Dissolution kinetics of copper oxide nanoparticles in presence of glyphosate.

Recently CuO nanoparticles (n-CuO) have been proposed as an alternative method to deliver a Cu-based pesticide for controlling fungal infestations. With the concomitant use of glyphosate as an herbicide, the interactions between n-CuO and this strong ligand need to be assessed. We investigated the dissolution kinetics of n-CuO and bulk-CuO (b-CuO) particles in the presence of a commercial glyphosate product and compared it to oxalate, a natural ligand present in soil water. We performed experiments at concentration levels representative of the conditions under which n-CuO and glyphosate would be used (∼0.9 mg/L n-CuO and 50 µM of glyphosate). As tenorite (CuO) dissolution kinetics are known to be surface controlled, we determined that at pH 6.5, T âˆ¼ 20 °C, using KNO3 as background electrolyte, the presence of glyphosate leads to a dissolution rate of 9.3 ± 0.7 ×10-3 h-1. In contrast, in absence of glyphosate, and under the same conditions, it is 2 orders of magnitude less: 8.9 ± 3.6 ×10-5 h-1. In a more complex multi-electrolyte aqueous solution the same effect is observed; glyphosate promotes the dissolution rates of n-CuO and b-CuO within the first 10 h of reaction by a factor of ∼2 to ∼15. In the simple KNO3 electrolyte, oxalate leads to dissolution rates of CuO about two times faster than glyphosate. However, the kinetic rates within the first 10 h of reaction are about the same for the two ligands when the reaction takes place in the multi-electrolyte solution as oxalate is mostly bound to Ca2+ and Mg2+.

Polyaniline-metal oxide coatings for biocidal applications: Mechanisms of activation and deactivation.

Metal oxide (MO) coatings (e.g. TiO2, ZnO, and CuO) have shown great promise to inactivate pathogenic bacteria, maintain self-cleaning surfaces, and prevent infectious diseases spread via surface contact. Under light illumination, the antibacterial performance of photoactive MO coatings is determined by reactive oxygen species (ROS) generation. However, several drawbacks, such as photo-corrosion and rapid electron-hole recombination, hinder the ROS production of MO coatings and diminish their antibacterial efficiency. In this study, we employed polyaniline (PANI), an inexpensive and easy-to-synthesize conductive polymer, to fabricate polyaniline-metal oxide composite (PMC) films. The antibacterial performance of PMC films was tested using E. coli as the model bacterium and Lake Michigan water (LMW) as the background medium and revealed enhanced antibacterial performance relative to MO coatings alone (approximately 75-90 % kill of E. coli by PMC coatings in comparison to 20-40 % kill by MO coatings), which is explained by an increase in the ROS yields of PMC. However, with repeated use, the antibacterial performance of the PMC coatings is diminished due to deprotonation of the PANI in the neutral/slightly basic aqueous environment of LMW. Overall, PANI can enhance the antibacterial performance of MO coatings, but efforts need to be directed to preserve or regenerate PMC stability under environmental conditions and applications.

An XAS study of Hg(II) sorption to Al-based drinking water treatment residuals.

Drinking water treatment residuals (DWTRs) are produced from the coagulation and flocculation processes in conventional drinking water treatment. The abundant metal oxide content of these materials resulting from the use of coagulants, like alum and ferric chloride, has driven strong research interest into the reuse of DWTRs as sorptive materials. Using a suite of aluminum-based DWTRs, we provide new insights into Hg(II) sorption mechanisms. Experiments performed at circum-neutral pH show that sorption capacities are related to the amount of organic carbon/matter present in DWTRs. We found that carbon rich samples can scavenge about 9000 mg/kg of Hg, in contrast to 2000 mg/kg for lime based DWTRs. X-ray absorption spectroscopy (XAS) at the Hg L3 edge further characterizes mercury coordination. X-ray absorption near edge structure (XANES) and extended x-ray absorption fine structure (EXAFS) results point to a partial association of mercury with sulfur at low mass loadings, transitioning to a full association with oxygen/carbon at higher concentrations of sorbed Hg(II) and in DWTRs with limited sulfur content. These results suggest that sorption of Hg(II) is primarily controlled by the carbon/organic matter fraction of DWTRs, but not by the coagulants.

An algorithm for the automatic deglitching of X-ray absorption spectroscopy data.

Analysis of X-ray absorption spectroscopy data often involves the removal of artifacts or glitches from the acquired signal, a process commonly known as deglitching. Glitches result either from specific orientations of monochromator crystals or from scattering by crystallites in the sample itself. Since the precise energy - or wavelength - location and the intensity of glitches in a spectrum cannot always be predicted, deglitching is often performed on a per spectrum basis by the analyst. Some routines have been proposed, but they are prone to arbitrary selection of spectral artifacts and are often inadequate for processing large data sets. Here, a statistically robust algorithm, implemented as a Python program, for the automatic detection and removal of glitches that can be applied to a large number of spectra, is presented. It uses a Savitzky-Golay filter to smooth spectra and the generalized extreme Studentized deviate test to identify outliers. Robust, repeatable, and selective removal of glitches is achieved using this algorithm.

The impacts of metal-based engineered nanomaterial mixtures on microbial systems: A review.

The last decade has witnessed tremendous growth in the commercial use of metal-based engineered nanomaterials (ENMs) for a wide range of products and processes. Consequently, direct and indirect release into environmental systems may no longer be considered negligible or insignificant. Yet, there is an active debate as to whether there are real risks to human or ecological health with environmental exposure to ENMs. Previous research has focused primarily on the acute effects of individual ENMs using pure cultures under controlled laboratory environments, which may not accurately reveal the ecological impacts of ENMs under real environmental conditions. The goal of this review is to assess our current understanding of ENM effects as we move from exposure of single to multiple ENMs or microbial species. For instance, are ENMs' impacts on microbial communities predicted by their intrinsic physical or chemical characteristics or their effects on single microbial populations; how do chronic ENM interactions compare to acute toxicity; does behavior under simplified laboratory conditions reflect that in environmental media; finally, is biological stress modified by interactions in ENM mixtures relative to that of individual ENM? This review summarizes key findings and our evolving understanding of the ecological effects of ENMs under complex environmental conditions on microbial systems, identifies the gaps in our current knowledge, and indicates the direction of future research.

Acute and Chronic Toxicity Testing of Drinking Water Treatment Residuals in Freshwater Systems.

The beneficial use of drinking water treatment residuals (DWTRs) faces barriers due primarily to uncertainties and concerns about their potential environmental impacts. We used total and water leachable toxic metal concentrations and 2 benthic organism-based bioassays to identify suitable DWTR substrates for introduction to freshwater systems. Using total metal contents and the consensus probable effect concentration concept, 3 DWTRs were selected and used in elutriate and toxicity studies. The concentrations of water leachable Ag, As, Cd, Cu, Cr, Ni, Pb, and Zn were below the US Environmental Protection Agency's ambient water quality criteria. Using the long-term 65-d life cycle Chironomus tentans test and 4 different endpoints (survival, adult emergence, egg case production, and number of eggs produced per female), no statistical differences were found between the DWTR treatments and the controls. Similarly, results obtained using the 10-d Hyalella azteca test showed no toxicity. However, although both survival and growth were recorded in all bioassays, the results of the 10-d C. tentans and the 28-d H. azteca tests were ambiguous. For C. tentans, 2 of the 3 DWTRs resulted in significantly lower survival rates compared to the controls. For H. azteca, no significant growth differences were observed between controls and DWTR treatments, but 2 of the 3 DWTRs resulted in significantly lower survival rates than the controls. Overall, these results suggest that certain DWTR substrates could be suitable for introduction to aquatic systems. Environ Toxicol Chem 2021;40:2005-2014. © 2021 SETAC.

A Screening Approach for the Selection of Drinking Water Treatment Residuals for Their Introduction to Marine Systems.

Drinking water treatment residuals (DWTRs) produced in large quantities worldwide show strong sorption capacities for several contaminants including metals. These by-products of the water-treatment process are primarily discharged as wastes, to either natural or engineered systems, based on the regulations in place in the country where they are produced. To assess how DWTRs can be repurposed to limit the mobility of metals in aquatic systems, we tested their propensity to release toxic metals and their potential ecotoxicity. To account for the wide variability in their physicochemical characteristics, DWTR samples were obtained from 15 water-treatment plants across the United States. A screening procedure based on a combination of 1) the toxicity characteristics leaching procedure (TCLP), 2) total metal contents and sediment quality guidelines, and 3) acute 10-d Americamysis bahia and chronic 28-d Neanthes arenaceodentata survival and growth bioassays was used. All tested samples were found to be nonhazardous based on TCLP results. However, the concentrations of As, Cu, and Ni exceeded the sediment quality guidelines in some samples, resulting in the exclusion of 7 DWTR samples. All of the DWTRs evaluated for toxicity were nontoxic to the tested organisms. The results of the present study suggest that certain DWTRs can be introduced safely into the marine environment and, therefore, used as potential amendments or capping materials to control the mobility of certain sediment contaminants. Environ Toxicol Chem 2021;40:1194-1203. © 2020 SETAC.

Cell-free biosensors for rapid detection of water contaminants.

Lack of access to safe drinking water is a global problem, and methods to reliably and easily detect contaminants could be transformative. We report the development of a cell-free in vitro transcription system that uses RNA Output Sensors Activated by Ligand Induction (ROSALIND) to detect contaminants in water. A combination of highly processive RNA polymerases, allosteric protein transcription factors and synthetic DNA transcription templates regulates the synthesis of a fluorescence-activating RNA aptamer. The presence of a target contaminant induces the transcription of the aptamer, and a fluorescent signal is produced. We apply ROSALIND to detect a range of water contaminants, including antibiotics, small molecules and metals. We also show that adding RNA circuitry can invert responses, reduce crosstalk and improve sensitivity without protein engineering. The ROSALIND system can be freeze-dried for easy storage and distribution, and we apply it in the field to test municipal water supplies, demonstrating its potential use for monitoring water quality.

Partitioning of copper at the confluences of Andean rivers.

The fate of copper (Cu) in rivers impacted by acid drainage remains poorly studied in waters with comparatively low Al and Fe concentrations. This work addresses the role of confluences in controlling the physical and chemical fate of Cu in a system with total molar ratio Cu/Al > 0.2 and Cu/Fe > 0.15. Two consecutive confluences were studied in the upper Mapocho watershed, a densely populated basin with intensive mining located in the Chilean Andes. The inflow had acidic conditions with seasonal variations and Cu up to 9 mg L-1. Lability measurements with diffusive gradient in thin films showed that Cu entered as a dissolved labile form. However, downstream from the confluences a higher pH shifted Cu toward nonlabile compounds and solid phases enriched with Cu. Measurements of x-ray absorption spectroscopy of freshly formed particles showed that composition was dominated by sorbed Cu and Cu(OH)2(s) precipitates, with a higher proportion of sorbed Cu downstream from confluences when pH < 5. Particle size distributions (PSD) measured in field showed that downstream from the confluences the total volume and average diameter of the suspended particles grew progressively, with estimated mean settling velocities increasing from 0.3 to 4.2 cm s-1. As a result, 7-30% of the influent Cu was removed from the river flow. These results highlight that shifts in chemical partition and PSDs in river confluences and the hydrodynamic environments at the river reach level control the mobility of Cu in systems with high Cu/Al and Cu/Fe.

Immobilization of mercury using high-phosphate culture-modified microalgae.

This study developed a novel Hg(II) immobilization strategy by firstly incubating algal cells in high-phosphate cultures for surface modification, followed by obtaining the P-rich biomass as adsorbents for enhanced Hg(II) removal and then charring the Hg-loaded biomass to prevent leaching of phosphate and to immobilize Hg(II). For algal surface modification, Scenedesmus obtusus XJ-15 were cultivated under different P concentrations and obtained the highest sites concentration of surface phosphoryl functional groups in 80 mg L-1 P cultures. For Hg(II) adsorption, biomass from 80 mg L-1 P cultures (B-80) achieved the highest saturated sorption capacity of 95 mg g-1 fitting to Langmuir isotherm model under the optimum pH of 5.0. For charring stabilization, the Hg-loaded B-80 was calcinated under different temperatures, and the product obtained from 300 °C charring showed the lowest Hg(II) leaching rate without P release. Moreover, FT-IR and XPS analysis indicate that the surge of surface phosphoryl functional groups dominated the enhancement of Hg(II) sorption and also Hg(II) charring immobilization. The above results suggested that the developed strategy is promising for both phosphate and mercury removal from water and for co-immobilization of P and Hg(II) to prevent leaching.

The role of cysteine and sulfide in the interplay between microbial Hg(ii) uptake and sulfur metabolism.

Biogenic thiols, such as cysteine, have been used to control the speciation of Hg(ii) in bacterial exposure experiments. However, the extracellular biodegradation of excess cysteine leads to the formation of Hg(ii)-sulfide species, convoluting the interpretation of Hg(ii) uptake results. Herein, we test the hypothesis that Hg(ii)-sulfide species formation is a critical step during bacterial Hg(ii) uptake in the presence of excess cysteine. An Escherichia coli (E. coli) wild-type and mutant strain lacking the decR gene that regulates cysteine degradation to sulfide were exposed to 50 and 500 nM Hg with 0 to 2 mM cysteine. The decR mutant released ∼4 times less sulfide from cysteine degradation compared to the wild-type for all tested cysteine concentrations during a 3 hour exposure period. We show with thermodynamic calculations that the predicted concentration of Hg(ii)-cysteine species remaining in the exposure medium (as opposed to forming HgS(s)) is a good proxy for the measured concentration of dissolved Hg(ii) (i.e., not cell-bound). Likewise, the measured cell-bound Hg(ii) correlates with thermodynamic calculations for HgS(s) formation in the presence of cysteine. High resolution X-ray absorption near edge structure (HR-XANES) spectra confirm the existence of cell-associated HgS(s) at 500 nM total Hg and suggest the formation of Hg-S clusters at 50 nM total Hg. Our results indicate that a speciation change to Hg(ii)-sulfide controls Hg(ii) cell-association in the presence of excess cysteine.

Solving the problem with stannous fluoride: Formulation, stabilization, and antimicrobial action.

BACKGROUND: Stannous fluoride (SnF2) is a compound present in many commercially available dentifrices; however, oxidative decomposition negatively impacts its efficacy. Stannous oxidation is often mitigated through the addition of complexing agents or sources of sacrificial stannous compounds. The authors have found that the addition of zinc phosphate significantly improved stannous stability more effectively than other stabilization methods. The authors evaluated the chemical speciation of stannous compounds within a variety of formulations using x-ray absorption near edge spectroscopy (XANES), a technique never used before in this manner. These data were compared and correlated with several antimicrobial experiments. METHODS: XANES data of various commercially available compounds and Colgate TotalSF were performed and analyzed against a library of reference compounds to determine the tin chemical speciation. The antibacterial assays used were salivary adenosine triphosphate, short-interval kill test, plaque glycolysis, and anaerobic biofilm models. RESULTS: XANES spectra showed a diverse distribution of tin species and varying degrees of SnF2 oxidation. In vitro antimicrobial assessment indicated significant differences in performance, which may be correlated to the differences in tin speciation and oxidation state. CONCLUSIONS: Driven by the excipient ingredients, SnF2 dentifrices contain a distribution of tin species in either the SnF2 or Sn(IV) oxidation state. The addition of zinc phosphate provided significant robustness against oxidation, which directly translated to greater efficacy against bacteria. PRACTICAL IMPLICATIONS: The choice of inactive ingredients in a dentifrice with active SnF2 can dramatically impact product stability.

Effects of resuspension on the mobility and chemical speciation of zinc in contaminated sediments.

Identifying and quantifying the processes governing the mobilization of metals during resuspension events is key to assessing long-term metals efflux from sediments and associated ecological impacts. We investigated the effects of sediment resuspension on the mobilization and chemical speciation of zinc in two-week-long batch experiments using metal-contaminated sediments from Lake DePue (IL, USA). Measurements of dissolved zinc and sulfate allowed us to characterize the kinetics of metal sulfide dissolution and the resulting net release of zinc to the aqueous phase. X-ray absorption spectroscopy (XAS) provided direct insights into the chemical speciation of iron and zinc and their dynamic transformations during resuspension. While ZnS rapidly oxidized during resuspension, dissolved zinc increased only after two days of resuspension. We proposed a kinetic model to explain changes in the chemical speciation of zinc during these experiments as constrained by the dissolved species concentrations and chemical speciation as informed by XAS. Only 15% of the zinc mobilized was released to the aqueous phase while the remaining fraction repartitioned the solid phase either as a carbonate precipitate or as a sorbed species. Our results show that zinc sorption onto particle surfaces and reprecipitation of zinc minerals limit zinc solubility during resuspension of metal-sulfide sediments.

Spectroscopic and Microscopic Evidence of Biomediated HgS Species Formation from Hg(II)-Cysteine Complexes: Implications for Hg(II) Bioavailability.

We investigated the chemistry of Hg(II) during exposure of exponentially growing bacteria ( Escherichia coli, Bacillus subtilis, and Geobacter sulfurreducens) to 50 nM, 500 nM, and 5 µM total Hg(II) with and without added cysteine. With X-ray absorption spectroscopy, we provide direct evidence of the formation of cell-associated HgS for all tested bacteria. The addition of cysteine (100-1000 µM) promotes HgS formation (>70% of total cell-associated Hg(II)) as a result of the biodegradation of added cysteine to sulfide. Cell-associated HgS species are also detected when cysteine is not added as a sulfide source. Two phases of HgS, cinnabar (α-HgS) and metacinnabar (ß-HgS), form depending on the total concentration of Hg(II) and sulfide in the exposure medium. However, α-HgS exclusively forms in assays that contain an excess of cysteine. Scanning transmission electron microscopy images reveal that nanoparticulate HgS(s) is primarily located at the cell surface/extracellular matrix of Gram-negative E. coli and G. sulfurreducens and in the cytoplasm/cell membrane of Gram-positive B. subtilis. Intracellular Hg(II) was detected even when the predominant cell-associated species was HgS. This study shows that HgS species can form from exogenous thiol-containing ligands and endogenous sulfide in Hg(II) biouptake assays under nondissimilatory sulfate reducing conditions, providing new considerations for the interpretation of Hg(II) biouptake results.

Structural characterization of metal complexes in aqueous solutions: a XAS study of stannous fluoride.

The identity and structure of tin(ii)-fluoride complexes formed in aqueous solutions is determined by combining X-ray absorption spectroscopy, thermodynamic modeling and quantum mechanical calculations. Spectroscopic measurements confirm the presence of 3 stannous fluoride complexes, SnF+, SnF02 and SnF3-, with mean Sn-F bond distances that increase linearly, from 1.98 to about 2.04 Å, as a function of the coordination number. Computational ab initio calculations indicate that the stannous fluoride complexes form localized σs-p bonds, with the stereochemically active lone pair of the Sn(ii) atom distorting the geometry of the complexes. In addition, the SnF3- complex exhibits loosely coordinated water, which is removed upon addition of glycerol to lower the solvent activity. Our results provide spectroscopic confirmation of the stannous fluoride complexes proposed in the literature, and explain why glycerol additions stabilize solutions of Sn(ii) against oxidation.

Synergistic Bacterial Stress Results from Exposure to Nano-Ag and Nano-TiO2 Mixtures under Light in Environmental Media.

Due to their widespread use and subsequent release, engineered nanomaterials (ENMs) will create complex mixtures and emergent systems in the natural environment where their chemical interactions may cause toxic stress to microorganisms. We previously showed that under dark conditions n-TiO2 attenuated bacterial stress caused by low concentrations of n-Ag (<20 µg L-1) due to Ag+ adsorption, yet, since both n-Ag and n-TiO2 are photoactive, their photochemistries may play a key role in their interactions. In this work, we study the chemical interactions of n-Ag and n-TiO2 mixtures in a natural aqueous medium under simulated solar irradiation to investigate photoinduced stress. Using ATP levels and cell membrane integrity as probes, we observe that n-Ag and n-TiO2 together exert synergistic toxic stress in Escherichia coli. We find increased production of hydrogen peroxide by the n-Ag/n-TiO2 mixture, revealing that the enhanced photocatalytic activity and production of ROS likely contribute to the stress response observed. Based on STEM-EDS evidence, we propose that a new composite Ag/TiO2 nanomaterial forms under these conditions and explains the synergistic effects of the ENM mixture. Overall, this work reveals that environmental transformations of ENM mixtures under irradiation can enhance biological stress beyond that of individual components.

Interplay between flow and bioturbation enhances metal efflux from low-permeability sediments.

Understanding the interplay effects between processes such as hydrodynamic forcing, sediment resuspension, and bioturbation is key to assessment of contaminated sediments. In the current study, effects of hydrodynamic forcing, sediment resuspension, and bioturbation by the marine polychaete Nereis virens were evaluated both independently and together in a six-month flume experiment. The results show that hydrodynamic forcing without resuspension or worm action slightly enhanced efflux of dissolved Cu to the water column, sediment resuspension released considerable amounts of dissolved Cu, and interactions between hydrodynamics and worm burrowing further enhanced Cu efflux. In non-bioturbated sediments, fine particles were only resuspended to the overlying water under the highest imposed shear stress, 0.58Pa. However, bioturbated sediments were resuspended under all shear stresses tested (0.11-0.58Pa), indicating that bioturbation destabilized the sediment bed. Further, increases in fluid shear following bioturbation caused rapid releases of dissolved Cu to the overlying water within a few hours. Cu efflux under fluid shears of 0.47Pa and 0.58Pa were 360× and 15× greater after the introduction of worms compared with the same flow conditions without their presence. Overall, our results indicate that the release of metals from low-permeability sediments is greatly enhanced by interactions between flow and bioturbation.

Cysteine Addition Promotes Sulfide Production and 4-Fold Hg(II)-S Coordination in Actively Metabolizing Escherichia coli.

The bacterial uptake of mercury(II), Hg(II), is believed to be energy-dependent and is enhanced by cysteine in diverse species of bacteria under aerobic and anaerobic conditions. To gain insight into this Hg(II) biouptake pathway, we have employed X-ray absorption spectroscopy (XAS) to investigate the relationship between exogenous cysteine, cellular metabolism, cellular localization, and Hg(II) coordination in aerobically respiring Escherichia coli (E. coli). We show that cells harvested in exponential growth phase consistently display mixtures of 2-fold and 4-fold Hg(II) coordination to sulfur (Hg-S2 and Hg-S4), with added cysteine enhancing Hg-S4 formation. In contrast, cells in stationary growth phase or cells treated with a protonophore causing a decrease in cellular ATP predominantly contain Hg-S2, regardless of cysteine addition. Our XAS results favor metacinnabar (ß-HgS) as the Hg-S4 species, which we show is associated with both the cell envelope and cytoplasm. Additionally, we observe that added cysteine abiotically oxidizes to cystine and exponentially growing E. coli degrade high cysteine concentrations (100-1000 µM) into sulfide. Thermodynamic calculations confirm that cysteine-induced sulfide biosynthesis can promote the formation of dissolved and particulate Hg(II)-sulfide species. This report reveals new complexities arising in Hg(II) bioassays with cysteine and emphasizes the need for considering changes in chemical speciation as well as growth stage.

Conformational changes during permeation of Na+ through a modified cyclic peptide nanotube promote energy landscape roughness.

Using a metadynamics approach, we investigate the potential of mean force for Na+ permeation inside a cyclic peptide nanotube (CPN) with modified interior as a function of ion position, coordination number, and lumen chemistry. We show that functionalizing the lumen of a CPN with a methyl-benzoic acid group introduces non-periodic variations in the internal energy of the nanotube, which dictate the overall free energy roughness during the permeation of Na+. These non-periodic variations arise from the structural dynamics of the functional group, where changes in the dihedral angles induced by the proximity of the ion give rise to conformational changes that increase landscape roughness and thereby decrease transport rate. Our computational framework emphasizes the advantages of using the coordination number as a collective variable to investigate the available conformations during ion permeation through CPNs, and reveals new structure-function relations for chemically tunable CPNs, paving the way for rational design of nano-porous systems with tunable selectivity and flux.

Attenuation of Microbial Stress Due to Nano-Ag and Nano-TiO2 Interactions under Dark Conditions.

Engineered nanomaterials (ENMs) are incorporated into thousands of commercial products, and their release into environmental systems creates complex mixtures with unknown toxicological outcomes. To explore this scenario, we probe the chemical and toxicological interactions of nanosilver (n-Ag) and nanotitania (n-TiO2) in Lake Michigan water, a natural aqueous medium, under dark conditions. We find that the presence of n-Ag induces a stress response in Escherichia coli, as indicated by a decrease in ATP production observed at low concentrations (in the µg L-1 range), with levels that are environmentally relevant. However, when n-Ag and n-TiO2 are present together in a mixture, n-TiO2 attenuates the toxicity of n-Ag at and below 20 µg L-1 by adsorbing Ag+(aq). We observe, however, that toxic stress cannot be explained by dissolved silver concentrations alone and, therefore, must also depend on silver associated with the nanoscale fraction. Although the attenuating effect of n-TiO2 on n-Ag's toxicity is limited, this study emphasizes the importance of probing the toxicity of ENM mixtures under environmental conditions to assess how chemical interactions between nanoparticles change the toxicological effects of single ENMs in unexpected ways.