Measurement of total body water (TBW) and total energy expenditure (TEE) using stable isotopes

Understanding the relationship between diet, physical activity and health in humans requires accurate measurement of body composition and daily energy expenditure. Stable isotopes provide a means of measuring total body water and daily energy expenditure under free-living conditions. While the use of isotope ratio mass spectrometry (IRMS) for the analysis of 2H (Deuterium) and 18O (Oxygen-18) is well established in the field of human energy metabolism research, numerous questions remain regarding the factors which influence analytical and measurement error using this methodology. This thesis was comprised of four studies with the following emphases. The aim of Study 1 was to determine the analytical and measurement error of the IRMS with regard to sample handling under certain conditions. Study 2 involved the comparison of TEE (Total daily energy expenditure) using two commonly employed equations. Further, saliva and urine samples, collected at different times, were used to determine if clinically significant differences would occur. Study 3 was undertaken to determine the appropriate collection times for TBW estimates and derived body composition values. Finally, Study 4, a single case study to investigate if TEE measures are affected when the human condition changes due to altered exercise and water intake. The aim of Study 1 was to validate laboratory approaches to measure isotopic enrichment to ensure accurate (to international standards), precise (reproducibility of three replicate samples) and linear (isotope ratio was constant over the expected concentration range) results. This established the machine variability for the IRMS equipment in use at Queensland University for both TBW and TEE. Using either 0.4mL or 0.5mL sample volumes for both oxygen-18 and deuterium were statistically acceptable (p>0.05) and showed a within analytical variance of 5.8 Delta VSOW units for deuterium, 0.41 Delta VSOW units for oxygen-18. This variance was used as “within analytical noise” to determine sample deviations. It was also found that there was no influence of equilibration time on oxygen-18 or deuterium values when comparing the minimum (oxygen-18: 24hr; deuterium: 3 days) and maximum (oxygen-18: and deuterium: 14 days) equilibration times. With regard to preparation using the vacuum line, any order of preparation is suitable as the TEE values fall within 8% of each other regardless of preparation order. An 8% variation is acceptable for the TEE values due to biological and technical errors (Schoeller, 1988). However, for the automated line, deuterium must be assessed first followed by oxygen-18 as the automated machine line does not evacuate tubes but merely refills them with an injection of gas for a predetermined time. Any fractionation (which may occur for both isotopes), would cause a slight elevation in the values and hence a lower TEE. The purpose of the second and third study was to investigate the use of IRMS to measure the TEE and TBW of and to validate the current IRMS practices in use with regard to sample collection times of urine and saliva, the use of two TEE equations from different research centers and the body composition values derived from these TEE and TBW values. Following the collection of a fasting baseline urine and saliva sample, 10 people (8 women, 2 men) were dosed with a doubly labeled water does comprised of 1.25g 10% oxygen-18 and 0.1 g 100% deuterium/kg body weight. The samples were collected hourly for 12 hrs on the first day and then morning, midday, and evening samples were collected for the next 14 days. The samples were analyzed using an isotope ratio mass spectrometer. For the TBW, time to equilibration was determined using three commonly employed data analysis approaches. Isotopic equilibration was reached in 90% of the sample by hour 6, and in 100% of the sample by hour 7. With regard to the TBW estimations, the optimal time for urine collection was found to be between hours 4 and 10 as to where there was no significant difference between values. In contrast, statistically significant differences in TBW estimations were found between hours 1-3 and from 11-12 when compared with hours 4-10. Most of the individuals in this study were in equilibrium after 7 hours. The TEE equations of Prof Dale Scholler (Chicago, USA, IAEA) and Prof K.Westerterp were compared with that of Prof. Andrew Coward (Dunn Nutrition Centre). When comparing values derived from samples collected in the morning and evening there was no effect of time or equation on resulting TEE values. The fourth study was a pilot study (n=1) to test the variability in TEE as a result of manipulations in fluid consumption and level of physical activity; the magnitude of change which may be expected in a sedentary adult. Physical activity levels were manipulated by increasing the number of steps per day to mimic the increases that may result when a sedentary individual commences an activity program. The study was comprised of three sub-studies completed on the same individual over a period of 8 months. There were no significant changes in TBW across all studies, even though the elimination rates changed with the supplemented water intake and additional physical activity. The extra activity may not have sufficiently strenuous enough and the water intake high enough to cause a significant change in the TBW and hence the CO2 production and TEE values. The TEE values measured show good agreement based on the estimated values calculated on an RMR of 1455 kcal/day, a DIT of 10% of TEE and activity based on measured steps. The covariance values tracked when plotting the residuals were found to be representative of “well-behaved” data and are indicative of the analytical accuracy. The ratio and product plots were found to reflect the water turnover and CO2 production and thus could, with further investigation, be employed to identify the changes in physical activity.

Raman spectroscopy of selected tsumcorite Pb(Zn,Fe3+)2(AsO4)2(OH,H2O) minerals –implications for arsenate accumulation

The presence of arsenic in the environment is a hazard. The accumulation of arsenate by a range of cations in the formation of minerals provides a mechanism for the accumulation of arsenate. The formation of the tsumcorite minerals is an example of a series of minerals which accumulate arsenate. There are about twelve examples in this mineral group. Raman spectroscopy offers a method for the analysis of these minerals. The structure of selected tsumcorite minerals with arsenate and sulphate anions were analysed by Raman spectroscopy. Isomorphic substitution of sulphate for arsenate is observed for gartrellite and thometzekite. A comparison is made with the sulphate bearing mineral natrochalcite. The position of the hydroxyl and water stretching vibrations are related to the strength of the hydrogen bond formed between the OH unit and the AsO43- anion. Characteristic Raman spectra of the minerals enable the assignment of the bands to specific vibrational modes.

Determination of coastal ground and surface water processes and character by use of hydrochemistry and stable isotopes, Fraser Coast, Queensland

This study was part of an integrated project developed in response to concerns regarding current and future land practices affecting water quality within coastal catchments and adjacent marine environments. Two forested coastal catchments on the Fraser Coast, Australia, were chosen as examples of low-modification areas with similar geomorphological and land-use characteristics to many other coastal zones in southeast Queensland. For this component of the overall project, organic , physico-chemical (Eh, pH and DO), ionic (Fe2+, Fe3+), and isotopic (ä13CDIC, ä15NDIN ä34SSO4) data were used to characterise waters and identify sources and processes contributing to concentrations and form of dissolved Fe, C, N and S within the ground and surface waters of these coastal catchments. Three sites with elevated Fe concentrations are discussed in detail. These included a shallow pool with intermittent interaction with the surface water drainage system, a monitoring well within a semi-confined alluvial aquifer, and a monitoring well within the fresh/saline water mixing zone adjacent to an estuary. Conceptual models of processes occurring in these environments are presented. The primary factors influencing Fe transport were; microbial reduction of Fe3+ oxyhydroxides in groundwaters and in the hyporheic zone of surface drainage systems, organic input available for microbial reduction and Fe3+ complexation, bacterial activity for reduction and oxidation, iron curtain effects where saline/fresh water mixing occurs, and variation in redox conditions with depth in ground and surface water columns. Data indicated that groundwater seepage appears a more likely source of Fe to coastal waters (during periods of low rainfall) via tidal flux. The drainage system is ephemeral and contributes little discharge to marine waters. However, data collected during a high rainfall event indicated considerable Fe loads can be transported to the estuary mouth from the catchment.

The effects of low degree alteration on Sm-Nd and U-Pb isotope systematics in Eoarchean basalts from the Doolena Gap and Warralong Greenstone belts, Pilbara Craton, Western Australia

This research led to the discovery of one of the best preserved remnants of the Earth's surficial environment 3.47 billion years ago. These ancient volcanic and sedimentary rocks contain original minerals and textures that are rare in rocks of this age. The research concentrated on chemical analysis of volcanic rocks to differentiate secondary alteration from the primary magmatic signature. This study contributes to our understanding of melting processes and geochemical reservoirs in the early Earth, which is vital for forward modelling of Earth's geodynamic evolution.

The Cressbrook Creek alluvial aquifer system, Southeast Queensland : hydrochemistry and isotopes to determine hydrological processes and response to floods

This study developed an understanding of hydrological processes within the Cressbrook Creek catchment of the upper Brisbane River, in particular for the alluvial aquifers. Those aquifers within the lower catchment are used for intensive irrigation, and have been impacted by long-term drought followed by flooding. The study utilised water chemistry, isotopic characters and hydraulic measurements to determine factors such as recharge, links between creeks and groundwater, and variations in water quality. The catchment-wide study will enable improved management of the local water resources.

Electrocatalytic and SERS activity of Pt rich Pt-Pb nanostructures formed via the utilisation of in-situ underpotential deposition of lead

The controlled synthesis of nanostructured materials remains an ongoing area of research, especially as the size, shape and composition of nanomaterials can greatly influence their properties and applications. In this work we present the electrodeposition of highly dendritic platinum rich platinum-lead nanostructures, where lead acetate acts as an inorganic shape directing agent via underpotential deposition on the growing electrodeposit. It was found that these nanomaterials readily oxidise at potentials below monolayer oxide formation, which significantly impacts on the methanol electrooxidation reaction and correlates with the incipient hydrous oxide adatom mediator (IHOAM) model of electrocatalysis. Additionally these materials were tested for their surface enhanced Raman scattering (SERS) activity, where the high density of sharp tips provides promise for their application as SERS substrates.

Stability of Fe3+ -VO defect pairs in polycrystalline Pb(Ti,Zr)O3

The intensity of the EPR signal with g = 5.985 arising from a ferric ion â oxygen vacancy defect pair (Fe3+ â VO) in PbTiO3, varies with the extent of PbO nonstoichiometry at constant Fe3+ content due to an increased oxygen vacancy concentration. In PZT solid solutions, the signal intensity decreases with an increase in Zr. A lower intensity is also noticed for Fe3+ â VO signals in PbZrO3. This behaviour is explained on the basis of PbO nonstoichiometry arising from independent Pb- and O-vacancies as well as the randomly distributed crystallographic shear (CS) plane defects. The contribution to PbO nonstoichiometry from CS planes is larger in high zirconium compositions of PZT.

Formation of Pb(Ti,Zr)O3, from thermal decomposition of oxalate precursors

Lead zir conyl oxalate hexahydrate (LZO) and lead titanyl zirconyl oxalate hydrate (LTZO) are prepared and characterized. Their thermal decompositions have been investigated by thermoanalytical and gas analysis techniques. The decomposition in air or oxygen has three steps — dehydration, decomposition of the oxalate to a carbonate and the decomposition of carbonate to PbZrO3. In non oxidising atmosphere, partial reduction of Pb(II) to Pb(0) takes place at the oxalate decomposition step. The formation of free metallic lead affects the stoichiometry of the intermediate carbonate and yields a mixture of Pb(Ti,Zr)O3 and ZrO2 as the final products. By maintaining oxidising atmosphere and low heating rate, direct preparation of stoichiometric, crystalline Pb(Ti,Zr)O3 at 550°C is possible from the corresponding oxalate precursor.

Microbial communities in Pb contaminated boreal forest soil

Lead contamination in the environment is of particular concern, as it is a known toxin. Until recently, however, much less attention has been given to the local contamination caused by activities at shooting ranges compared to large-scale industrial contamination. In Finland, more than 500 tons of Pb is produced each year for shotgun ammunition. The contaminant threatens various organisms, ground water and the health of human populations. However, the forest at shooting ranges usually shows no visible sign of stress compared to nearby clean environments. The aboveground biota normally reflects the belowground ecosystem. Thus, the soil microbial communities appear to bear strong resistance to contamination, despite the influence of lead. The studies forming this thesis investigated a shooting range site at Hälvälä in Southern Finland, which is heavily contaminated by lead pellets. Previously it was experimentally shown that the growth of grasses and degradation of litter are retarded. Measurements of acute toxicity of the contaminated soil or soil extracts gave conflicting results, as enchytraeid worms used as toxicity reporters were strongly affected, while reporter bacteria showed no or very minor decreases in viability. Measurements using sensitive inducible luminescent reporter bacteria suggested that the bioavailability of lead in the soil is indeed low, and this notion was supported by the very low water extractability of the lead. Nevertheless, the frequency of lead-resistant cultivable bacteria was elevated based on the isolation of cultivable strains. The bacterial and fungal diversity in heavily lead contaminated shooting sectors were compared with those of pristine sections of the shooting range area. The bacterial 16S rRNA gene and fungal ITS rRNA gene were amplified, cloned and sequenced using total DNA extracted from the soil humus layer as the template. Altogether, 917 sequenced bacterial clones and 649 sequenced fungal clones revealed a high soil microbial diversity. No effect of lead contamination was found on bacterial richness or diversity, while fungal richness and diversity significantly differed between lead contaminated and clean control areas. However, even in the case of fungi, genera that were deemed sensitive were not totally absent from the contaminated area: only their relative frequency was significantly reduced. Some operational taxonomic units (OTUs) assigned to Basidiomycota were clearly affected, and were much rarer in the lead contaminated areas. The studies of this thesis surveyed EcM sporocarps, analyzed morphotyped EcM root tips by direct sequencing, and 454-pyrosequenced fungal communities in in-growth bags. A total of 32 EcM fungi that formed conspicuous sporocarps, 27 EcM fungal OTUs from 294 root tips, and 116 EcM fungal OTUs from a total of 8 194 ITS2 454 sequences were recorded. The ordination analyses by non-parametric multidimensional scaling (NMS) indicated that Pb enrichment induced a shift in the EcM community composition. This was visible as indicative trends in the sporocarp and root tip datasets, but explicitly clear in the communities observed in the in-growth bags. The compositional shift in the EcM community was mainly attributable to an increase in the frequencies of OTUs assigned to the genus Thelephora, and to a decrease in the OTUs assigned to Pseudotomentella, Suillus and Tylospora in Pb-contaminated areas when compared to the control. The enrichment of Thelephora in contaminated areas was also observed when examining the total fungal communities in soil using DNA cloning and sequencing technology. While the compositional shifts are clear, their functional consequences for the dominant trees or soil ecosystem remain undetermined. The results indicate that at the Hälvälä shooting range, lead influences the fungal communities but not the bacterial communities. The forest ecosystem shows apparent functional redundancy, since no significant effects were seen on forest trees. Recently, by means of 454 pyrosequencing , the amount of sequences in a single analysis run can be up to one million. It has been applied in microbial ecology studies to characterize microbial communities. The handling of sequence data with traditional programs is becoming difficult and exceedingly time consuming, and novel tools are needed to handle the vast amounts of data being generated. The field of microbial ecology has recently benefited from the availability of a number of tools for describing and comparing microbial communities using robust statistical methods. However, although these programs provide methods for rapid calculation, it has become necessary to make them more amenable to larger datasets and numbers of samples from pyrosequencing. As part of this thesis, a new program was developed, MuSSA (Multi-Sample Sequence Analyser), to handle sequence data from novel high-throughput sequencing approaches in microbial community analyses. The greatest advantage of the program is that large volumes of sequence data can be manipulated, and general OTU series with a frequency value can be calculated among a large number of samples.

Ab initio electronic structures of rhombohedral and cubic HgXO3 (X = Ti, Pb)

First-principles calculations were performed for orthorhombic HgO, rhombohedral and cubic phases of HgTiO3 (HTO) and HgPbO3 (HPO). The calculations show that in the rhombohedral phase HTO is a direct gap insulator with a gap of ~1.6 eV. The rhombohedral phase of HPO, on the other hand, shows a weak metallic character. The results provide an explanation for the electrical properties of these compounds. The cubic phases of HTO and HPO are invariably metallic in nature, thereby suggesting that for HTO the rhombohedral–cubic transition must also be accompanied by a change in the electrical state. Examination of the electronic density of states of these systems revealed no significant on-site mixing of Hg 5d and Hg 6s states in any of these materials.

Thermodynamic activities in the Pb(Zr1-XTiX)O-3 solid solution at 1373 K

Although Pb(Zr1-XTiX)O-3 solid solution is the cornerstone of the piezoelectric ceramics, there is no information in the literature on thermodynamic activities of the component phases in the solid solution. Using inter-crystalline ion exchange equilibria between Pb(Zr1-XTiX)O-3 solid solution with cubic perovskite structure and (Zr1-YTiY)O-2 solid solutions with monoclinic and tetragonal structures, activities of PbTiO3 and PbZrO3 in the perovskite solid solution have been derived at 1373 K using the modified Gibbs-Duhem integration technique of Jacob and Jeffes. Tie-lines from the cubic solid solution are skewed towards the ZrO2 corner. Activities in the zirconia-rich (Zr1-YTiY)02 solid solutions are taken from a recent emf study. The results for the perovskite solid solution at 1373 K can be represented by a sub-regular solution model: Delta G(E.M) (J mol(-1)) = X-PbTiO3 X-PbZrO3(5280X(PbTiO3) - 1980X(PbZrO3)) where Delta G(E.M) is the excess Gibbs energy of mixing of the cubic solid solution and Xi represents the mole fraction of component i. There is a significant positive deviation from ideality for PbTiO3-rich compositions and mild negative deviation near the PbZrO3 corner. The cubic solid solution is intrinsically stable against composition fluctuations at temperatures down to 840 K. The results contrast sharply with the recent calorimetric data on enthalpy of mixing which signal instability of the cubic perovskite solid solution. (C) 2007 Elsevier B.V. All rights reserved.