R. David Holbrook

Copper Oxide Nanoparticle Mediated DNA Damage in Terrestrial Plant Models

Engineered nanoparticles, due to their unique electrical, mechanical, and catalytic properties, are presently found in many commercial products and will be intentionally or inadvertently released at increasing concentrations into the natural environment. Metal- and metal oxide-based nanomaterials have been shown to act as mediators of DNA damage in mammalian cells, organisms, and even in bacteria, but the molecular mechanisms through which this occurs are poorly understood. For the first time, we report that copper oxide nanoparticles induce DNA damage in agricultural and grassland plants. Significant accumulation of oxidatively modified, mutagenic DNA lesions (7,8-dihydro-8-oxoguanine; 2,6-diamino-4-hydroxy-5-formamidopyrimidine; 4,6-diamino-5-formamidopyrimidine) and strong plant growth inhibition were observed for radish (Raphanus sativus), perennial ryegrass (Lolium perenne), and annual ryegrass (Lolium rigidum) under controlled laboratory conditions. Lesion accumulation levels mediated by copper ions and macroscale copper particles were measured in tandem to clarify the mechanisms of DNA damage. To our knowledge, this is the first evidence of multiple DNA lesion formation and accumulation in plants. These findings provide impetus for future investigations on nanoparticle-mediated DNA damage and repair mechanisms in plants.

Polyelectrolyte and Silver Nanoparticle Modification of Microfiltration Membranes To Mitigate Organic and Bacterial Fouling

Membrane fouling remains one of the most problematic issues surrounding membrane use in water and wastewater treatment applications. Organic and biological fouling contribute to irreversible fouling and flux decline in these processes. The aim of this study was to reduce both organic and biological fouling by modifying the surface of commercially available poly(ether sulfone) (PES) membranes using the polyelectrolyte multilayer modification method with poly(styrenesulfonate) (PSS), poly(diallyldimethylammonium chloride) (PDADMAC), and silver nanoparticles (nanoAg) integrated onto the surface as stable, thin (15 nm) films. PSS increases the hydrophilicity of the membrane and increases the negative surface charge, while integration of nanoAg into the top PSS layer imparts biocidal characteristics to the modified surface. Fouling was simulated by filtering aqueous solutions of humic acid (5 and 20 mg L(-1)), a suspension of Escherichia coli (10(6) colony-forming units (CFU) mL(-1)), and a mixture of both foulants through unmodified and modified PES membranes under batch conditions. Filtration and cleaning studies confirmed that the modification significantly reduced organic and biological fouling.

Evaluation of membrane bioreactor process capabilities to meet stringent effluent nutrient discharge requirements

A six-stage membrane bioreactor (MBR) pilot plant was operated to determine and demonstrate the capability of this process to produce a low-nutrient effluent, consistent with the nutrient reduction goals for the Chesapeake Bay. Biological nitrogen removal was accomplished using a multistage configuration with an initial anoxic zone (using the carbon in the influent wastewater), an aerobic zone (where nitrification occurred), a downstream anoxic zone (where methanol was added as a carbon source), and the aerated submerged membrane zone. The capability to reliably reduce effluent total nitrogen to less than 3 mg/L as nitrogen (N) was demonstrated. A combination of biological (using an initial anaerobic zone) and chemical (using alum) phosphorus removal was used to achieve effluent total phosphate concentrations reliably less than 0.1 mg/L as phosphorus (P) and as low as 0.03 mg/L as P. Alum addition also appeared to enhance the filtration characteristics of the MBR sludge and to reduce membrane fouling. Aeration of the submerged membranes results in thickened sludge with a high dissolved oxygen concentration (approaching saturation), which can be recycled to the main aeration zone rather than to an anoxic or anaerobic zone to optimize biological nutrient removal. Biological nutrient removal was characterized using the International Water Association Activated Sludge Model No. 2d. The stoichiometry of chemical phosphorus removal was also consistent with conventional theory and experience. The characteristics of the solids produced in the MBR were compared with those of a parallel full-scale conventional biological nitrogen removal process and were generally found to be similar. These results provide valuable insight to the design and operating characteristics of MBRs intended to produce effluents with very low nutrient concentrations.

Impact of activated sludge‐derived colloidal organic carbon on behavior of estrogenic agonist recombinant yeast bioassay

The impact of size-fractionated colloidal organic carbon (COC) originating from a biological wastewater treatment facility on the sensitivity of the yeast estrogen screen (YES) bioassay containing the human estrogen receptor (hER) gene was evaluated. Dose-response curves of serially diluted 17beta-estradiol (E2), both in the presence and absence of COC, were generated by the YES bioassay. The concentration of E2 leading to a 50% YES response (effective concentration 50%, or EC50) was used to evaluate quantitatively the estrogenic activity of the different COC-E2 mixtures. The EC50 values for all COC size fractions, including COC-free samples (<1 kD), were statistically greater than the controls using Milli-Q water. Normalized EC50 values significantly increased as a function of COC concentration for the larger size fractions (>0.22 microm), but were not significantly affected by smaller COC material at environmental levels (1-5 mg/L), while both colloidal polysaccharide concentrations and colloidal fluorophores (measured at an excitation/emission wavelength pair of 350 nm/450 nm) appear to have an important role in the sensitivity of the YES bioassay. Estimates of the colloid-associated E2 fraction did not predict accurately increases in EC50 values. Matrix effects of the specific environment being tested with the YES bioassay need to be evaluated closely due to the sensitivity of the hER and reporter plasmid.

Excitation—Emission Matrix Fluorescence Spectroscopy for Natural Organic Matter Characterization: A Quantitative Evaluation of Calibration and Spectral Correction Procedures:

The influence of different data collection procedures and of wavelength-dependent instrumental biases on fluorescence excitation–emission matrix (EEM) spectral analysis of aqueous organic matter samples was investigated. Particular attention was given to fluorescence contours (spectral shape) and peak fluorescence intensities. Instrumental bias was evaluated by independently applying excitation and emission correction factors to the raw excitation and emission data, respectively. The peak fluorescence intensities of representative natural organic matter and tryptophan were significantly influenced by the application of excitation and emission spectral correction factors and by the manner in which the raw data was collected. Humification and fluorescence indices were also influenced by emission correction factors but were independent of reference (excitation) intensity normalization or correction. EEM surface contours were dependent on normalization of the fluorescence intensity to the reference intensity but were not influenced by either excitation or emission spectral correction factors. Authors should be explicit in how excitation and emission spectral correction procedures are implemented in their investigations, which will help to facilitate intra-laboratory comparisons and data sharing.

Multiple Method Analysis of TiO2 Nanoparticle Uptake in Rice (Oryza sativa L.) Plants

Understanding the translocation of nanoparticles (NPs) into plants is challenging because qualitative and quantitative methods are still being developed and the comparability of results among different methods is unclear. In this study, uptake of titanium dioxide NPs and larger bulk particles (BPs) in rice plant (Oryza sativa L.) tissues was evaluated using three orthogonal techniques: electron microscopy, single-particle inductively coupled plasma mass spectroscopy (spICP-MS) with two different plant digestion approaches, and total elemental analysis using ICP optical emission spectroscopy. In agreement with electron microscopy results, total elemental analysis of plants exposed to TiO2 NPs and BPs at 5 and 50 mg/L concentrations revealed that TiO2 NPs penetrated into the plant root and resulted in Ti accumulation in above ground tissues at a higher level compared to BPs. spICP-MS analyses revealed that the size distributions of internalized particles differed between the NPs and BPs with the NPs showing a distribution with smaller particles. Acid digestion resulted in higher particle numbers and the detection of a broader range of particle sizes than the enzymatic digestion approach, highlighting the need for development of robust plant digestion procedures for NP analysis. Overall, there was agreement among the three techniques regarding NP and BP penetration into rice plant roots and spICP-MS showed its unique contribution to provide size distribution information.

Separation, Sizing, and Quantitation of Engineered Nanoparticles in an Organism Model Using Inductively Coupled Plasma Mass Spectrometry and Image Analysis.

For environmental studies assessing uptake of orally ingested engineered nanoparticles (ENPs), a key step in ensuring accurate quantification of ingested ENPs is efficient separation of the organism from ENPs that are either nonspecifically adsorbed to the organism and/or suspended in the dispersion following exposure. Here, we measure the uptake of 30 and 60 nm gold nanoparticles (AuNPs) by the nematode, Caenorhabditis elegans, using a sucrose density gradient centrifugation protocol to remove noningested AuNPs. Both conventional inductively coupled plasma mass spectrometry (ICP-MS) and single particle (sp)ICP-MS are utilized to measure the total mass and size distribution, respectively, of ingested AuNPs. Scanning electron microscopy/energy-dispersive X-ray spectroscopy (SEM/EDS) imaging confirmed that traditional nematode washing procedures were ineffective at removing excess suspended and/or adsorbed AuNPs after exposure. Water rinsing procedures had AuNP removal efficiencies ranging from 57 to 97% and 22 to 83%, while the sucrose density gradient procedure had removal efficiencies of 100 and 93 to 98%, respectively, for the 30 and 60 nm AuNP exposure conditions. Quantification of total Au uptake was performed following acidic digestion of nonexposed and Au-exposed nematodes, whereas an alkaline digestion procedure was optimized for the liberation of ingested AuNPs for spICP-MS characterization. Size distributions and particle number concentrations were determined for AuNPs ingested by nematodes with corresponding confirmation of nematode uptake via high-pressure freezing/freeze substitution resin preparation and large-area SEM imaging. Methods for the separation and in vivo quantification of ENPs in multicellular organisms will facilitate robust studies of ENP uptake, biotransformation, and hazard assessment in the environment.