticle layer. However, the fraction of citrate-AuNPs that had entered the leaf was translocated efficiently to the plant vasculature. In contrast, for similar sizes, virtually all of the PVP-AuNPs crossed the cuticle layer after 2 weeks, but its transport through the mesophyll cells was lower. As a consequence of PVP-AuNP accumulation in the leaf mesophyll, wheat photosynthesis was impaired. Regardless of their coating and sizes, the majority of the transported AuNPs accumulated in younger shoots (10-30%) and in roots (10-25%), and 5-15% of the NPs <50 nm were exuded into the rhizosphere soil. A greater fraction of larger sizes AuNPs (presenting lower ζ potentials) was transported to the roots. The key hypotheses about the NPs physical-chemical and plant physiology parameters that may matter to predict leaf-to-rhizosphere transport are also discussed.

Utilization of nanoparticles (NP) in agriculture as fertilizers or pesticides requires an understanding of the NP properties influencing their interactions with plant roots. To evaluate the influence of the solubility of Cu-based NP on Cu uptake and NP association with plant roots, wheat seedlings were hydroponically exposed to 1 mg/L of Cu NPs with different solubilities [CuO, CuS, and Cu(OH) 2 ] for 1 h then transferred to a Cu-free medium for 48 h. Fresh, hydrated roots were analyzed using micro X-ray fluorescence (μ-XRF) and imaging fluorescence X-ray absorption near edge spectroscopy (XANES imaging) to provide laterally resolved distribution and speciation of Cu in roots. Higher solubility Cu(OH) 2 NPs provided more uptake of Cu after 1 h of exposure, but the lower solubility materials (CuO and CuS) were more persistent on the roots and continued to deliver Cu to plant leaves over the 48 h depuration period. These results demonstrate that NPs, by associating to the roots, have the potential to play a role in slowly providing micronutrients to plants. Thus, tuning the solubility of NPs may provide a long-term slow delivery of micronutrients to plants and provide important information for understanding mechanisms responsible for plant uptake, transformation, and translocation of NPs.

With advances in nanotechnology, particles with various size, shape, surface chemistry, and composition can be easily produced. Nano- and microparticles have been extensively explored in many industrial and clinical applications. Ensuring that the particles themselves are not possessing toxic effects to the biological system is of paramount importance. This paper describes a proof of concept method, in which a microfluidic system is used in conjunction with a cell microarray technique aiming to streamline the analysis of particle-cell interaction in a high throughput manner. Polymeric microparticles, with different particle surface functionalities, were first used to investigate the efficiency of particle-cell adhesion under dynamic flow. Silver nanoparticles (AgNPs, 10 nm in diameter) perfused at different concentrations (0 to 20 μg/mL) in parallel streams over the cell microarray exhibited a higher toxicity compared to the static culture in the 96-well-plate format. This developed microfluidic system can be easily scaled up to accommodate a larger number of microchannels for high throughput analysis of the potential toxicity of a wide range of particles in a single experiment.

Increasing consumer use of engineered nanomaterials has led to significantly increased efforts to understand their potential impact on the environment and living organisms. Currently, no individual technique can provide all the necessary information such as their size, distribution, and chemistry in complex biological systems. Consequently, there is a need to develop complementary instrumental imaging approaches that provide enhanced understanding of these "bio-nano" interactions to overcome the limitations of individual techniques. Here we used a multimodal imaging approach incorporating dark-field light microscopy, high-resolution electron microscopy, and nanoscale secondary ion mass spectrometry (NanoSIMS). The aim was to gain insight into the bio-nano interactions of surface-functionalized silver nanoparticles (Ag-NPs) with the green algae Raphidocelis subcapitata, by combining the fidelity, spatial resolution, and elemental identification offered by the three techniques, respectively. Each technique revealed that Ag-NPs interact with the green algae with a dependence on the size (10 nm vs 60 nm) and surface functionality (tannic acid vs branched polyethylenimine, bPEI) of the NPs. Dark-field light microscopy revealed the presence of strong light scatterers on the algal cell surface, and SEM imaging confirmed their nanoparticulate nature and localization at nanoscale resolution. NanoSIMS imaging confirmed their chemical identity as Ag, with the majority of signal concentrated at the cell surface. Furthermore, SEM and NanoSIMS provided evidence of 10 nm bPEI Ag-NP internalization at higher concentrations (40 μg/L), correlating with the highest toxicity observed from these NPs. This multimodal approach thus demonstrated an effective approach to complement dose-response studies in nano-(eco)-toxicological investigations.

Quantification of cell-associated nanoparticles (NPs) is a paramount question in both nanomedicine and nanotoxicology. Inductively coupled plasma mass spectrometry is a well-established method to resolve cell-associated (metal) NPs in bulk cell populations, however, such analysis at single cell level remains a challenge. Here we used mass cytometry, a technique that combines single cell analysis and time-of-flight mass spectrometry, to quantitatively analyze extra- and intracellular silver (Ag) in individual Ag NP exposed human T-lymphocytes. The results revealed significant population heterogeneity: for example, in lymphocytes exposed to 3 μg of 30 nm branched polyethylene imine coated Ag NPs/mL the extracellularly bound Ag varied from 79 to 560 fg and cellular uptake from 17 to 121 fg. Similar amplitude of heterogeneity was observed in cells exposed to various doses of Ag NPs with other sizes and surface coatings, demonstrating the importance of single cell analysis when studying NP-cell interactions. Although mass cytometry has some shortcomings such as inability to analyze potential transformation or dissolution of NPs in cells, we consider this method as the most promising for quantitative assessment of cell-NP interaction at single cell level.

While the importance of nanoparticle (NP) characterization under relevant test conditions is widely recognized in nanotoxicology, few studies monitor NPs behavior in the presence of exposed organisms. Here we studied the behavior of nine types of silver nanoparticles (AgNPs) during the 48 h algal toxicity test. In particular, we investigated NP aggregation and dissolution by time-resolved inductively coupled plasma mass spectrometry and ultrafiltration and performed mass balance measurements to study the distribution of Ag in the test system. We also determined the amount of extra- and intracellular Ag by chemically etching AgNPs on the surface of algal cells and used dark field microscopy for their imaging. We observed that positively charged branched polyethilenimine (bPEI)-coated AgNPs tend to aggregate in the presence of algae and interact with test vessels and algal cells, while citrate-coated AgNPs have a tendency to dissolve. On the other hand, with large variation of half-maximum effective concentration (EC50) across tested NPs (5.4 to 300 ngAg mL -1 ), Ag internalized by the algal cells at EC50 was similar (0.8 to 3.6 ngAg mL -1 ) for all AgNP types. These data show that while sorption to the vessels, dissolution, and aggregation impact on the distribution of AgNPs in the test system and on interactions with algal cells, AgNP toxicity is strongly correlated with the NP-cell surface interaction and internalization of Ag.

The use of zero-valent iron nanoparticles (nZVI) has been advocated for the remediation of both soils and groundwater. A key parameter affecting nZVI remediation efficacy is the mobility of the particles as this influences the reaction zone where remediation can occur. However, by engineering nZVI particles with increased stability and mobility we may also inadvertently facilitate nZVI-mediated contaminant transport away from the zone of treatment. Previous nZVI mobility studies have often been limited to model systems as the presence of background Fe makes detection and tracking of nZVI in real systems difficult. We overcame this problem by synthesising Fe-59 radiolabelled nZVI. This enabled us to detect and quantify the leaching of nZVI-derived Fe-59 in intact soil cores, including a soil contaminated by Chromated-Copper-Arsenate. Mobility of a commercially available nZVI was also tested. The results showed limited mobility of both nanomaterials; <1% of the injected mass was eluted from the columns and most of the radiolabelled nZVI remained in the surface soil layers (the primary treatment zone in this contaminated soil). Nevertheless, the observed breakthrough of contaminants and nZVI occurred simultaneously, indicating that although the quantity transported was low in this case, nZVI does have the potential to co-transport contaminants. These results show that direct injection of nZVI into the surface layers of contaminated soils may be a viable remediation option for soils such as this one, in which the mobility of nZVI below the injection/remediation zone was very limited. This Fe-59 experimental approach can be further extended to test nZVI transport in a wider range of contaminated soil types and textures and using different application methods and rates. The resulting database could then be used to develop and validate modelling of nZVI-facilitated contaminant transport on an individual soil basis suitable for site specific risk assessment prior to nZVI remediation., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Here, we evaluate the extent of sorption of silver nanoparticles (AgNPs) with different primary sizes (30 and 70 nm) and surface properties (branched polyethylene imine, "bPEI" and citrate coating) to laboratory plastic during (eco)toxicological testing. Under conditions of algal growth inhibition assay, up to 97% of the added AgNPs were sorbed onto the test vessels whereas under conditions of in vitro toxicological assay with mammalian cells, the maximum loss of AgNPs was 15%. We propose that the high concentration of proteins and biomolecules in the in vitro toxicological assay originating from serum-containing cell culture medium prevented NP sorption due to steric stabilisation. The sorption of AgNPs to test vessels was clearly concentration dependent. In the conditions of algal growth inhibition assay at 10 ng AgNPs/mL, up to 97% of AgNPs were lost from the test while at higher concentrations (1000 ng AgNPs/mL), the loss of AgNPs was remarkably smaller, up to 64%. Sorption of positively charged bPEI-coated AgNPs was more extensive than the sorption of negatively charged citrate-coated AgNPs and, when calculated on a mass basis, more 70 nm-sized Ag than 30 nm Ag sorbed to plastic surfaces. In summary, this study demonstrates that the loss of AgNPs during (eco)toxicological tests due to sorption on test vessel surfaces is significant, especially in diluted media (e.g. in algal growth medium) and at low NP concentrations. Thus, to ensure the accurate interpretation of (eco)toxicological results, the loss of AgNPs due to adsorption to test vessels should not be overlooked and considered for each specific case.

Engineered zinc oxide (ZnO) and silver (Ag) nanoparticles (NPs) used in consumer products are largely released into the environment through the wastewater stream. Limited information is available regarding the transformations they undergo during their transit through sewerage systems before reaching wastewater treatment plants. To address this knowledge gap, laboratory-scale systems fed with raw wastewater were used to evaluate the transformation of ZnO- and Ag-NPs within sewerage transfer networks. Two experimental systems were established and spiked with either Ag- and ZnO-NPs or with their dissolved salts, and the wastewater influent and effluent samples from both systems were thoroughly characterised. X-ray absorption spectroscopy (XAS) was used to assess the extent of the chemical transformation of both forms of Zn and Ag during transport through the model systems. The results indicated that both ZnO- and Ag-NPs underwent significant transformation during their transport through the sewerage network. Reduced sulphur species represented the most important endpoint for these NPs in the sewer with slight differences in terms of speciation; ZnO converted largely to Zn sulfide, while Ag was also sorbed to cysteine and histidine. Importantly, both ionic Ag and Ag-NPs formed secondary Ag sulfide nanoparticles in the sewerage network as revealed by TEM analysis. Ag-cysteine was also shown to be a major species in biofilms. These results were verified in the field using recently developed nanoparticle in situ deployment devices (nIDDs) which were exposed directly to sewerage network conditions by immersing them into a municipal wastewater network trunk sewer and then retrieving them for XAS analysis., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Long-term speciation and lability of silver (Ag-), silver chloride (AgCl-), and silver sulfide nanoparticles (Ag2S-NPs) in soil were studied by X-ray absorption spectroscopy (XAS), and newly developed “nano” Diffusive Gradients in Thin Films (DGT) devices. These nano-DGT devices were designed specifically to avoid confounding effects when measuring element lability in the presence of nanoparticles. The aging profile and stabilities of the three nanoparticles and AgNO3 (ionic Ag) in soil were examined at three different soil pH values over a period of up to 7 months. Transformation of ionic Ag, Ag-NP and AgCl-NPs were dependent on pH. AgCl formation and persistence was observed under acidic conditions, whereas sulfur-bound forms of Ag dominated in neutral to alkaline soils. Ag2S-NPs were found to be very stable under all conditions tested and remained sulfur bound after 7 months of incubation. Ag lability was characteristically low in soils containing Ag2S-NPs. Other forms of Ag were linked to higher DGT-determined lability, and this varied as a function of aging and related speciation changes as determined by XAS. These results clearly indicate that Ag2S-NPs, which are the most environmentally relevant form of Ag that enter soils, are chemically stable and have profoundly low Ag lability over extended periods. This may minimize the long-term risks of Ag toxicity in the soil environment.

In order to accurately assess the potential environmental risk posed by silver nanoparticles (Ag-NPs), their transformation and fate must be investigated in natural systems. This has proven to be very challenging due to the difficulties encountered in retrieving/analyzing NPs dispersed in complex and heterogeneous environmental matrices at relevant (i.e., low) concentrations. In this study, we overcame this challenge by immobilizing functionalized Ag-NPs onto plasma polymerized solid substrates to form "nano in situ deployment devices" (nIDDs). This method allowed us to retrieve and analyze the Ag-NPs after 48 h of direct exposure in freshwater-sediment and saltwater-sediment environments. The type and extent of Ag-NPs transformation was expected to vary along the water-sediment continuum as sediments typically contain steep gradients in solute concentrations and redox potential. To trace the distribution of redox sensitive elements (e.g., Fe, Mn), Diffusive Equilibration in Thin-films (DET) devices were inserted into the sediments alongside the nIDDs. Chemical transformation of the immobilized Ag-NPs across the water-sediment continuum was investigated after retrieval by synchrotron radiation X-ray Absorption Spectroscopy. Linear combination fitting of Ag K-edge X-ray absorption spectra indicated that the chemical transformations of Ag-NPs in both freshwater and saltwater sediments were strongly affected by the redox conditions over the investigated range. Silver bound to reduced sulfur was the principal product of Ag-NP transformations but different extents of transformation were observed for Ag-NPs exposed to different depths in the sediment. These field results add important insights about the transformation of Ag-NPs in heterogeneous environments.

Silver nanoparticles (NPs) are used in more consumer products than any other nanomaterial and their release into the environment is unavoidable. Of primary concern is the wastewater stream in which most silver NPs are transformed to silver sulfide NPs (Ag2S-NPs) before being applied to agricultural soils within biosolids. While Ag2S-NPs are assumed to be biologically inert, nothing is known of their effects on terrestrial plants. The phytotoxicity of Ag and its accumulation was examined in short-term (24 h) and longer-term (2-week) solution culture experiments with cowpea (Vigna unguiculata L. Walp.) and wheat (Triticum aestivum L.) exposed to Ag2S-NPs (0-20 mg Ag L(-1)), metallic Ag-NPs (0-1.6 mg Ag L(-1)), or ionic Ag (AgNO3; 0-0.086 mg Ag L(-1)). Although not inducing any effects during 24-h exposure, Ag2S-NPs reduced growth by up to 52% over a 2-week period. This toxicity did not result from their dissolution and release of toxic Ag(+) in the rooting medium, with soluble Ag concentrations remaining below 0.001 mg Ag L(-1). Rather, Ag accumulated as Ag2S in the root and shoot tissues when plants were exposed to Ag2S-NPs, consistent with their direct uptake. Importantly, this differed from the form of Ag present in tissues of plants exposed to AgNO3. For the first time, our findings have shown that Ag2S-NPs exert toxic effects through their direct accumulation in terrestrial plant tissues. These findings need to be considered to ensure high yield of food crops, and to avoid increasing Ag in the food chain.

The use of nanoscale Ag in textiles is one the most often mentioned uses of nano-Ag. It has previously been shown that significant amounts of the Ag in the textiles are released upon washing. However, the form of Ag present in the textiles remains largely unknown as product labelling is insufficient. The aim of this study was therefore to investigate the solid phase speciation of Ag in original and washed silver textiles using XANES. The original Ag speciation in the textiles was found to vary greatly between different materials with Ag(0), AgCl, Ag2S, Ag-phosphate, ionic Ag and other species identified. Furthermore, within the same textile a number of different species were found to coexist. This is likely due to a combination of factors such as the synthesis processes at industrial scale and the possible reaction of Ag with atmospheric gases. Washing with two different detergents resulted in marked changes in Ag-speciation. For some textiles the two detergents induced similar transformation, in other textiles they resulted in very different Ag species. This study demonstrates that in functional Ag textiles a variety of different Ag species coexist before and after washing. These results have important implications for the risk assessment of Ag textiles because they show that the metallic Ag is only one of the many silver species that need to be considered., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The transformation and environmental fate of engineered nanomaterials (ENMs) is the focus of intense research due to concerns about their potential impacts in the environment as a result of their uniquely engineered properties. Many approaches are being applied to investigate the complex interactions and transformation processes ENMs may undergo in aqueous and terrestrial environments. However, major challenges remain due to the difficulties in detecting, separating, and analyzing ENMs from environmental matrices. In this work, a novel technique capable of in situ study of ENMs is presented. By exploiting the functional interactions between surface modified silver nanoparticles (AgNPs) and plasma-deposited polymer films, AgNPs were immobilized on to solid supports that can be deployed in the field and retrieved for analysis. Either negatively charged citrate or polyethylene glycol, or positively charged polyethyleneimine were used to cap the AgNPs, which were deployed in two field sites (lake and marina), two standard ecotoxicity media, and in primary sewage sludge for a period of up to 48 h. The chemical and physical transformations of AgNPs after exposure to different environments were analyzed by a combination of XAS and SEM/EDX, taken directly from the substrates. Cystine- or glutathione-bound Ag were found to be the dominant forms of Ag in transformed ENMs, but different extents of transformation were observed across different exposure conditions and surface charges. These results successfully demonstrate the feasibility of using immobilized ENMs to examine their likely transformations in situ in real environments and provide further insight into the short-term fate of AgNPs in the environment. Both the advantages and the limitations of this approach are discussed.

The increasing use of silver (Ag) nanoparticles [containing either elemental Ag (Ag-NPs) or AgCl (AgCl-NPs)] in commercial products such as textiles will most likely result in these materials reaching wastewater treatment plants. Previous studies indicate that a conversion of Ag-NPs to Ag2S is to be expected during wastewater transport/treatment. However, the influence of surface functionality, the nature of the core structure and the effect of post-processing on Ag speciation in sewage sludge/biosolids has not been investigated. This study aims at closing these knowledge gaps using bench scale anaerobic digesters spiked with Ag nitrate, three different types of Ag-NPs, and AgCl-NPs at environmentally realistic concentrations. The results indicate that neither surface functionality nor the different compositions of the NP prevented the formation of Ag2S. Silver sulfides, unlike the sulfides of other metals present in sewage sludge, were stable over a six month period simulating composting/stockpiling., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

The rapid development and commercialization of nanomaterials will inevitably result in the release of nanoparticles (NPs) to the environment. As NPs often exhibit physical and chemical properties significantly different from those of their molecular or macrosize analogs, concern has been growing regarding their fate and toxicity in environmental compartments. The wastewater-sewage sludge pathway has been identified as a key release pathway leading to environmental exposure to NPs. In this study, we investigated the chemical transformation of two ZnO-NPs and one hydrophobic ZnO-NP commercial formulation (used in personal care products), during anaerobic digestion of wastewater. Changes in Zn speciation as a result of postprocessing of the sewage sludge, mimicking composting/stockpiling, were also assessed. The results indicated that "native" Zn and Zn added either as a soluble salt or as NPs was rapidly converted to sulfides in all treatments. The hydrophobicity of the commercial formulation retarded the conversion of ZnO-NP. However, at the end of the anaerobic digestion process and after postprocessing of the sewage sludge (which caused a significant change in Zn speciation), the speciation of Zn was similar across all treatments. This indicates that, at least for the material tested, the risk assessment of ZnO-NP through this exposure pathway can rely on the significant knowledge already available in regard to other "conventional" forms of Zn present in sewage sludge.