Engineered nanomaterials (NMs), defined as artificially made particles possessing at least one dimension within 1 – 100 nm range, have different physical and biological properties from bulk materials of the same chemistry due to their increased surface areas. Their novel properties have facilitated the prolific growth of commercial NM-incorporating products. NMs may be leached into the aquatic environment during the product life-cycle, but their ecological impacts on marine ecosystems are currently largely unknown. Therefore, this study primarily aimed to investigate the physicochemical characteristics (particle and aggregate sizes, dissolution rate) and in vivo toxicities of commonly-used metallic NMs to marine organisms under various environmental scenarios. First, in vivo ecotoxicity screening tests, using the marine diatom Skeletonema costatum and the rotifer Brachionus sp., were conducted for nine common nano metal oxides which had been applied in various commercial products. Among them, nano zinc oxide (nZnO) and nano magnesia (nMgO) were found to be the most toxic NMs to both organisms, as they induced oxidative stress by increasing reactive oxygen species (ROS) production in the organism. The endocrine disrupting potential of nZnO was revealed by its modulation of transcriptions of the genes for retinoid X receptor (RXR) in the rotifer. Conversely, nano alumina (nAl2O3) and nano indium oxide (nIn2O3) were the least toxic NMs. Due to its high potency, toxicity of nZnO was further evaluated using five marine organisms (i.e., microalgae Thalassiosira pseudonana and S. costatum, copepod Tigriopus japonicus, amphipod Elasmopus rapax, and medaka Oryzias melastigma). Crustaceans were generally more sensitive to nZnO. Toxicity of nZnO was mainly attributed to metal ion dissolution, although nanoparticulate effects such as aggregation and adhesion of nZnO onto the animal’s exoskeleton as well as physical disruption of cell structures could not be discounted. Due to the fact that the dissolution of nZnO decreased from 16 mg Zn L-1 at 4°C to 1.4 mg Zn L-1 at 35°C, and Zn ion was the main contributing factor for nZnO toxicity at 25°C, it was postulated that nZnO toxicity would increase with decreasing temperature. This hypothesis was tested with S. costatum, O. melastigma and the amphipod Melita longidactyla through a factorial design experiment (i.e., 2 concentrations x 4 temperatures). In agreement with the hypothesis, the growth of S. costatum was significantly inhibited by nZnO at the lowest test temperature (15°C). However, contradictive results were observed in the two animal species. For instance, the amphipod could reduce the nZnO uptake and its toxicity by undergoing metabolic depression and dormancy at lower temperatures. As the morphology and coating of NMs utilized in commercial products may differ from those employed in toxicity studies, T. japonicus was exposed to nZnO-containing sunscreens to assess the effects of nZnO and Zn2+ released in seawater during epidermal applications. Based on their genetic biomarker responses, the results suggested that other components in sunscreens could react synergistically or antagonistically on nZnO toxicity. Clearly, there is a need for further study of the combined effects of NMs and other common chemical contaminants to marine organisms. / published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

Nanotoxicology is a nascent field of study concerned with the potential for nanotechnology to adversely impact human health or result in ecological damage. Nanomaterials can display unique physicochemical properties not present in the parent bulk material and it is these properties that may be a potential source of toxicity. There are a growing number of examples of nanomaterials functioning differently in biosystems compared to the parent bulk material. With the rapid growth of nanotechnology and increasing exposure of people to novel nanomaterials there is an urgent need to evaluate the toxicity of nanomaterials. In this study the toxicities of silver and zinc oxide nanoparticles were assessed. The effects of size and surface coating on the cytotoxicity and immunogenicity of silver nanoparticles were investigated, with cytotoxicity found to be inversely proportional to nanoparticle size. The subcutaneous penetration of zinc oxide nanoparticles was assessed to determine whether this material can be safely used as a UV filter in sunscreens and cosmetics. No dermal penetration was detected using a porcine in vitro model. Zinc oxide nanoparticles were also used as a model material to investigate nano-specific toxicity by comparing cytotoxicity and changes to gene expression with bulk scale zinc oxide. In both cases cytotoxicity and changes to gene expression were greater for zinc oxide nanoparticles. Methods and techniques to test the toxicity of nanomaterials in vitro and the implication for in vivo toxicity are only beginning to be elucidated. The methods and techniques used in this study, particularly nanomaterial stabilization in biofluids and toxicity testing using blood cell cultures, may assist the establishment of standard in vitro testing protocols for nanomaterials.

Cardiovascular disease (CVD) is the leading cause of death for people of most ethnicities on a global scale, and countless research efforts on the pathology of CVD has been well-characterized over the years. However, advancement in modern technologies, such as nanotechnology, has generated environmental and occupational health concerns within the scientific community. Current investigation of nanotoxicity calls into question the negative effects nanomaterials may invoke from their environmental, commercial, and therapeutic usage. As a result, further research is needed to investigate and characterize the toxicological implications associated with nanomaterial-exposure and CVD. We investigated the toxicity of multi-walled carbon nanotubes (MWCNT) and titanium dioxide (TiO2), which are two prominently used nanomaterials that have been previously linked to upregulation of inflammatory and atherogenic factors. However, the mechanistic pathways involved in these nanomaterials mediating detrimental effects on the heart and/or coronary vasculature have not yet been fully determined. Thus, we utilized two different routes of exposure in rodent models to assess alterations in proinflammatory and proatherogenic signaling pathways, which are represented in contrast throughout the dissertation. In our MWCNT study, we used C57Bl/6 mice exposed to MWCNTs (1 mg/m3) or filtered air (FA-Controls), via inhalation, for 6 hr/d for 14d. Conversely, intravenous TiO2 was administered to F344 male fisher rats, following 24h and 28d post-exposure to a single injection of TiO2-NPs (1 mg/kg), compared to control animals. MWCNT-exposed endpoints investigated the alterations in cholesterol transport, such as lectin-like oxidized low-density lipoprotein receptor (LOX)-1 and ATP-binding cassette transporter (ABCA)-1, inflammatory markers [tumor necrosis factor (TNF)-α], interleukin (IL)-1β/IL-6, nuclear-factor kappa-light-chain-enhancer of activated B cells (NF-κB) and signaling factors involved in activation of the pathway, as well as intracellular/vascular adhesion molecule(s) (VCAM-1, ICAM-1), and miRNAs (miR-221/-21/-1), associated with CVD, were analyzed in cardiac tissue and coronary vasculature. Cardiac fibrotic deposition, matrix-metalloproteinases (MMP)-2/9, and reactive oxygen species (ROS) were also assessed. TiO2-exposure endpoints also involved alterations on cholesterol transport proteins via LOX-1 and ABCA-1, factors of inflammation, namely intracellular macrophages and interleukin (IL)-1β, MMP-2/9 activity and protein expression, fibrotic deposition, and ROS generation were analyzed via quantitative detection or histologically in both cardiac tissue and coronary vasculature. Results from both studies found alterations in fibrotic deposition, upregulation in LOX-1 expression and MMP-2/9 activity, and ROS generation; with a concurrent decrease in ABCA-1 expression in cardiac tissue and coronary vasculature. Individually, MWCNT-exposed endpoints had shown induction of cardiac TNF-α, MMP-9, IκB Kinase (IKK)-α/β, and miR-221 mRNAs; as well as increased coronary expression of TNF-α and VCAM-1. TiO2 studies found increases in IL-1β and MMP-9 protein expression, as well as intracellular macrophage induction. Both studies also found, through pre-treatment of NADPH oxidase inhibitor, apocynin, resulted in attenuation of nanomaterial-exposure mediated ROS production; with nitric oxide synthase inhibitor, L-NNA, also showing attenuation, but only in our MWCNT-exposed inhalation study. The results from both studies have demonstrated, through different routes of administration, exposures, and rodent models; that exposure to nanomaterials can mediate signaling pathways involved in initiation and/or progression of CVD.

The development of analytical methods for emerging contaminants creates many unique challenges for analytical chemists. By their nature, emerging contaminants have inherent data gaps related to their environmental occurrence, fate, and impact. This dissertation is a compilation of three studies related to method development for the structural identification of emerging contaminants, the detection and quantification of chemicals used in unprecedented quantities and applications, and the extraction of compounds from complex matrices where the solvent-solute-matrix interactions are not completely understood. The three studies present analytical methods developed for emerging contaminants in complex matrices, including: fluorochemical surfactants in aqueous film-forming foams, oil dispersant surfactants in seawater, and fullerene nanomaterials in carbonaceous solids. Aqueous film-forming foams, used in military and commercial firefighting, represent environmentally-relevant commercial mixtures that contain a variety of fluorochemical surfactants. Combining the surfactant-selective ionization of fast atom bombardment mass spectrometry with high resolution mass spectrometry, chemical formulas for 11 different fluorochemical classes were identified. Then AFFF-related patents were used to determine the structures. Of the eleven classes of fluorochemicals, ten have little, if any, data on their environmental occurrence, fate, and potential impacts in the peer-reviewed literature. In addition, nine of the identified classes had either cationic or zwitterionic functionalities and are likely to have different transport properties compared to the well-studied anionic fluorochemicals, such as perfluorooctanoate. After the Deepwater Horizon oil spill in the summer of 2010, one of the emergency response methods for the mitigation of the oil's environmental impact was the use of unprecedented amounts of oil dispersant to break down the oil slick and encourage biodegradation. This event illustrated the need for rapid analytical method development in order to respond to the potential environmental disaster in a timely manner. Using large volume injection liquid chromatography with tandem mass spectrometry, an analytical method was developed for the trace analysis of the multiple dispersant surfactant classes and the potential degradation products of the primary surfactant. Limits of detection ranged from 49 ��� 3,000 ng/L. The method provided excellent recovery (86 ��� 119%) and precision (10 ��� 23% RSD), while also accommodating for the high salinity of seawater samples and analyte contamination. Despite the fact that fullerene nanomaterials have been studied for almost three decades, research is still being conducted to fully understand the environmental properties of these materials. Previous studies to extract fullerenes from environmental matrices have resulted in low efficiency, high variability, or the extraction efficiencies have gone unreported. Extraction by ultrasonication with toluene and 1-methylnaphthalene increased the recovery 5-fold of a spiked, isotopically-labeled C������ surrogate from carbon lampblack as compared to that of the conventional approach of extracting with 100% toluene. The study revealed the importance of evaluating experimental variables such as extraction solvent composition and volume, and sample mass, as they have a significant impact on the quantitative extraction of fullerenes from environmental matrices. / Graduation date: 2013 / Access restricted to the OSU Community at author's request from Aug. 15, 2012 - Aug. 15, 2013