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Synthesis of an Unnatural Phospholipid for use in Pulmonary Surfactant TherapyBest, Natasha 02 May 2012 (has links)
Neonatal respiratory distress syndrome (RDS) is a disease that affects premature infants born prior to 32 weeks gestation. The main cause is a deficiency in pulmonary surfactant due to immature type II pneumocyte cells found in the alveoli. These cells are not capable of producing the required surfactant which normally functions to reduce the surface tension at the air-liquid interface of the lungs, as well as reduce the work of breathing and prevent alveolar collapse. A current treatment method for RDS is exogenous surfactant replacement therapy involving application of an exogenous surfactant preparation directly into the lungs of premature infants. Current surfactant preparations are animal-derived and very costly. Synthetic preparations, on the other hand, are an attractive alternative. The goal of this research is to synthesize a diether phosphonolipid analogue of dipalmitoyl phosphatidylcholine (DPPC), designated DEPN-8. When incorporated into a synthetic exogenous surfactant mixture, DEPN-8 exhibits greater adsorption and surface activity compared to its natural counterpart, DPPC. The synthesis of several components related to the re-tailored synthesis of DEPN-8 will be presented and discussed below. / National Institute of Health, NSERC
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NMR studies of metabolites and xenobiotics: From time-points to long-term metabolic regulationEhlers, Ina January 2015 (has links)
Chemical species carry information in two dimensions, in their concentrations and their isotopic signatures. The concentrations of metabolites or synthetic compounds describe the composition of a chemical or biological system, while isotopic signatures describe processes in the system by their reaction pathways, regulation, and responses to external stimuli. Stable isotopes are unique tracers of these processes because their natural abundances are modulated by isotope effects occurring in physical processes as well as in chemical reactions. Nuclear magnetic resonance (NMR) spectroscopy is a prime technique not only for identification and quantification of small molecules in complex systems but also for measuring intramolecular distribution of stable isotopes in metabolites and other small molecules. In this thesis, we use quantitative NMR in three fields: in food science, environmental pollutant tracing, and plant-climate science. The phospholipid (PL) composition of food samples is of high interest because of their nutritional value and technological properties. However, the analysis of PLs is difficult as they constitute only a small fraction of the total lipid contents in foods. Here, we developed a method to identify PLs and determine their composition in food samples, by combining a liquid-liquid extraction approach for enriching PLs, with specialized 31P,1H-COSY NMR experiments to identify and quantify PLs. Wide-spread pollution with synthetic compounds threatens the environment and human health. However, the fate of pollutants in the environment is often poorly understood. Using quantitative deuterium NMR spectroscopy, we showed for the nitrosamine NDMA and the pesticide DDT how intramolecular distributions (isotopomer patterns) of the heavy hydrogen isotope deuterium reveal mechanistic insight into transformation pathways of pollutants and organic compounds in general. Intramolecular isotope distributions can be used to trace a pollutant’s origin, to understand its environmental transformation pathways and to evaluate remediation approaches. The atmospheric CO2 concentration ([CO2]) is currently rising at an unprecedented rate and plant responses to this increase in [CO2] influence the global carbon cycle and will determine future plant productivity. To investigate long-term plant responses, we developed a method to elucidate metabolic fluxes from intramolecular deuterium distributions of metabolites that can be extracted from historic plant material. We show that the intramolecular deuterium distribution of plant glucose depends on growth [CO2] and reflects the magnitude of photorespiration, an important side reaction of photosynthesis. In historic plant samples, we observe that photorespiration decreased in annual crop plants and natural vegetation over the past century, with no observable acclimation, implying that photosynthesis increased. In tree-ring samples from all continents covering the past 60 – 700 years, we detected a significantly smaller decrease in photorespiration than expected. We conclude that the expected “CO2 fertilization” has occurred but was significantly less pronounced in trees, due to opposing effects. The presented applications show that intramolecular isotope distributions not only provide information about the origin and turnover of compounds but also about metabolic regulation. By extracting isotope distributions from archives of plant material, metabolic information can be obtained retrospectively, which allows studies over decades to millennia, timescales that are inaccessible with manipulation experiments.
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Electrostatic Modification of Phospholipid and Lipopolysaccharide MembranesMa, Zheng 22 May 2012 (has links)
Biological membranes are quasi two-dimensional self-assembled structure, primarily serving as a barrier to the leakage of cell’s contents. The main constituents of biological membrane are various amphiphilic lipids that form bilayers in an aqueous environment. These lipids carry acidic and/or basic functional groups that ionize in water, giving some of them a net electrical charge. Such a lipid molecule, when integrated into the membrane, experiences electrostatic forces from all other charged objects around it, including ions, surrounding lipids, and other molecules such as cationic peptides. The electrostatic interaction can profoundly influence the membrane, to which many phenomena with physiological significance as well as biophysical interest can be ascribed.
In this thesis, we concentrate on investigating the electrostatic properties of lipid membranes. First, we study how the electrostatic interaction affects their preferred structure. To this end, we adopt a coarse-grain model that preserves the dominant characteristics of the lipids, in which the electrostatic interaction is treated within the “renormalized” Debye-H¨uckel theory. In particular, we calculate the spontaneous curvature of a phospholipid monolayer, along with other associated quantities. Our results suggest that such divalent ions as Mg2+ can stabilize HII phases of lipids (inverted hexagonal phases), which would otherwise form lamellar phases.
Second,we investigate the competitive binding of ions and cationic peptides onto a monolayer of lipopolysaccharide (LPS) molecules, a class of highly charged bio-molecules found in the outer leaflet of the outer membranes of gram-negative (G-) bacteria. Cationic anti-microbial peptides (AMPs) can selectively kill bacteria, and it is suggested that they destabilize the LPS layer, easing their permeation across it, a process of great physiological and clinical interest. To this end, we model the LPS layer as a collection of charged “binding sites”, based on which we study the binding of cations (monovalent and divalent) and cationic peptides onto the layer. Our calculations suggest that the peptides can compete with divalent ions on the binding to the layer. It has been empirically known that since the stability of an LPS layer relies greatly on the bridging of divalent ions, the substitution of these ions by the peptides significantly compromises its stability. Our results offer a quantitative basis for this observation, thus providing a possible mechanism of an important step in the action of AMPs against G- bacteria.
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Investigations on beta 2-glycoprotein I and antiphospholipid antibodiesGiannakopoulos, Bill, Clinical School - St George Hospital, Faculty of Medicine, UNSW January 2008 (has links)
An outline of the work contained in this thesis is presented. The first chapter is a critical review of the literature pertaining to the pathophysiological mechanisms operational with regards to the antiphospholipid syndrome (APS). The syndrome is characterised by venous and arterial thrombosis, and recurrent fetal loss, in association with the persistent presence of antibodies targeting the main autoantigen beta 2-glycoprotein I (β2GPI). The second chapter reviews the literature delineating the diverse physiological functions of β2GPI, and then relates them to its role in our current understanding of the pathophysiology of APS. The third chapter presents a critical review of the evidence base for the diagnosis and management of APS. The fourth chapter describes the interaction between β2GPI and the glycoprotein Ib alpha (GPIbα) subunit of the platelet receptor GPIb-IX-V. GPIbα is an important platelet adhesion receptor, which mediates multiple additional functions on the platelet surface, including binding coagulation factor XI (FXI). The implication of the interaction between β2GPI and GPIbα on platelet activation and the release of thromboxane in the presence of anti-β2GPI antibodies is explored, as well as the intracellular pathways via which this activation occurs. The relevance of these findings to understanding APS pathogenesis, in particular thrombosis, is discussed. The fifth chapter delineates the interaction between the fifth domain of β2GPI and FXI and its activated form factor XIa (FXIa). The ability of FXIa to cleave β2GPI between lysine (Lys) 317 and threonine (Thr) 318, and modulate its function is reported. The sixth chapter describes the ability of β2GPI to inhibit FXIa autoproteolytic hydrolysis at the specific FXIa residues arginine (Arg) 507, Arg532 and Lys539. This interaction with β2GPI stabilizes FXIa activity over time, and leads to enhanced FXIa mediated fibrin formation. This is a novel physiological function of β2GPI with important implications. Recent epidemiological studies by others have emphasized the critical role of FXIa in pathological thrombus propagation. The seventh chapter defines the relevance of the FXIa residues Arg507, Arg532 and Lys539 to FXIa mediated inactivation by the main FXIa inhibitor Protease Nexin 2 (PN2), and by Antithrombin III (ATIII). Insights into future directions for research are presented and discussed within each individual chapter.
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Characterizations of the anti-obesity and anti-adipogenic effects of the limonoid prieurianinSaunders, Rudel Anton. January 2010 (has links)
Dissertation (Ph.D.)--University of Toledo, 2010. / "Submitted to the Graduate Faculty as partial fulfillment of the requirement for the Doctoral of Philosophy Degree in Biomedical Sciences." Title from title page of PDF document. "A Dissertation entitled"--at head of title. Bibliography: p. 80-98.
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Various aspects of soil microbial ecology as revealed by phospholipid fatty acid (PLFA) analysis.KOTAS, Petr January 2018 (has links)
The PLFA profiling method was adopted and used to determine changes in microbial community structure and abundance along natural and human-induced environmental gradients. The presented studies were based on field sampling campaigns combined with targeted laboratory experiments. According to the aims of particular studies, microbial PLFA fingerprinting was combined with the auxiliary below- and aboveground ecosystem characteristics to identify the drivers of microbial responses to environmental changes or with 13C-labelling and metagenomics to obtain more complex information about running processes and involved microorganisms.
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Design and Synthesis of a Macrocyclic PhospholipidMitchell, Gavin Maxwell 08 September 2014 (has links)
The membrane-spanning phospholipids of the domain Archaea are postulated to provide membrane stability; this thesis reports the design and synthesis of a synthetic membrane-spanning macrocyclic lipid to test this hypothesis. Protected glutaric anhydride reacted with 10-undecyn-1-ol to produce a glutarate monoester. Copper-mediated azide-alkyne coupling (CuAAC) using 1,5-diazido-3-oxapentane(bis-azide) afforded a dicarboxylicacid with a hydrophobic chain of sufficient length to span a 35 Å bilayer membrane. The carboxylic acids were each esterified with an equivalent of 10-undecyn-1-ol. After optimization an 87% yield was obtained in the closure of the 72-membered macrocycle with the bis-azide using CuAAC. Deprotection and coupling with p-nitrophenyl phosphorodichloridate completed the synthesis. Two other phospholipids, a linear bolaamphiphile derived from the precursor to the second click reaction, and a linear amphiphile created from a glutarate bis-dodecyl ester, were also synthesized to provide controls for probing the orientation of the macrocyclic phospholipid in the bilayer membrane of vesicles. The amphiphile, linear bolaamphiphile, and macrocyclic bolaamphiphile were synthesized in 4, 5, and 6 steps with yields of 19, 4, and 5% respectively.
The hydrophilic head group of the macrocyclic phospholipid was designed to release p-nitrophenolate in basic conditions from the p-nitrophenylphosphate head group to produce an absorbance at 400 nm. This assay was expected to elucidate the membrane-spanning orientation of the phospholipid in the bilayer membrane of vesicles. The final target compound failed to release p-nitrophenolate under basic conditions and underwent phosphate elimination to produce an α, β-unsaturated ester instead. Although the macrocyclic lipid produced associates with membranes and may be membrane-spanning, this lipid design was unable to reveal its membrane orientation. / Graduate
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Studies of lysophosphatidic acid acyltransferases generating membrane lipid diversity in bacteria / 細菌膜脂質の多様性を形成するリゾホスファチジン酸アシル基転移酵素群に関する研究Toyotake, Yosuke 25 March 2019 (has links)
京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第21841号 / 農博第2354号 / 新制||農||1069(附属図書館) / 学位論文||H31||N5213(農学部図書室) / 京都大学大学院農学研究科応用生命科学専攻 / (主査)教授 栗原 達夫, 教授 植田 充美, 教授 小川 順 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DGAM
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Tocopherol regeneration by phospholipids in soybean oil-in-water emulsions: effect of tocopherol homologue and emulsifier typeSamdani, Gautam 21 March 2018 (has links)
Phospholipids can regenerate oxidized tocopherols and help delay lipid oxidation. The impact of emulsifier type, tocopherol homologue and phospholipid head group on tocopherol-phospholipid interaction was investigated in this study.
Three µmol tocopherol/kg emulsion and 15.0µmol/kg emulsion of PE or PS were dissolved in oil and emulsions were prepared. Tween 20 or bovine serum albumin(BSA) was used as emulsifier and the continuous phase contained 10mM imidazole/acetate buffer at pH 7. Lipid hydroperoxides and hexanal were measured as lipid oxidation products and the lag phase was determined. With Tween 20 as the emulsifier, α and δ-tocopherol had a hexanal lag phase of 2 and 4 days respectively. PE and PS both extended the lag phase to 7 and 10 days respectively in presence of δ-tocopherol. Whereas, PS extended the lag phase to 6 days and PE could not exhibit any synergism with α-tocopherol. With BSA as the emulsifier, α and δ-tocopherol had a lag phase of 4 days. PE and PS extended the lag phase to 11 days and 10 days respectively in presence of δ-tocopherol and to 7 and 8 days respectively in presence of α-tocopherol. PE and PS both exhibited synergism with mixed tocopherol and the extent of synergism was in less than δ-tocopherol but more than α-tocopherol.
Phospholipids could potentially be used with tocopherols to improve the oxidative stability of emulsions. PE was more effective with BSA whereas PS was equally effective with both emulsifiers.
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Preparation, Characterization, and Use of Antioxidant-LiposomesYang, Hongsong, Paromov, Victor, Smith, Milton, Stone, William L. 01 December 2008 (has links)
Antioxidant liposomes provide a unique means of delivering both water and/or lipid soluble antioxidants to tissues thereby affecting disease states or signal transduction pathways modulated by oxidative stress. Considerable evidence suggests that liposome-encapsulated antioxidants can be superior to the corresponding free antioxidants in this regard. This chapter will provide practical details on the preparation, characterization, and use of antioxidant liposomes. Methods will be described for the small-scale preparation (1 ml) and large-scale (100 ml/hour) preparation of antioxidant liposomes as well as the techniques for characterizing their size distribution and their physical and chemical stability. The use of antioxidant liposomes in an in vitro situation will also be detailed.
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