Salmonella enterica serovar Typhimurium is responsible for a variety of diseases in domestic animals and humans. The infection of mice causes similar disease progression to human typhoid fever, thus representing a model for this systemic disease. The ability of S. Typhimurium to reside in a macrophage phagosome is important for their survival and spread to different organs. The antimicrobial mechanisms in this compartment include reactive oxygen species, reactive nitrogen species and elevated copper levels. S. Typhimurium possesses two copper-exporting P1B-type ATPases, CopA and GolT, both of which contribute to copper resistance. A previous study has shown that copper export by CopA and GolT confers a survival advantage in resting macrophage phagosomes. In this study the role of copper resistance systems has been examined further. The reduced survival of ΔcopA/ΔgolT in macrophages is detected beyond 8 hours post infection and coincides with increased nitrite production by macrophages. We have established that ΔcopA/ΔgolT display some increased sensitivity to reactive nitrogen species. However, whilst treatment of macrophages with the iNOS inhibitor L-NMMA reduced macrophage bactericidal activity against wildtype S. Typhimurium, this was not the case for ΔcopA/ΔgolT. In contrast, survival of ΔcopA/ΔgolT was not impaired in macrophages treated with the copper-chelator BCS. Furthermore real-time PCR confirmed the expression of copA and golT is elevated during infection of macrophages treated with IFN-γ or L-NMMA, but is reduced during infection of BCS treated macrophages. This indicates that bactericidal activity in macrophages is associated with copper availability and this is unaffected by reactive nitrogen species released due to iNOS activity. In contrast to Escherichia coli Salmonella lacks a cus system associated with export across the outer membrane and hence the mechanism of copper export from the periplasm is not known. TolC was investigated as a potential outer membrane copper exporter based on clustering of TolC dependent systems to genes with sequence similarity to the S. typhimurium periplasmic copper chaperone CueP, across several bacteria. Mutation of tolC gave reduced copper tolerance and over-accumulation of copper at non-lethal concentrations under aerobic conditions. However TolC does not provide a role in copper tolerance or homeostasis under anaerobic conditions. TolC also does not provide tolerance or homeostasis to other divalent cations: Zn, Ni and Co. TolC therefore provides specific transport of copper under aerobic conditions in S. Typhimurium.

The work presented in this thesis is aimed to design efficient reactive crystallization operations that could potentially be implemented in the manufacture of enantiomerically pure compounds and β-lactam antibiotics. Multiple aspects of solution thermodynamics, reaction engineering and crystallization from complex solutions are involved and will be discussed in detail through the following chapters. The first piece of this work utilizes reactive crystallization for the manufacture of enantiomerically pure amino acids. Chemo-enzymatic stereoiversion reactions are used to enrich saturated or supersaturated solutions to favor the selection of a desired enantiomer. L-Methionine and L-Phenylalanine were resolve successfully from racemic mixtures by cyclic stereoinversion. r D-amino acids were oxidized by D-amino acid oxidase (D-AAO) and the resulting ketoacid was subsequently reduced by ammonia borane producing a racemic-mixture After the necessary enantiomeric enrichment was reached, system conditions were changed to induce supersaturation and promote crystal formation. In each case crystals with chemical and enantiomeric purities greater than 99% wt. were recovered. experimental information about reaction and crystallization kinetics was used to developed models. Such models were used to design model-based optimizations in which the productivity of the operation was enhanced by selecting an optimal temperature profile. The second example is a reactive crystallization towards the manufacture of β-lactam antibiotics. One of the major drawbacks of the utilization of enzymes towards the manufacture of β-lactam antibiotics is the fact that the same enzyme that catalyzes the synthesis of the antibiotic also catalyzes its hydrolysis and thus, its degradation. The reaction scheme is a kinetically controlled synthesis in which the desired product is an intermediate within the network. Hence, the focus of this work is to design an efficient reactive crystallization in which the product is crystallized before it is consumed by hydrolysis. In order to accomplish this goal we have study solution equilibria, reaction kinetics, and crystallization kinetics. Even though crystallization kinetics of ampicillin has been previously reported; the reported models are not applicable to a reactive crystallization scheme for a variety of reasons. In this work, we have developed a robust model that can be applied to multiple crystallization protocols that are consistent with the conditions at which the enzymatic reaction can be performed. Finally, a reactive-crystallization scheme in which ampicillin was successfully recovered from solution was developed. In this work, crystal seeds were used to promote crystallization of the desired product from the complex media. The results indicated that is possible to perform the reaction and crystallization in parallel, and still recover crystals with high purity. This work is the first example in which ampicillin was produced and recovered with high purity in a single stage. Previous work on reaction crystallization of antibiotics reported ampicillin crystallization; however, this was accompanied by precipitation of by-products which greatly reduces the applicability of the operation as product purification is required after the reaction.

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Hard-burnt magnesium oxide (MgO) has been successfully used in large volume concrete, mainly dam construction in China, for thermal shrinkage reduction since the 1970’s. At ~4% addition to the cement by weight, it is much simpler than conventional shrinkage measures and in particular, reduces the quantity of crack control reinforcement used. The mechanism is similar to conventional ettringite-based and lime-based expansive admixture relying on their expansion to offset concrete shrinkage. In other words, the tensile stresses induced by concrete shrinkage are offset by the compressive stresses induced by MgO expansion under a restrained condition so that shrinkage and cracking can be reduced. While there are technical specifications in China for this application, the production of MgO is not robust enough to produce specified MgOs, and thus specific performance cannot be specified or guaranteed. In addition, there has been very little rigorous scientific research work in the literature on the performance mechanisms and controlling variables. These issues have hindered the application of the unique designable expansion characteristics of MgO and have limited the application in thermal shrinkage reduction. Hence, this PhD study aims to investigate the expansive characteristics of a number of commercially available MgOs used as an additive and evaluate their shrinkage reduction in cement and concrete. This work also presents a literature review of MgOs, particularly from Chinese sources, and a thorough evaluation of the expansion characteristics and shrinkage reduction capabilities of MgOs with different reactivities in both cement paste and concrete. To this end, four different reactivity MgOs obtained from seawater, brine and magnesite were tested. First, cement paste and concrete prisms - both cured in water - were used for correlating MgO reactivities with expansion characteristics under unrestrained and restrained conditions. The changes in length in cement paste and concrete prisms showed that the MgO expansion rate and magnitude increased with MgO content and reactivity as well as water content. Provision of restraint in prisms both decreased the magnitude and rate of expansion. Highly reactive MgOs showed higher early age expansion and plateaued in a relatively short period while less reactive MgOs exhibited delayed expansion. Temperature increase accelerated all MgOs hydration activation. In particular, less reactive MgOs displayed rapid expansion at an early age instead of delayed expansion. With regards to unrestrained compressive strength development, it was found that the higher the MgO expansion, the greater the compressive strength reduction. Microstructural analyses confirmed the existence of MgO hydration product brucite which contributed to the expansion. While the brucite quantity increased in cement paste systems, it had an insignificant influence on cement paste pH and thus potential reinforcement corrosion resistance. Similar expansion characteristic was found in the PC-Slag cement pastes containing different reactivities of MgOs as compared to that of PC cement paste except that the PC-Slag cement pastes displayed a slightly higher expansion than PC cement at an early age but lower ultimate expansion. One high reactivity MgO (MgO-N50) and one medium reactivity MgO (MgO-92/200), with distinct expansion characteristics, were selected for evaluating the shrinkage reduction performance of cement paste in autogenous and drying curing conditions. Autogenous shrinkage of unrestrained cement paste prisms was measured using laser sensors and length comparator at an early age (< 24 hours) and long-term respectively. The results showed that autogenous shrinkage in the cement paste containing the MgOs was reduced and further decreased with MgO reactivity, content, as well as water content. Compared to PC, the slower strength development in PC-Slag cement resulted in smaller autogenous shrinkage reduction. Drying shrinkage reduction was observed at an early age in both PC and PC-Slag unrestrained cement paste containing the MgOs at water-to-bonder (w/b) ratio of 0.5, although the long-term shrinkage was higher than the control. To further investigate the benefit of early age MgO expansion in drying shrinkage reduction, the final part of the study focused on restrained concrete specimens using the restrained ring test. The results showed that drying shrinkage was reduced in PC concrete with the addition of the highly reactive MgO-N50 when the w/b ratio was increased from 0.40 and 0.50. On the contrary, drying shrinkage reduction was not observed in the PC concrete with the less reactive MgO-92/200. Neither MgOs in PC-Slag concrete showed drying shrinkage reduction. The results reaffirmed the earlier findings that MgO expansion is dependent on strength development. The combination of pre-compressive stresses, delay in tensile stress development and smaller modulus of elasticity of MgO concrete contributed to a lower stresses-to-strength ratio and thus higher resistance to cracking caused by drying shrinkage. From an engineering point of view, the high reactivity MgO-N50 characterised by early age expansion can be used for reducing concrete autogenous shrinkage as it is used for drying shrinkage effect. The less reactive MgO-92/200 with delayed expansion can be utilised in mass concrete to address thermal shrinkage.

In airway epithelia, lactoperoxidase (LPO) constitutes an important anti-microbial system to protect the host against infection and inflammation. LPO uses hydrogen peroxide and thiocyanate anion to form the biocidal compound, hypothiocyanite. The rate-limiting factor is hydrogen peroxide substrate availability. This study was conducted to identify the major source of hydrogen peroxide and to characterize its regulation in the human airway. Two homologues of the phagocytic NADPH oxidase, Duox1 and Duox2, were shown to be highly expressed and functional in human airway epithelial cells re-differentiated at the air liquid interface (ALI). Duox activity is regulated by intracellular calcium concentration via its two EF-hand motifs. A rise of intracellular calcium concentration exhibited kinetics that correlated with increase of Duox-generated hydrogen peroxide production, which was inhibited by DPI, a NADPH oxidase inhibitor. Additionally, the involvement of Duox activity in the LPO system was investigated. Bacterial products such as flagellin or inflammatory mediators were used to challenge ALI cultures. As a result, mRNAs from Duox2, LPO and DUOXA2, but not Duox1, were up-regulated in response to stimuli. This study provided new information about the regulation of the anti-microbial LPO system in innate immune host defense.

Abstract AlN thin films grown on Si¡BGlass¡BSiO2 and Si3N4 substrate by RF magnetron sputtering technique with AlN target has been studied.Room temperature growth was applied to this study.During thin film growing, sputtering work pressure, sputtering power, sputtering working distance and sputtering time are those key parameters to be adjusted in order to highly C-axis prefer orientation AlN thin films. The microstructures of the AlN films were examined by x-ray diffraction. TEM was adopted to observe grain growth of the AlN films. The results was compared with the results of reactive RF magnetron sputtering. The results of the X-ray patterns showed that the strong C-axis prefer orientation of the AlN films were obtained with AlN target. in a 17¢Q long sputtering working distance condition, chemical composition of substrate can help to growth of AlN films. The column structures of AlN films can be observed by TEM.

Thesis (Ph. D.)--School of Civil and Environmental Engineering, Georgia Institute of Technology, 2004. Directed by Spyros G. Pavlostathis. / Vita. Includes bibliographical references (leaves 381-393).

Thesis (MMedVet (Med) (Veterinary Science))--University of Pretoria, 2009. / Includes bibliographical references. Also available in print format.