This study examines three possible methods for improving the digestibility and bioavailability of zinc to rainbow trout (Oncorhynchus mikiss). The first method was to examine the availability of the zinc utilisation from commonly used protein sources; the second was to assess the efficacy of the upstream use of an enzyme treatment of the raw materials; the third was to assess the use of organically complexed mineral supplements as opposed to the inorganic salts widely used at present. The first section indicated that the zinc from the soyabean meal was the most available (49.4%). The zinc digestibilities of the animal based protein used in this current study were 15.1% for LT94 fish meal, 26.6% for the Provimi 66 white fishmeal and 15.8% for poultry meat meal. The zinc in the maize/corn gluten meal was 31.9% digestible and from the NuPro 26.1%. Gram for gram maize gluten meal supplied the least amount of zinc to the fish (3.66 mg per kg). Based on these results the diets for the subsequent supplementation trials were formulated. The liver, eye and caudal fin were identified biomarkers of a severe zinc deficiency. The second part of the study revealed a soybean product, treated by exogenous enzymes, had a higher phosphorus digestibility, (49.0%, vs. 36.6%) and zinc digestibility (30.7% vs. 7.9%) The treatment did not improve the protein digestibility (85%). The third part of the study showed the organic source proved more digestible than the inorganic, 37.4% and 26.9% respectively. The fish fed the organic source maintained a higher level of zinc in both the eye and caudal fin. The liver zinc levels were unaffected by both dietary level and zinc source. Analysis of the liver for a zinc dependant protein showed that under stress conditions only the organic supplemented fish were able to synthesis this protein. The analysis of the mRNA levels coding for this protein indicate the fish on both zinc forms up regulated the production of the mRNA to the same extent when stressed. Finally this study also examined the viability of using a stable isotope to identify different ‘preferences’ for one form of supplementation over the other in different tissues. This method illustrated a tissue dependant difference to how the fish attempted physiologically to compensate for zinc deficiency. The rate of turnover was fastest in the liver, then the caudal fin and then the eye, and also showed that when the diet was more deficient there was an increased ability for the tissues to take up the organic form.

Polycyclic aromatic hydrocarbons (PAHs) are common soil contaminants due to their lipophilic nature which limits partitioning to water or air. Soil properties such as organic carbon can affect PAH release from soil, and thus affect PAH bioavailability of ingested soil. Risk assessment of PAHs in soil generally assumes equal bioavailability of PAHs ingested in soil compared to PAHs ingested in reference dose media, leading to environmental cleanup guidelines that are potentially too conservative. This research intended to use the juvenile swine model to assess PAH bioavailability from impacted soil to better inform bioavailability estimates for risk assessment. This was done by assessing PAH bioavailability from single and repeated exposure to PAHs in different spiked exposure media, assessing PAH bioavailability from soil collected from PAH impacted sites, and assessing biomarkers of exposure and effect following PAH exposure. The effect of exposure duration on bioavailability was assessed because people are usually chronically exposed to PAHs, rather than acutely exposed, as most bioavailability studies are performed, and chronic exposure may lead to increases in xenobiotic metabolizing enzymes and transporters which may affect bioavailability. This research found that exposure duration did not significantly affect anthracene and benzo[a]pyrene bioavailability (p>0.075), but exposure media did (p<0.004). These results suggest that exposure medium has a more important effect on bioavailability than exposure duration, and also bioavailability calculated from a single exposure is appropriate for use in risk assessment. Bioavailability from 24 naturally impacted soils was assessed to determine which soil characteristics had the greatest effect on PAH bioavailability. Area under the curve (AUC) measurements for benzo[a]pyrene (BaP) and anthracene in swine blood after oral exposure from a soil matrix for benzo[a]pyrene and anthracene in soils had s very poor relationship with soil concentrations in soils collected from impacted sites (r2<0.15), but a very strong relationship with soil concentrations from spiked artificial soils (r2<0.95). As spiked soils had much higher concentrations of PAH, these results suggest there is a point of departure in soil concentrations where internal exposure becomes linearly related to soil concentration. Point of departure modeling indicates that this point occurs at soil PAH concentrations greater than 1,900 mg kg-1. Thus, risk assessment can assume a constant exposure to PAHs at soil concentrations lower than the point of departure. Comparison of terminal rate constants from intravenous (IV) exposure to PAHs and oral exposure to PAHs in a soilmatrix suggest that flip-flop kinetics occur in swine, where absorption occurs at a slower rate than elimination. Flip-flop kinetics likely explains the lack of relationship between real world soil concentrations and area under the curve measurements as absorption is the rate limiting step of elimination. Biomarkers of exposure and effect were assessed in swine liver and ileum tissue, as well as blood following single and subchronic exposure to PAHs to determine if relationships could be drawn between exposure magnitude and duration and biomarker formation. Biomarkers included cytochrome P450 (P450) 1A1, 1A2, and 1B1 expression and activity as biomarkers of exposure and DNA adducts, carbonylated proteins, and micronucleated reticulocytes as biomarkers of effect. Biomarkers of exposure were not affected by exposure magnitude or duration, indicating that they would serve best as exposure markers rather than indicators of bioavailability or other effects. However, DNA adduct and protein carbonyl formation was significantly affected by exposure duration (p<0.045), but micronuclei formation was not. The micronuclei results suggest the liver was effective at clearing PAHs to non-toxic metabolites at the study doses, while tissue biomarkers of effect may correlate more effectively with exposure iv length and magnitude of dose. This work indicates that PAH bioavailability from soil is lower than 100%, but additional work needs to be done to determine soil characteristics that affect bioavailability and to determine a bioavailability value relative to reference material.

With the advance of combinational chemistry and high throughput screening, an increasing number of pharmacologically active compounds have been discovered and developed. A significant proportion of those drug candidates are poorly water-soluble, thereby exhibiting limited absorption profiles after oral administration. Therefore, advanced formulation and processing technologies are demanded in order to overcome the biopharmaceutical limits of poorly water-soluble drugs. A number of pharmaceutical technologies have been investigated to address the solubility issue, such as particle size reduction, salt formation, lipid-based formulation, and solubilization. Within the scope of this dissertation, two of the pharmaceutical technologies were investigated names thin film freezing and hot-melt extrusion. The overall goal of the research was to improve the oral bioavailability of poorly water-soluble drugs by producing amorphous solid dispersion systems with enhanced wetting, dissolution, and supersaturation properties. In Chapter 1, the pharmaceutical applications of hot-melt extrusion technology was reviewed. The formulation and process development of hot-melt extrusion was discussed. In Chapter 2, we investigated the use of thin film freezing technology combined with template emulsion system to improve the dissolution and wetting properties of itraconazole (ITZ). The effects of formulation variables (i.e., the selection of polymeric excipients and surfactants) and process variables (i.e., template emulsion system versus cosolvent system) were studied. The physic-chemical properties and dissolution properties of thin film freezing compositions were characterized extensively. In Chapter 3 and Chapter 4, we investigated hot-melt extrusion technology for producing amorphous solid dispersion systems and improving the dissolution and absorption of ITZ. Formulation variables (i.e., the selection of hydrophilic additives, the selection of polymeric carriers) and process variables (i.e., the screw configuration of hot-melt extrusion systems) were investigated in order to optimize the performance of ITZ amorphous solid dispersions. The effects of formulation and process variables on the properties of hot-melt extrusion compositions were investigated. In vivo studies revealed that the oral administration of advanced ITZ amorphous solid dispersion formulations rendered enhanced oral bioavailability of the drug in the rat model. Results indicated that novel formulation and processing technologies are viable approaches for enhancing the oral absorption of poorly water-soluble drugs. / text

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The effects of predicted future climate change was investigated with special emphasis on the association of organic contaminants with dissolved organic carbon (DOC) in the Baltic Sea. An automated method was developed for the measurements of DOC - water distribution constants at realistic DOC concentrations in brackish water. The method proved to be valid for 30 organic contaminants with different structural elements in the 5 – 100 mg car bon/L DOC concentration range. There were limitations of this method. Firstly, its applicability is limited towards contaminants with lower affinity to DOC. Secondly, at higher (&gt;100 mg carbon/L) DOC concentrations the sorption of contaminants was underest imated. Afterwards, water samples were collected from 15 points within the Baltic Sea in a north - south gradient t o examine the spatial differences in DOC characteristics and sorption properties . The DOC samples were analyzed using proton nuclear magnetic resonance and ultraviolet spectroscopy. Results from both techniques indicated that the aromatic nature of the DOC pool increased towards the northern Baltic Sea. This was expected as the freshwater inflow has high significance in controlling the hydrograp hic conditions in the Bothnian Bay. Sorption of organic contaminants was subsequently measured in the same samples. The results showed decreased sorption from north to south for hydrophobic contaminants such as chlorinated benzenes but for contaminants lik e tributyl - phosphate no spatial tendencies were observed. The data generated was used to determine molecular descriptors of DOC using linear free energy relationships. The results indicated a higher significance of hy drogen bond donor/acceptor functional g roups of the DOC in the south. Changes in contaminant distribution were simulated in model pelagic ecosystems at possible endpoints predicted by future climate change scenarios. Separate and combined effects of temperature a nd DOC were studied in mesocosms. The results indicated interesting tendencies. Increased temperature resulted in increased losses in the amounts of organic contaminants. Increased DOC levels promoted sedimentation and sorption of contaminants to particulate matter and biota. Hi gher amounts of contaminants were retained. The combined effects of the two factors led to and overall decrease in dissolved amounts. Higher losses or increased sedimentation and sorption to particles were also observed depending on contaminant properties. / EcoChange

The aim of this study was to develop a suitable monitoring protocol for mediated amperometric whole-cell biosensors for in situ assessment of heavy metals in soil. E. coli 8277, Pseudomonas 9773, Pseudomonas 9046 and Pseudomonas 8917 were screened as biosensor catalysts to select the sensitive biosensor configurations to heavy metals. A new protocol was developed for monitoring heavy metals in defined solution, soil pore water, and in situ in soil. This study also demonstrated the applications of mediated amperometric bacterial biosensors for in situ assessing the bioavailability and toxicity of heavy metals in freshly spiked soils or historically contaminated soils, and mixture toxicities of heavy metals. It was found that the biosensors incorporating selected bacterial strains were appropriately sensitive to copper, but less sensitive to Zn, Pb, and Hg, compared to Microtox assay. The advantage of the mediated amperometric bacterial biosensor system is its in situ application in soils. The present study demonstrated that soil pore water does not accurately reflect conditions of soil ecosystem, and that in situ bioassays are more reliable for determining the bioavailability and toxicity of heavy metals. This is the first reported use of disposable whole cell biosensors for in situ heavy metal bioavailability and toxicity assessment. The biosensor protocol developed here can be adapted to allow the incorporation of dfferent bacterial biocatalysts for applications in soil quality assessment, screening of sites for contamination ‘hot spots’, and the evaluation of soil degradation or rehabilitation from metal pollution. Mediated amperometric bacterial biosensors are not analyte specific, their response reflecting the metabolic impact of the combined chemical and physical properties of the environment to which they are exposed. In assessing the toxicity of soil samples from fields using these biosensors, it is vital to get appropriate control soil samples. The conditions of soil samples also need to be well defined. The sensitivity of the mediated amperometric whole-cell biosensors to heavy metals need to be further improved. Investigations are also required to determine how the natural conditions affect the application of the biosensor system in the field.

There is significant concern about the accumulation of potentially toxic elements (PTEs) in soils because of both direct and indirect impacts on human and ecosystem health. Knowledge of the fate and distribution of such contamination can lead to an effective assessment of the hazards to soil biota and the need for protective or mitigation activities. This is a particular challenge due to the heterogeneity of the soil matrix and complexity of the processes that determine PTE availability to soil biota. While whole-cell bacterial biosensors have been proposed as tools in enabling greater confidence in addressing such biological and chemical interfaces their genuine value remains to be realised. The underpinning objective of this work was to link the response of microbial biosensors to detailed chemical analysis and to relate the dose response sensitivity to other biological measurements. To better understand the phenomena of PTE bioavailability, the study considered changes in toxicity within the context of ion competition in both freshly amended and historically impacted soils. The interaction of test bacteria with both free (soil pore water) and sorbed (solid phase) fractions of the target analytes (copper, nickel and zinc) has enabled a better estimation of bioavailability/toxicity of PTEs in soils. In comparison to other assays, the responses of the microbial sensor to Cu, Ni and Zn highlighted its relative sensitivity to PTE contamination. The use of luminescence marked microbial sensors complements the performance of rigorous analytical soil chemistry approaches. Their value in soil pollution should be considered a technique that should be interpreted alongside chemical analysis rather than an alternative as their performance in complex environmental matrixes is yet to be validated.

The general aims of the thesis were to study GH responsiveness by estimation of pharmacokinetics and bioavailability of injected recombinant human GH (rhGH), of growth response as gain in heightSDS during childhood and puberty, and IGF-I response as change in circulating IGF-ISDS and IGFBP3SDS. Methods Short children were recruited during 1988–1999 into two national randomized multicentre clinical trials on growth until adult height. A group of 117 GHD patients who had been treated from prepuberty with a single GH dose of 33μg/kg/day for at least 1 year were randomized at onset of puberty either to remain on this dose regimen or to an increased dose, GH67μg/kg/day, administered once daily or divided into two doses, GH33x2μg/kg/day. Data on IGF-ISDS and IGF binding protein 3 (IGFBP3)SDS were available from 111 patients and analysed as stated below. The 151 short prepubertal non-GHD patients were randomized into three groups: untreated controls, GH33 or GH67μg/kg/day. A subpopulation from both trials, 128 patients examined annually in Gothenburg, formed the study sample on GH uptake. They received sc GH injections to obtain 16–24 hour GH curves and the GH pharmacokinetics and bioavailability was calculated. Results: A dose-dependent effect on Cmax was found with great intra- and inter-individual variability. Of the Cmax variability, 43% was explained by the rhGH dose and proxies for injection depth. Median bioavailability of the injected dose was 71%, with great variation, mainly dependent on injection depth. In the IGHD group a dose-dependent difference in pubertal gain in heightSDS was found, with mean of 0.8 for the GH67 group and 0.4 for GH33, p&lt;0.01. The mean total gain in heightSDS during treatment was 1.9 for GH67 and 1.4 for GH33, p&lt;0.01. A dose-dependent pubertal ΔIGF-ISDS was 0.5 vs −0.1, p=0.007, correlating to pubertal gain in heightSDS, p=0.003; and was the most important variable to explain the variation in pubertal gain in heightSDS. In the non-GHD group the ΔIGF-ISDS from baseline to mean study level was dose-dependent 2.07 vs 1.20, p=0.001; and correlated negatively with baseline values of IGF-ISDS, rho= -0.56 for GH67, p=0.001, vs rho= -0.82 for GH33, p=0.0001, and correlated positively with gain in heightSDS in both GH-treated groups, rho= 0.42, p&lt;0.001. In multivariable regression analyses, ΔIGF-ISDS was always an important explanatory variable for long-term growth response from the prepubertal period until adult height, while the IGF-ISDS study level per se was not. Conclusion: Growth response to GH treatment was dose dependent with great variability between patients. More pubertal growth was attained by an increased rhGH dose, mimicking the physiology of healthy children, in whom GH secretion rate increases during puberty. This resulted in a gain in IGF-ISDS closely correlating to pubertal gain in heightSDS in both IGHD and non-GHD patients. A broad range in GH responsiveness was found for both growth and IGF response in both diagnostic groups, but lower in the non-GHD group. Higher uptake of a given GH dose was observed after a deep injection and a higher GH concentration. These results are clinically applicable for individuals who remain short close to onset of puberty; by identifying and deeply injecting a rhGH dose that accounts for individual responsiveness, we can stimulate an increment in IGF-ISDS that correlates to gain in heightSDS during puberty.

National Water Quality Criteria intended to protect aquatic life and their uses from the adverse effects of pollutants may not be appropriate due to site-specific factors that alter chemical bioavailability. The Indicator Species Procedure may be used to derive site-specific criteria in order to account for differences in site-specific bioavailability. This procedure was implemented using zinc for three chemically different site (river) waters. The purpose of this study was to quantify the bioavailability of zinc in each site water and correlate results to water quality parameters and/or zinc speciation. Results demonstrated that national criteria for zinc accurately predicted the experimentally derived site-specific values within a factor of two when adjusted for water hardness. Particulate forms of zinc were shown to be biologically unavailable under conditions tested.

Les travaux menés durant cette thèse ont visé à évaluer la pertinence de la plante aquatique Myriophyllum aquaticum pour l'étude de la biodisponibilité du cuivre dans des sédiments ainsi que l'intérêt de la technique du gradient de diffusion en couche mince (DGT) comme outil biomimétique de cette plante. Une première expérimentation réalisée avec la plante M.aquaticum exposée selon le protocole du biotest normalisé (sous forme de verticilles) dans des sédiments artificiels dopés en cuivre a montré que ce biotest semble être un outil intéressant pour l'évaluation de la toxicité de sédiments contaminés en cuivre. En outre, aucune corrélation n'a été observée entre les mesures DGT et la bioaccumulation dans la plante M.aquaticum. Ceci est probablement lié aux conditions d'exposition des plantes. En effet, le développement des racines se faisant au cours de l'expérimentation, les processus d'incorporation du métal dans la plante ne sont donc pas constants au cours de l'expérience. Suite à ces résultats nous avons réalisé deux autres séries d'expérimentations (hydroponie et Rhizotest) dans lesquelles, les plantes ont été exposées selon un protocole modifié (plantes ayant déjà développé des racines avant leur exposition dans le sédiment). Ces expérimentations nous ont permis de montrer que la plante M.aquaticum est plutôt sensible au cuivre et qu'elle a une forte capacité à l'accumuler. De plus, les résultats obtenus ont permis de montrer que la technique DGT semble être un bon outil biomimétique de cette plante dans le cas où elle est exposée avec des racines car le prélèvement par la plante est limité par la capacité de la phase solide à réapprovisionner le métal en solution. / The work carried out during this thesis aimed to assess the relevance of the aquatic plant Myriophyllum aquaticum to study the bioavailability of copper in sediments as well as the interest of the technique of diffusion gradient in thin film (DGT) as a biomimetic tool of this plant. A first experiment, performed with the plant M.aquaticum exposed according to the protocol of the standardized bioassay (as whorls) in an artificial sediment spiked with copper, showed that this bioassay appears to be a useful tool for assessing the toxicity of contaminated sediments copper. Furthermore, no correlation was observed between DGT measurements and bioaccumulation in the M.aquaticum plant. This is probably due to the exposure conditions of the plants. In fact, as the root development occurs during the experiment, the process of incorporating the metal into the plant is therefore not constant throughout the experiment.Following these results we conducted two other series of experiments (hydropony and Rhizotest) in which, plants were exposed following a modified protocol (plants having already developed roots before their exposure in the sediment). These experiments showed that the plant M.aquaticum is rather sensitive to copper and has a strong ability to accumulate it. Futhermore, the results have shown that the DGT technique seems to be a good biomimetic tool for this plant when it is exposed by roots because the plant uptake is limited by the capacity of the solid phase to replenish the metal in solution.