Sonically induced narrowing of the NMR spectra of solids and other novel NMR techniques

A typical liquid state NMR spectrum is composed of a number of discrete absorptions which can be readily interpreted to yield detailed information about the chemical environment of the nuclei found within the sample. The same cannot be said about the spectra of solid samples. For these the absorptions are typically broad, featureless and yield little information directly. This situation may be further exacerbated by the characteristically long T1 values of nuclei bound within a solid lattice which, consequently, require long inter-sequence delays that necessitate lengthy experiments. This work attempts to address both of these inherent problems. Classically, the resolution of the broad-line spectra of solids into discrete resonances has been achieved by imparting to the sample coherent rotation about specific axes in relation to the polarising magnetic field, as implemented in the magic-angle spinning (MAS) [1], dynamic angle spinning (DAS) [2] and double rotation (DOR) [3] NMR experiments. Recently, an alternative method, sonically induced narrowing of the NMR spectra of solids (SINNMR) [4], has been reported which yields the same well resolved solid-state spectra as the classic solid-state NMR experiments, but which achieves the resolution of the broad-line spectra through the promotion of incoherent motion in a suspension of solid particles. The first part of this work examines SINNMR and, in particular, concentrates on ultrasonically induced evaluation, a phenomenon which is thought to be essential to the incoherent averaging mechanism. The second part of this work extends the principle of incoherent motion, implicit in SINNMR, to a new genre of particulate systems, air fluidized beds, and examines the feasibility of such systems to provide well resolved solid state NMR spectra. Samples of trisodium phosphate dodecahydrate and of aluminium granules are examined using the new method with partially resolved spectra being reported in the case of the latter.

Intumescent formulations incorporating melamine phosphate

Once familiar with the fire test rig constructed by M Kay, and modified to allow incorporation of both video and computer facilities, Melamine Phosphate production was scaled up from small to large laboratory scale, and then commercial scale production was considered. Samples produced at each stage were compared analytically, visually and in fire testing. The separation and drying stages on a commercial scale lay unresolved practically, due to lack of test facilities. Different cure regimes for the Araldite MY753 and Versamid system were investigated along with weathering tests and cured samples. Surface priming is suggested for large scale application, though on a small scale a clean unprimed surface was thought sufficient. Some samples heat, aired, cracked at the edges but remained bonded on fire testing. An intumescent sample containing Melamine Phosphate, Araldite and Versamid could not be applied to a vertical surface successfully, the viscosity had to be increased to allow application and curing, various additives were tested, two successful ones being fumed silica and a solvent, isopropanol. The low percentages fumed silica used was incorporated into the sample and the viscosity and fire test results compared with a `standard sample'. An expanding graphite incorporated into a standard sample made mixing and application increasingly difficult, due to the lubricating affect of graphite, but the char produced was a good quality, stable char. A suitable formulation could now be mixed, applied and cured, and assuming no adverse interaction between the additives would protect the sample in the event of a fire.

Pharmacogenomic studies of the anticancer and immunosuppressive thiopurines mercaptopurine and azathioprine

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • 6-Mercaptopurine (6-MP) and azathioprine (AZA) are both inactive prodrugs that require intracellular activation into the active 6-thioguanine nucleotides (6-TGNs). • This metabolic process undergoes three different competitive pathways that are catalysed by three different enzymes; xanthine oxidase (XO), thiopurine methyltransferase (TPMT) and inosine triphosphatase (ITPA), all of which exhibit genetic polymorphisms. • Although the impact of genetic variation in the TPMT gene on treatment outcome and toxicity has been demonstrated, the role of other polymorphisms remains less well known. WHAT THIS STUDY ADDS • New information on the allelic variation of these three enzymes (XO, TPMT and ITPA) and their influence on 6-MP/AZA metabolism and toxicity. • Confirmation of the association of TPMT polymorphism with haematological toxicity. • Identified potential genetic characteristics that may contribute to higher risk of adverse events (such as ITPA IVS2+21A→C mutation). AIMS - To examine the allelic variation of three enzymes involved in 6-mercaptopurine/azathioprine (6-MP/AZA) metabolism and evaluate the influence of these polymorphisms on toxicity, haematological parameters and metabolite levels in patients with acute lymphoblastic leukaemia (ALL) or inflammatory bowel disease (IBD). METHODS - Clinical data and blood samples were collected from 19 ALL paediatric patients and 35 IBD patients who were receiving 6-MP/AZA therapy. All patients were screened for seven genetic polymorphisms in three enzymes involved in mercaptopurine metabolism [xanthine oxidase, inosine triphosphatase (C94→A and IVS2+21A→C) and thiopurine methyltransferase]. Erythrocyte and plasma metabolite concentrations were also determined. The associations between the various genotypes and myelotoxicity, haematological parameters and metabolite concentrations were determined. RESULTS - Thiopurine methyltransferase variant alleles were associated with a preferential metabolism away from 6-methylmercaptopurine nucleotides (P = 0.008 in ALL patients, P = 0.038 in IBD patients) favouring 6-thioguanine nucleotides (6-TGNs) (P = 0.021 in ALL patients). Interestingly, carriers of inosine triphosphatase IVS2+21A→C variants among ALL and IBD patients had significantly higher concentrations of the active cytotoxic metabolites, 6-TGNs (P = 0.008 in ALL patients, P = 0.047 in IBD patients). The study confirmed the association of thiopurine methyltransferase heterozygosity with leucopenia and neutropenia in ALL patients and reported a significant association between inosine triphosphatase IVS2+21A→C variants with thrombocytopenia (P = 0.012). CONCLUSIONS - Pharmacogenetic polymorphisms in the 6-MP pathway may help identify patients at risk for associated toxicities and may serve as a guide for dose individualization.

New biomarkers and risk stratification in atrial fibrillation:simplicity and practicality matter

Advances in our understanding of pathological mechanisms can inform the identification of various biomarkers for risk stratification, monitoring drug efficacy and toxicity; and enabling careful monitoring of polypharmacy. Biomarkers in the broadest sense refer to 'biological markers' and this can be blood-based (eg. fibrin D-dimer, von Willebrand factor, etc) urine-based (eg. thromboxane), or even related to cardiac or cerebral imaging(1). Most biomarkers offer improvements over clinical risk scores in predicting high risk patients - at least statistically - but usually at the loss of simplicity and practicality for easy application in everyday clinical practice. Given the various biomarkers can be informed by different aspects of pathophysiology (e.g. inflammation, clotting, collagen turnover) they can nevertheless contribute to a better understanding of underlying disease processes(2). Indeed, many age-related diseases share common modifiable underpinning mechanisms e.g. inflammation, oxidative stress and visceral adiposity.

Novel sol–gel preparation of (P2O5)0.4–(CaO)0.25–(Na2O)X–(TiO2)(0.35−X) bioresorbable glasses (X = 0.05, 0.1, and 0.15)

Quaternary phosphate-based glasses in the P2O5–CaO–Na2O–TiO2 system with a fixed P2O5 and CaO content of 40 and 25 mol% respectively have been successfully synthesised via sol–gel method and bulk, transparent samples were obtained. The structure, elemental proportion, and thermal properties of stabilised sol–gel glasses have been characterised using X-ray diffraction (XRD), energy dispersive X-ray spectroscopy (EDX), 31P nuclear magnetic resonance (31P NMR), titanium K-edge X-ray absorption near-edge structure (XANES), fourier transform infrared (FTIR) spectroscopy, and differential thermal analysis (DTA). The XRD results confirmed the amorphous nature for all stabilized sol–gel derived glasses. The EDX result shows the relatively low loss of phosphorus during the sol–gel process and Ti K-edge XANES confirmed titanium in the glass structure is in mainly six-fold coordination environment. The 31P NMR and FTIR results revealed that the glass structure consist of mainly Q1 and Q2 phosphate units and the Ti4+ cation was acting as a cross-linking between phosphate units. In addition DTA results confirmed a decrease in the glass transition and crystallisation temperature with increasing Na2O content. Ion release studies also demonstrated a decrease in degradation rates with increasing TiO2 content therefore supporting the use of these glasses for biomedical applications that require a degree of control over glass degradation. These sol–gel glasses also offer the potential to incorporate proactive molecules for drug delivery application due to the low synthesis temperature employed.

Anticancer effects of Curcuma C20-dialdehyde against colon and cervical cancer cell lines

Background: Recent attention on chemotherapeutic intervention against cancer has been focused on discovering and developing phytochemicals as anticancer agents with improved efficacy, low drug resistance and toxicity, low cost and limited adverse side effects. In this study, we investigated the effects of Curcuma C20-dialdehyde on growth, apoptosis and cell cycle arrest in colon and cervical cancer cell lines. Materials and Methods: Antiproliferative, apoptosis induction, and cell cycle arrest activities of Curcuma C20-dialdehyde were determined by WST cell proliferation assay, flow cytometric Alexa fluor 488-annexin V/propidium iodide (PI) staining and PI staining, respectively. Results: Curcuma C20 dialdehyde suppressed the proliferation of HCT116, HT29 and HeLa cells, with IC50 values of 65.4±1.74 μg/ml, 58.4±5.20 μg/ml and 72.0±0.03 μg/ml, respectively, with 72 h exposure. Flow cytometric analysis revealed that percentages of early apoptotic cells increased in a dose-dependent manner upon exposure to Curcuma C20-dialdehyde. Furthermore, exposure to lower concentrations of this compound significantly induced cell cycle arrest at G1 phase for both HCT116 and HT29 cells, while higher concentrations increased sub-G1 populations. However, the concentrations used in this study could not induce cell cycle arrest but rather induced apoptotic cell death in HeLa cells. Conclusions: Our findings suggest that the phytochemical Curcuma C20-dialdehyde may be a potential antineoplastic agent for colon and cervical cancer chemotherapy and/or chemoprevention. Further studies are needed to characterize the drug target or mode of action of the Curcuma C20-dialdehyde as an anticancer agent.

Relative significance of pelagic, benthic and allochthonous organic matter to microbial carbon cycling in a phosphorus-depleted subtropical estuary (Florida Bay)

Heterotrophic bacteria are important decomposers and transformers of primary production and provide an important link between detritus and the aquatic food web. In seagrass ecosystems, much of seagrass primary production is unavailable through direct grazing and must undergo microbial reworking before seagrass production can enter the aquatic food web. The goal of my dissertation research is to understand better the role heterotrophic bacteria play in carbon cycling in seagrass estuaries. My dissertation research focuses on Florida Bay, a seagrass estuary that has experienced recent changes in carbon source availability, which may have altered ecosystem function. My dissertation research investigates the importance of seagrass, algal and/or cyanobacterial, and allochthonous-derived organic matter to heterotrophic bacteria in Florida Bay and helps establish the carbon base of the estuarine food web. ^ A three tiered approach to the study of heterotrophic bacterial carbon cycling and trophic influences in Florida Bay was used: (1) Spatiotemporal observations of environmental parameters (hydrology, nutrients, extracellular enzymes, and microbial abundance, biomass, and production); (2) Microbial grazing experiments under different levels of top-down and bottom-up influence; and (3) Bulk and compound-specific (bacteria-biomarker fatty acid analysis) stable carbon isotope analysis. ^ In Florida Bay, spatiotemporal patterns in microbial extracellular enzyme (also called ectoenzyme) activities indicate that microorganisms hydrolyzed selectively fractions of the estuarine organic matter pool. The microbial community hydrolyzed organic acids, peptides, and phosphate esters and did not use storage and structural carbohydrates. Organic matter use by heterotrophic bacterioplankton in Florida Bay was co-regulated by bottom-up (resource availability) and top-down (grazer mediated) processes. A bacterial carbon budget based on bacterial, epiphytic, and seagrass production indicates that heterotrophic bacterial carbon cycles are supported primarily through epiphytic production with mixing from seagrass production. Stable carbon isotope analysis of bacteria biomarkers and carbon sources in Florida Bay corroborate the results of the bacterial carbon budget. These results support previous studies of aquatic consumers in Florida Bay, indicating that epiphytic/benthic algal and/or cyanobacterial production with mixing from seagrass-derived organic matter is the carbon base of the seagrass estuarine food web. ^

The relationship of thymulin activity, immune parameters of HIV disease progression and substance use in a cohort of HIV seropositive alcohol and drug users

Zinc is essential for the activity of thymulin, a thymic hormone involved in T-lymphocyte differentiation and activation. Zinc deficiency is widespread in populations with HIV infection, and HIV+ drug users are particularly susceptible to zinc deficiency and immune suppression. This dissertation explored the relationship of zinc-bound active thymulin to plasma zinc, CD4+ and CD8+ cell count, the CD4+/CD8+ ratio, and drug use in HIV-infected drug users. Zinc-bound active thymulin was assessed in plasma of HIV+ drug users who were participating in a 30 month zinc supplementation trial. Plasma from 80 participants at the 12 month visit, and 40 of these same participants, randomly selected, at the baseline visit were assessed for zinc-bound active thymulin levels using a modification of the rosette inhibition assay. Thymulin activity was directly associated with CD4+ cell count (β = 0.127, p = 0.002) and inversely associated with cocaine use (β = −0.908, p = 0.026; R2 = 0.188, p = 0.019) independent of HIV viral load, age, gender and antiretroviral use. An increase in thymulin activity was 1.4 times more likely when CD4+ cell count increased (OR = 1.402, 95%CI: 1.006–1.956), independent of change in viral load, antiretroviral use, and age. Participants who used cocaine consistently, were 7.6 times less likely to have an increase in thymulin activity (OR = 0.133, 95%CI: 0.017–1.061). There was a direct correlation between change in plasma zinc and change in zinc-bound active thymulin (r = 0.243, p = 0.13). Analysis of CD4+ cell count decline in 222 participants in the zinc supplementation trial across the 30 months showed that both crack cocaine use and heavy alcohol use accelerated CD4+ cell count decline. Thymulin activity is directly associated with HIV disease progression, measured by CD4+ cell count, and is depressed with cocaine use independent of antiretroviral use and HIV viral load. Cocaine and heavy alcohol accelerate CD4+ cell count decline. The effect of cocaine on thymic output requires further evaluation as a mechanism for the association of cocaine use with faster HIV disease progression.

AFM and SPR studies of protein adsorption at solid/liquid interface

Membrane-like structure formed by surfactant molecules of didodecyldimethylammonium bromide (DDAB) on both HOPG and gold electrodes were studied with AFM and SPR techniques. The study shows that the thickness of the adsorbed layer of DDAB is strongly dependent on the concentration of the vesicle solution. We have also investigated the adsorption of redox protein, Cytochrome c, on graphite electrode with in situ tapping mode AFM. The protein adsorbs spontaneously onto the electrode covered with an adsorbed phosphate layer and forms a uniform monolayer. The adsorbed protein exhibits a reversible electron transfer at 0.17 V (Ag/AgCI) once the electrode potential has been increased to 0.75V. Using surface plasmon resonance spectroscopy we have measured subtle conformational change in protein, Cyt c, due to electron transfer of a single electron on MPA-coated gold electrode. The electron transfer induced change in the resonant angle is about 0.006 deg., which corresponds to ~ 0.2 A decreases in the thickness. This is consistent with that reduced state is more compact than the oxidized state.

Effects of Natural Organic Matter on the Dissolution Kinetics and Bioavailability of Metal Oxide Nanoparticles

The rapid development of nanotechnology and wider applications of engineered nanomaterials (ENMs) in the last few decades have generated concerns regarding their environmental and health risks. After release into the environment, ENMs undergo aggregation, transformation, and, for metal-based nanomaterials, dissolution processes, which together determine their fate, bioavailability and toxicity to living organisms in the ecosystems. The rates of these processes are dependent on nanomaterial characteristics as well as complex environmental factors, including natural organic matter (NOM). As a ubiquitous component of aquatic systems, NOM plays a key role in the aggregation, dissolution and transformation of metal-based nanomaterials and colloids in aquatic environments.

The goal of this dissertation work is to investigate how NOM fractions with different chemical and molecular properties affect the dissolution kinetics of metal oxide ENMs, such as zinc oxide (ZnO) and copper oxide (CuO) nanoparticles (NPs), and consequently their bioavailability to aquatic vertebrate, with Gulf killifish (Fundulus grandis) embryos as model organisms.

ZnO NPs are known to dissolve at relatively fast rates, and the rate of dissolution is influenced by water chemistry, including the presence of Zn-chelating ligands. A challenge, however, remains in quantifying the dissolution of ZnO NPs, particularly for time scales that are short enough to determine rates. This dissertation assessed the application of anodic stripping voltammetry (ASV) with a hanging mercury drop electrode to directly measure the concentration of dissolved Zn in ZnO NP suspensions, without separation of the ZnO NPs from the aqueous phase. Dissolved zinc concentration measured by ASV ([Zn]ASV) was compared with that measured by inductively coupled plasma mass spectrometry (ICP-MS) after ultracentrifugation ([Zn]ICP-MS), for four types of ZnO NPs with different coatings and primary particle diameters. For small ZnO NPs (4-5 nm), [Zn]ASV was 20% higher than [Zn]ICP-MS, suggesting that these small NPs contributed to the voltammetric measurement. For larger ZnO NPs (approximately 20 nm), [Zn]ASV was (79±19)% of [Zn]ICP-MS, despite the high concentrations of ZnO NPs in suspension, suggesting that ASV can be used to accurately measure the dissolution kinetics of ZnO NPs of this primary particle size.

Using the ASV technique to directly measure dissolved zinc concentration, we examined the effects of 16 different NOM isolates on the dissolution kinetics of ZnO NPs in buffered potassium chloride solution. The observed dissolution rate constants (kobs) and dissolved zinc concentrations at equilibrium increased linearly with NOM concentration (from 0 to 40 mg-C L-1) for Suwannee River humic acid (SRHA), Suwannee River fulvic acid and Pony Lake fulvic acid. When dissolution rates were compared for the 16 NOM isolates, kobs was positively correlated with certain properties of NOM, including specific ultraviolet absorbance (SUVA), aromatic and carbonyl carbon contents, and molecular weight. Dissolution rate constants were negatively correlated to hydrogen/carbon ratio and aliphatic carbon content. The observed correlations indicate that aromatic carbon content is a key factor in determining the rate of NOM-promoted dissolution of ZnO NPs. NOM isolates with higher SUVA were also more effective at enhancing the colloidal stability of the NPs; however, the NOM-promoted dissolution was likely due to enhanced interactions between surface metal ions and NOM rather than smaller aggregate size.

Based on the above results, we designed experiments to quantitatively link the dissolution kinetics and bioavailability of CuO NPs to Gulf killifish embryos under the influence of NOM. The CuO NPs dissolved to varying degrees and at different rates in diluted 5‰ artificial seawater buffered to different pH (6.3-7.5), with or without selected NOM isolates at various concentrations (0.1-10 mg-C L-1). NOM isolates with higher SUVA and aromatic carbon content (such as SRHA) were more effective at promoting the dissolution of CuO NPs, as with ZnO NPs, especially at higher NOM concentrations. On the other hand, the presence of NOM decreased the bioavailability of dissolved Cu ions, with the uptake rate constant negatively correlated to dissolved organic carbon concentration ([DOC]) multiplied by SUVA, a combined parameter indicative of aromatic carbon concentration in the media. When the embryos were exposed to CuO NP suspension, changes in their Cu content were due to the uptake of both dissolved Cu ions and nanoparticulate CuO. The uptake rate constant of nanoparticulate CuO was also negatively correlated to [DOC]×SUVA, in a fashion roughly proportional to changes in dissolved Cu uptake rate constant. Thus, the ratio of uptake rate constants from dissolved Cu and nanoparticulate CuO (ranging from 12 to 22, on average 17±4) were insensitive to NOM type or concentration. Instead, the relative contributions of these two Cu forms were largely determined by the percentage of CuO NP that was dissolved.

Overall, this dissertation elucidated the important role that dissolved NOM plays in affecting the environmental fate and bioavailability of soluble metal-based nanomaterials. This dissertation work identified aromatic carbon content and its indicator SUVA as key NOM properties that influence the dissolution, aggregation and biouptake kinetics of metal oxide NPs and highlighted dissolution rate as a useful functional assay for assessing the relative contributions of dissolved and nanoparticulate forms to metal bioavailability. Findings of this dissertation work will be helpful for predicting the environmental risks of engineered nanomaterials.

The Effects of Brominated Flame Retardants on Thyroid Hormone Homeostasis in Human Placenta Tissues and Cell Culture

Polybrominated diphenyl ethers (PBDEs) are a class of brominated flame retardants (BFRs) that have been heavily used in consumer products such as furniture foams, plastics, and textiles since the mid-1970’s. BFRs are added to products in order to meet state flammability standards intended to increase indoor safety in the event of a fire. The three commercial PBDE mixtures, Penta-, Octa-, and DecaBDE, have all been banned in the United States, however, limited use of DecaBDE is still permitted. PBDEs were phased out of production and added to the Stockholm Convention due to concerns over their environmental persistence and toxicity. Human exposure to PBDEs occurs primarily through the inadvertent ingestion of contaminated house dust, as well as though dietary sources. Despite the phase-out and discontinued use of PBDEs, human exposure to this class of chemicals is likely to continue for decades due to the continued use of treated products and existing environmental reservoirs of PBDEs. Extensive research over the years has shown that PBDEs disrupt thyroid hormone (TH) levels and neurodevelopmental endpoints in rodent and fish models. Additionally, there is growing epidemiological evidence linking PBDE exposure in humans to altered TH homeostasis and neurodevelopmental impairments in children. Due to the importance of THs throughout gestation, there is a great need to understand the effects of BFRs on the developing fetus. Specifically, the placenta plays a critical role in the transport, metabolism, and delivery of THs to the fetal compartment during pregnancy and is a likely target for BFR bioaccumulation and endocrine disruption. The central hypothesis of this dissertation research is that BFRs disrupt the activity of TH sulfotransferase (SULT) enzymes, thereby altering TH concentrations in the placenta.

In the first aim of this dissertation research, the concentrations of PBDEs and 2,4,6-TBP were measured in a cohort of 102 placenta tissue samples from an ongoing pregnancy cohort in Durham, NC. Methods were developed for the extraction and analysis of the BFR analytes. It was found that 2,4,6-TBP was significantly correlated with all PBDE analytes, indicating that 2,4,6-TBP may share common product applications with PBDEs or that 2,4,6-TBP is a metabolite of PBDE compounds. Additionally, this was the first study to measure 2,4,6-TBP in human placenta tissues.

In the second aim of this dissertation research, the placenta tissue concentrations of THs, as well as the endogenous activity of deiodinase (DI) and TH SULT enzymes were quantified using the same cohort of 102 placenta tissue samples. Enzyme activity was detected in all samples and this was the first study to measure TH DI and SULT activity in human placenta tissues. Enzyme activities and TH concentrations were compared with BFR concentrations measured in Aim 1. There were few statistically significant associations observed for the combined data, however, upon stratifying the data set based on infant sex, additional significant associations were observed. For example, among males, those with the highest concentrations of BDE-99 in placenta had T3 levels 0.80 times those with the lowest concentration of BDE-99 (95% confidence interval (CI): 0.59, 1.07). Whereas females with the highest concentrations of BDE-99 in placenta had T3 levels 1.50 times those with the lowest concentration of BDE-99 (95% CI: 1.10, 2.04). Additionally, all BFR analyte concentrations were higher in the placenta of males versus females and they were significantly higher for 2,4,6-TBP and BDE-209. 3,3’-T2 SULT activity was significantly higher in female placenta tissues, while type 3 DI activity was significantly higher in male placenta tissues. This research is the first to show sex-specific differences in the bioaccumulation of BFRs in human placenta tissue, as well as differences in TH concentrations and endogenous DI and SULT activity. The underlying mechanisms of these observed sex differences warrant further investigation.

In the third aim of this dissertation research, the effects of BFRs were examined in a human choriocarcinoma placenta cell line, BeWo. Michaelis-Menten parameters and inhibition curves were calculated for 2,4,6-TBP, 3-OH BDE-47, and 6-OH BDE-47. 2,4,6-TBP was shown to be the most potent inhibitor of 3,3’-T2 SULT activity with a calculated IC50 value of 11.6 nM. It was also shown that 2,4,6-TBP and 3-OH BDE-47 exhibit mixed inhibition of 3,3’-T2 sulfation in BeWo cell homogenates. Next, a series of cell culture exposure experiments were performed using 1, 6, 12, and 24 hour exposure durations. Once again, 2,4,6-TBP was shown to be the most potent inhibitor of basal 3,3’-T2 SULT activity by significantly decreasing activity at the high and medium dose (1 M and 0.5 M, respectively) at all measured time points. Interestingly, BDE-99 was also shown to inhibit basal 3,3’-T2 SULT activity in BeWo cells following the 24 hour exposure, despite exhibiting no inhibitory effects in the BeWo cell homogenate experiments. This indicates that BDE-99 must act through a pathway other than direct enzyme inhibition. Following exposures, the TH concentrations in the cell culture growth media and mRNA expression of TH-related genes were also examined. There was no observed effect of BFR treatment on these endpoints. Future work should focus on determining the downstream biological effects of TH SULT disruption in placental cells, as well as the underlying mechanisms of action responsible for reductions in basal TH SULT activity following BFR exposure.

This was one of the first studies to measure BFRs in a cohort of placenta tissue samples from the United States and the first study to measure THs, DI activity, and SULT activity in human placenta tissues. This research provides a novel contribution to our growing understanding of the effects of BFRs on TH homeostasis within the human placenta, and provides further evidence for sex-specific differences within this important organ. Future research should continue to investigate the effects of environmental contaminants on TH homeostasis within the placenta, as this represents the most critical and vulnerable stage of human development.

Iron status in early life and associations with developmental outcomes

Infants and young children are at particular risk of iron deficiency and its associated consequences for growth and development. The main objectives of this thesis were to quantify iron intakes, status and determinants of status in two year olds; explore determinants of neonatal iron stores; investigate associations between iron status at birth and two years with neurodevelopmental outcomes at two years and explore the influence of growth on iron status in early childhood, using data from the Cork BASELINE (Babies after SCOPE: Evaluating Longitudinal Impact using Neurological and Nutritional Endpoints) Birth Cohort Study (n=2137). Participants were followed prospectively with interviewer-led questionnaires and clinical assessments at day 2 and at 2, 6, 12 and 24 months. At two years, there was a low prevalence of iron deficiency and iron deficiency anaemia in this cohort, representing the largest study of iron status in toddlers in Europe, to date. The increased consumption of iron-fortified products and compliance with recommendations to limit unmodified cows’ milk intakes in toddlers has contributed to the observed improvements in status. Low serum ferritin concentrations at birth, which reflect neonatal iron stores, were shown to track through to two years of age; delivery by Caesarean section, being born small-for-gestational age and maternal obesity and smoking in pregnancy were all associated with significantly lower neonatal iron stores. Despite a low prevalence of iron deficiency in this cohort, both a mean corpuscular volume <74fl and ferritin concentrations <20μg/l were associated with lower neurodevelopmental outcomes at two years. An inverse association between growth in the second year of life and iron status at two years was also observed. This thesis has presented data from one of the largest, extensively-characterised cohorts of young children, to date, to explore iron and its associations with growth and development.

Seawater carbonate chemistry, nutrient uptake and biological processes of coral Stylophora pistillata during experiments, 2011

The effects of ocean acidification and elevated seawater temperature on coral calcification and photosynthesis have been extensively investigated over the last two decades, whereas they are still unknown on nutrient uptake, despite their importance for coral energetics. We therefore studied the separate and combined impacts of increases in temperature and pCO2 on phosphate, ammonium, and nitrate uptake rates by the scleractinian coral S. pistillata. Three experiments were performed, during 10 days i) at three pHT conditions (8.1, 7.8, and 7.5) and normal temperature (26°C), ii) at three temperature conditions (26°, 29°C, and 33°C) and normal pHT(8.1), and iii) at three pHT conditions (8.1, 7.8, and 7.5) and elevated temperature (33°C). After 10 days of incubation, corals had not bleached, as protein, chlorophyll, and zooxanthellae contents were the same in all treatments. However, photosynthetic rates significantly decreased at 33°C, and were further reduced for the pHT 7.5. The photosynthetic efficiency of PSII was only decreased by elevated temperature. Nutrient uptake rates were not affected by a change in pH alone. Conversely, elevated temperature (33°C) alone induced an increase in phosphate uptake but a severe decrease in nitrate and ammonium uptake rates, even leading to a release of nitrogen into seawater. Combination of high temperature (33°C) and low pHT(7.5) resulted in a significant decrease in phosphate and nitrate uptake rates compared to control corals (26°C, pHT = 8.1). These results indicate that both inorganic nitrogen and phosphorus metabolism may be negatively affected by the cumulative effects of ocean warming and acidification.

Seawater carbonate chemistry and growth rate during experiments with coral Acropora cervicornis, 2005

The growth rate of Acropora cervicornis branch tips maintained in the laboratory was measured before, during, and after exposure to elevated nitrate (5 and 10 µM NO3-), phosphate (2 and 4 µM P-PO43) and/or pCO2 (CO2 ~700 to 800 µatm). The effect of increased pCO2 was greater than that of nutrient enrichment alone. High concentrations of nitrate or phosphate resulted in significant decreases in growth rate, in both the presence and absence of increased pCO2. The effect of nitrate and phosphate enrichment combined was additive or antagonistic relative to nutrient concentration and pCO2 level. Growth rate recovery was greater after exposure to increased nutrients or CO2 compared to increased nutrients and CO2. If these results accurately predict coral response in the natural environment, it is reasonable to speculate that the survival and reef-building potential of this species will be significantly negatively impacted by continued coastal nutrification and projected pCO2 increases.

(Table 2) Contents of U, P2O5, CO2, and TOC in phosphorites from Pacific seamounts

Uranium content of in phosphorites from Pacific seamounts does not exceed 10ppm; it is significantly lower than in phosphorites from submarine continental margins and deposits on land. Phosphate is not the main carrier of uranium, which is inhomogeneously distributed in ferromanganese hydroxide-, phosphate-, silicate- and carbonate materials. Uranium associated with phosphate is not isomorphic admixture. Uranium occurs in rocks in fine particles of unknown composition. Ultramicroscopic inclusions of U(IV) oxides have been also found.