Thesis (Ph.D.)--Ohio University, November, 2008. / Release of full electronic text on OhioLINK has been delayed until December 1, 2010. Includes bibliographical references (leaves 140-162)

Metallophthalocyanine complexes containing non-transition metals are very useful as sensitizers for photodynamic therapy, a cure for cancer that is based on visible light activation of tumour localized photo sensitizers. Excited sensitizers generate singlet oxygen as the main hyperactive species that destroy the tumour. Water soluble sensitizers are sought after for the convenience of delivery into the body. Thus, phthalocyanine (pc), tetrapyridinoporphyrazines (tppa) and tetramethyltetrapyridinoporphyrazines (tmtppa) with non-transition central metal atoms of Ge, Si, Sn and Zn were studied. First was the synthesis of these complexes, followed by their characterisation. The characterisation involved the use of ultraviolet and visible absorption spectroscopy, infrared spectroscopy, nuclear magnetic resonance spectroscopy, electrochemical properties and elemental analysis. Photochemical properties of the complexes were then investigated. Photolysis of these macrocycles showed two processes; -reduction of the dye and photobleaching, which leads to the disintegration of the conjugated chromophore structure of the dye. Photobleaching is the reductive quenching of the excited state of the sensitizers. The intensity of the quenching decreased progressively from tmtppa, tppa to pc metal complexes with photobleaching quantum yields, 6.6 x 10.5⁻¹, 1.8 x 10.5⁻¹ and 5.4 x 10⁻⁶ for Zntmtppa, Zntppa and Znpc, respectively. Efficiency of singlet oxygen sensitization is solvent dependent with very different values obtained for the same compound in different solvents, for example, 0.25 and 0.38 were observed as singlet oxygen quantum yields for Gepc complex in DMSO and DMF respectively. In DMSO the efficiency of ¹O₂ generation decrease considerably from pc to tppa and finally tmtppa. In water Getmtppa exhibits much higher singlet oxygen quantum yield, hence promising to be effective as a sensitizer for photodynamic therapy.

A series of structurally related BODIPY dyes were synthesised and characterised. Their photophysical properties were studied in order to determine whether they would be suitable candidates for use as photosensitisers in the photodynamic therapy (PDT) treatment of cancer. The synthesis of two highly fluorescent BODIPY cores was achieved via the acid-catalysed condensation of a pyrrole and a functionalised aldehyde. In order to promote intersystem crossing, and hence improve the singlet oxygen generation of these dyes, bromine atoms were added at the 2,6-positions of the BODIPY core. These dibrominated analogues showed good singlet oxygen quantum yields, and excellent photostability in ethanol. In order to red-shift the main spectral bands of the BODIPY dyes towards the therapeutic window, vinyl/ styryl groups were introduced at the 3-, 5-, and 7-positions via a modified Knoevengal condensation reaction. The addition of vinyl/ styryl groups to the BODIPY core caused an increase in fluorescence quantum yield as well as a decrease in singlet oxygen quantum yield with respect to the dibrominated analogues. However, two of the red-shifted BODIPY dyes still showed moderate singlet oxygen quantum yields. The use of BODIPY dyes in nonlinear optics (NLO) was explored. The nonlinear optical characterisations and optical limiting properties of a series of 3,5-dithienylenevinylene BODIPY dyes were studied, both in dimethylformamide (DMF) solution and when embedded in poly(bisphenol A carbonate) (PBC) as thin films. The 3,5-dithienylenevinylene BODIPY dyes showed typical nonlinear absorption behaviour, with reverse saturable absorption (RSA) profiles, indicating that they have potential as optical limiters. The second-order hyperpolarizability (Y), and third-order nonlinear susceptibility (/m[/(3)]) values are also reported for these dyes. The optical limiting values of one of the BODIPY dyes in solution, and two of the BODIPY-embedded PBC films, were below the maximum threshold of 0.95 J-cm-2. The effect of addition of substituents on the electronic structure of the BODIPY dyes was investigated using TD-DFT calculations. The calculated trends closely followed those determined experimentally.

Mitochondria are dynamic organelles that harbor their own stream-lined genome and generate much of the ATP necessary to sustain eukaryotic life via an electron transport chain (ETC). Because of the central role for mitochondria in organismal physiology, mitochondrial genetic and phenotypic variation can alter organismal fitness and affect population genetic and evolutionary outcomes. Despite the far-reaching relevance of mitochondria to evolutionary processes and human health, we lack a basic understanding of the causes and consequences of mitochondrial genetic and phenotypic variation. In this thesis, I quantified mitochondrial reactive oxygen species (ROS), membrane potential (δΨM), and mitochondrial morphological traits within Caenorhabditis briggsae natural isolates and mutation-accumulation (MA) lines of both C. briggsae and Caenorhabditis elegans. Substantial natural variation was discovered for most mitochondrial form and function traits measured for a set of C. briggsae isolates known to harbor mitochondrial DNA structural variation in the form of a heteroplasmic nad5 gene deletion (nad5δ) that correlates negatively with organismal fitness. Most among-isolate phenotypic variation could be accounted for by phylogeographic clade membership rather than nad5δ level. Analysis of mitochondrial-nuclear hybrid strains provided support for both mtDNA and nuclear genetic variation as drivers of natural mitochondrial phenotype variation. An MA experimental approach revealed that average levels of both ROS and nad5δ heteroplasmy evolved in remarkably linear ways in C. briggsae maintained under extreme inbreeding. In particular, among C. briggsae isolates prone to acquiring the nad5δ deletion, nad5δ level increased to a plateau of ~50% during successive generations of MA treatment. Conversely, mitochondrial ROS level increased or declined in a strain-specific fashion, which also meant that the relationship between ROS and nad5δ was strain-specific. Further, all lines generated from the isolate with the highest starting level of nad5δ heteroplasmy went extinct prior to generation 20 of MA treatment. Patterns of among-line variance in ROS level were also strain-specific but generally did not conform to the canonical pattern of increasing among-line variance expected for MA experiments. MA lines of C. elegans that had previously been subjected to whole-genome sequencing were found to vary significantly in ROS levels but not in 8-oxo-dG content. Despite a significant positive correlation between 8-oxo-dG and ROS levels, no relationship between oxidative stress measures and base substitution rate or G-to-T transversion rate was revealed. Finally, analysis of patterns of phenotypic correlation for a suite of 24 mitochondrial traits measured in C. briggsae natural isolates support a role for ΔΨM in shaping mitochondrial dynamics, but no such role for mitochondrial ROS. Further, our study suggests a novel model of mitochondrial population dynamics dependent upon cellular environmental context and with implications for mitochondrial genome integrity. This work identifies extensive natural variation and capacity for evolution in organellar traits within multicellular eukaryotic species, with a central role for δΨM in mitochondrial dynamics that may have implications for evolutionary adaptation to thermal niches.

No description available.

Qian, Wei. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 184-199). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Objectives: This investigation adapted the LIVE/DEAD� Baclight[TM] bacterial viability stain for the quantitative determination of bacterial cell viability of the aerotolerant anaerobes Porphyromonas gingivalis ATCC 33277 and Prevotella intermedia ATCC 25611. The Live/Dead stain was used to determine the influence of haem availability on the resistance of P. gingivalis and P. intermedia to the reactive oxygen species (ROS) superoxide anion and hydrogen peroxide and compare the sensitivities between the haem-requiring periodontal bacteria to ROS. Neutrophils use oxidative and non-oxidative killing mechanisms. During phagocytosis, neutrophils kill bacteria via a respiratory burst, producing ROS. P. gingivalis and P. intermedia are oxygen-tolerant gram-negative bacteria found in the gingival crevice. These bacteria express superoxide dismutase (SOD) activity, which extends some protection against superoxide radicals. Methods: Initially, experiments were performed to validate the reliability and accuracy of the fluorogenic Live/Dead stain using Escherichia coli ATCC 10798 (K-12), followed by experiments using P. gingivalis. The Live/Dead stain distinguishes viable:non-viable proportions of bacteria using mixtures of green (SYTO 9) and red (propidium iodide) fluorescent nucleic acid stains respectively. Bacterial cell viability was assessed with fluorescence microscopy and subsequently quantitative measurement using a fluorescence microplate reader (BMG Fluorostar plus Optima). P. gingivalis and P. intermedia colonies were subcultured from frozen cultures, in Tryptic soy broth (TSB) (Difco) and incubated anaerobically for approximately five days. They were further subcultured in pre-reduced TSB, supplemented with menadione 0.5[mu]g/ml (TSB-M) and either 5 [mu]g/ml haemin (Haem 5), 50 [mu]g/ml haemin (Haem 50) or without supplemental haemin (Haem 0). Cultures were grown anaerobically at 37�C to early stationary phase (approximately 48 hours). For experimental purposes, bacteria were harvested, washed and resuspended in 10 mM Tris-buffered saline (pH 7.5) containing peptone (TBS-P) (0.1 mg/ml), with a final adjustment to OD₅₄₀ [approximately equals] 2.0 (which corresponds to 1 x 10⁹ bacteria/ml). Bacterial suspensions were diluted ([approximately equals] 10⁸/ml) into TBS-P containing the fluorogenic viability stain (BacLight, Molecular Probes). Either pyrogallol (0.02 - 2 mM) or hydrogen peroxide (0.01 - 100 mM) was added (except to control tubes); tubes were vortexed for ten seconds and incubated at 37�C. Viability was monitored fluorimetrically for three hours. Results: For both P. gingivalis and P. intermedia, a pyrogallol concentration of 0.2 mM resulted in 80 to 90% cell death; and a hydrogen peroxide concentration of 10 mM killed approximately 80 to 90% of cells. Irrespective of the haem status, no significant difference was determined between the overall maximum rate of killing of P. gingivalis and P. intermedia, in their response to either superoxide or hydrogen peroxide; with the exception that the P. intermedia Haem 0 group was significantly less susceptible to hydrogen peroxide than the P. gingivalis Haem 0 group. For the majority of the experiments, there was no significant difference between final bacterial cell viability in the Haem 0 and Haem 5 cells for both species, after 3 hours exposure to various concentrations of ROS. However, the Haem 50 cells showed a significant increased susceptibility (albeit, a small difference) to both hydrogen peroxide and superoxide. Conclusions: The Live/Dead bacterial viability stain provided a valuable method to monitor "real-time" killing, avoiding the difficulties associated with culture-based methods for assessing viability. Haem availability had no clear influence on the resistance to ROS of either P. gingivalis or P. intermedia Haem 0 and Haem 5 cells. The Haem 50 cells showed a very slight increase in susceptibility to hydrogen peroxide and superoxide. Although P. intermedia may be isolated in significant numbers from healthy gingivae, as well as from periodontally diseased sites, it was no more resistant to ROS than was P. gingivalis, which is associated with periodontal lesions and difficult to cultivate from relatively healthy (more oxygenated) sites. This suggests that resistance to ROS does not contribute to the ecological distinction between these two species. The finding that haem availability did not influence sensitivity implies that these bacteria do not accumulate haem for the purpose of protection from ROS.

The overproduction of reactive oxygen species (ROS) have been linked to diseases and other pathologies. As therapeutic agents, antioxidants have been tested and some shown to attenuate these diseases by relieving oxidative stress. The May laboratory has previously developed a family of phenylaminoalkyl selenides and has demonstrated the antihypertensive and antioxidant properties of these compounds. To further understand the antioxidant property of these selenide compounds, the two step mechanism of the reaction between the selenoxide form and glutathione was investigated by stopped-flow and mass spectrometry, leading to the detection and characterization of a novel thioselenurane intermediate. Mass spectrometry studies supported the redox cycle of the selenide compounds as a straightforward cycle with no byproducts or side reactions and was the first evidence reported of a thioselenurane intermediate present in a reduction reaction of a selenoxide. The therapeutic potential of these compounds was further supported by cell and histological studies demonstrating their ability to alleviate the cardiotoxic effect of anthracyclines without affecting the anti-cancer property of the drugs. Codosage of a phenylaminoethyl selenide with Doxorubicin decreased the infiltration of inflammation cells in the myocardium of mice. Phenylaminoethyl selenides were also able to maintain the body weight of mice treated with Doxorubicin, compared to mice treated with Doxorubicin alone. In order to make the possibility of using Phenylaminoalkyl selenides as therapeutic agents or supplements with other agents, delivery of the compounds was investigated. N acetyl phenylaminoethyl selenides were successfully encapsulated into poly(lactic-co-glycolic) (PLGA) nanoparticles using the nanoprecipitation technique. An attempt was made to demonstrate the ability of these selenide- nanoparticles to reduce cellular oxidative stress caused by incubation with LPS. Future studies are needed to optimize the loading of the selenide compounds into nanocarriers and to demonstrate the ability of the encapsulated drug to work as the free drug. The long term goal of this research is to fully understand the potential of phenylaminoalkyl selenides as an efficient therapeutic agent for ailments derived from increased levels of ROS and a state of oxidative stress. As a supplemental project funded by the National Science Foundation, the Center for Innovative Biomaterial Education and Research (CIBER) was created. Enzymatically catalyzed reaction and polymerizations were investigated using Candida antarctica Lipase B (CALB). Several CALB catalyzed Michael addition reactions were successful and yielded compounds that could be used as future reactants and monomers. As an education requirement of the project a website was created in order to educate the public of the importance, sources and uses of biomaterials. The website provides information for all levels of students and educators. This center has allowed The Georgia Institute of Technology to form relationships and exchange programs with leading universities around the world allowing the exchange of knowledge and research in biomaterials.

Thesis (Ph. D.)--Oregon State University, 2010. / Printout. Includes bibliographical references (leaves 190-208). Also available on the World Wide Web.

Thesis (Ph. D.)--West Virginia University, 2002. / Title from document title page. Document formatted into pages; contains xi, 148 p. : ill. (some col.). Vita. Includes abstract. Includes bibliographical references.