Measurement artifacts in the dithiothreitol (DTT) oxidative potential assay caused by interactions between aqueous metals and phosphate buffer.

Metals in particulate matter (PM) are hypothesized to have enhanced toxicity based on their ability to catalyze reactive oxygen species (ROS) formation. Acellular assays are used to measure the oxidative potential (OP) of PM and its individual components. Many OP assays, including the dithiothreitol (DTT) assay, use a phosphate buffer matrix to simulate biological conditions (pH 7.4 and 37 °C). Prior work from our group observed transition metal precipitation in the DTT assay, consistent with thermodynamic equilibria. In this study, we characterized the effects of metal precipitation on OP measured by the DTT assay. Metal precipitation was affected by aqueous metal concentrations, ionic strength, and phosphate concentrations in ambient PM sampled in Baltimore, MD and a standard PM sample (NIST SRM-1648a, Urban Particulate Matter). Critically, differences in metal precipitation induced differing OP responses of the DTT assay as a function of phosphate concentration in all PM samples analyzed. These results indicate that comparison of DTT assay results obtained at differing phosphate buffer concentrations is highly problematic. Further, these results have implications for other chemical and biological assays that use phosphate buffer for pH control and their use to infer PM toxicity.

Precipitation of aqueous transition metals in particulate matter during the dithiothreitol (DTT) oxidative potential assay.

Transition metals in particulate matter (PM) are hypothesized to have enhanced toxicity based on their oxidative potential (OP). The acellular dithiothreitol (DTT) assay is widely used to measure the OP of PM and its chemical components. In our prior study, we showed that the DTT assay (pH 7.4, 0.1 M phosphate buffer, 37 °C) provides favorable thermodynamic conditions for precipitation of multiple metals present in PM. This study utilizes multiple techniques to characterize the precipitation of aqueous metals present at low concentrations in the DTT assay. Metal precipitation was identified using laser particle light scattering analysis, direct chemical measurement of aqueous metal removal, and microscopic imaging. Experiments were run with aqueous metals from individual metal salts and a well-characterized urban PM standard (NIST SRM-1648a, Urban Particulate Matter). Our results demonstrated rapid precipitation of metals in the DTT assay. Metal precipitation was independent of DTT but dependent on metal concentration. Metal removal in the chemically complex urban PM samples exceeded the thermodynamic predictions and removal seen in single metal salt experiments, suggesting co-precipitation and/or adsorption may have occurred. These results have broad implications for other acellular assays that study PM metals using phosphate buffer, and subsequently, the PM toxicity inferred from these assays.

Fate of transition metals in PO4-based in vitro assays: equilibrium modeling and macroscopic studies.

Transition metals are thought to be among the most toxic components in atmospheric particulate matter (PM) due to their role in catalyzing reactive oxygen species (ROS) formation. We show that precipitation of the transition metals Fe(ii), Fe(iii), and Mn(ii) are thermodynamically favored in phosphate-based assays used to measure the oxidative potential (OP) - a surrogate for toxicity - of PM. Fe and Mn precipitation is likely to occur at extremely low metal concentrations (<0.5 µM), levels that are imperceptible to the naked eye. The concentration of each metal (other than Cu) in aqueous PM filter extracts often exceeds the solubility limit in OP assays, indicating favorable thermodynamic conditions for precipitation. Macroscopic experimental results at higher metal concentrations (>100 µM) with visible precipitates provide quasi-validation of the thermodynamic modeling. Oxidation of Fe(ii) to Fe(iii) is likely to be rapid in all in vitro OP assays, transforming Fe to a much less soluble form. Fe precipitates are likely to increase the rate of precipitation of other metals and possibly induce co-precipitation. These results have direct relevance for all PO4-based assays; the implications for studies of PM toxicity are discussed.

Evaluation of sorbent amendments for in situ remediation of metal-contaminated sediments.

The present study evaluated sorbent amendments for in situ remediation of sediments contaminated with two divalent metals. A literature review screening was performed to identify low-cost natural mineral-based metal sorbents and high-performance commercial sorbents that were carried forward into laboratory experiments. Aqueous phase metal sorptivity of the selected sorbents was evaluated because dissolved metals in sediment porewater constitute an important route of exposure to benthic organisms. Based on pH-edge sorption test results, natural sorbents were eliminated due to inferior performance. The potential as in situ sediment amendment was explored by comparing the sorption properties of the engineered amendments in freshwater and saltwater (10 PPT salinity estuarine water) matrices. Self-assembled monolayers on mesoporous supports with thiols (Thiol-SAMMS) and a titanosilicate mineral (ATS) demonstrated the highest sorption capacity for cadmium (Cd) and lead (Pb), respectively. Sequential extraction tests conducted after mixing engineered sorbents with contaminated sediment demonstrated transfer of metal contaminants from a weakly bound state to a more strongly bound state. Biouptake of Cd in a freshwater oligochaete was reduced by 98% after 5-d contact of sediment with 4% Thiol-SAMMS and sorbed Cd was not bioavailable. While treatment with ATS reduced the small easily extractable portion of Pb in the sediment, the change in biouptake of Pb was not significant because most of the native lead was strongly bound. The selected sorbents added to sediments at a dose of 5% were mostly nontoxic to a range of sensitive freshwater and estuarine benthic organisms. Metal sorbent amendments in conjunction with activated carbon have the potential to simultaneously reduce metal and hydrophobic contaminant bioavailability in sediments.

Modeling As(V) removal by a iron oxide impregnated activated carbon using the surface complexation approach.

The objective of this research was to model As(V) removal onto a iron oxide impregnated activated carbon (FeAC) using the surface complexation model (SCM) approach. As(V) removal by FeAC was due to the impregnated Fe oxide, not the base carbon material and was a strong function of pH. The two-monoprotic site-triple layer model adequately described As(V) removal using 2 fitting parameters compared with the 3 parameters needed for the diprotic site model. This, along with a better representation of the recognized As(V) removal mechanism (ligand exchange with -OH) as well as the acid-base behavior makes the two-monoprotic approach the better model for As(V) removal by the impregnated iron oxide although the diprotic model was able to describe the pH dependent removal of As(V). Both models were also able to predict As(V) removal at different adsorbent/adsorbate ratios using K(As) determined from a single FeAC adsorption experiment. Thus, fewer adsorption experiments are required in order to model As(V) removal in equilibrium and column systems. The results described in this work will be used as a foundation in developing a dynamic model to predict As(V) adsorption in a fixed-bed adsorber.