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A MULTI-DIMENSIONAL METHOD FOR THE ANALYSIS OF HUMAN BLOOD PLASMA METABOLOMEAmoateng, Catherine, Amoateng, Catherine 10 1900 (has links)
<p>The comprehensive analysis of human blood plasma metabolome has been completed using derivatization gas chromatography-mass spectrometry (GC-MS) analysis, liquid chromatography-mass spectrometry (LC-MS) analysis and a comprehensive LC-GC-MS analysis approach wherein LC fractions were collected, derivatized and analyzed using GC-MS. In all cases blood plasma samples were deproteinized using solvent precipitation prior to chromatography and MS analysis.</p> <p>In GC-MS analyses, the progress of all derivatization reactions was monitored by adding 9-anthracenemethanol and 1,3-diphenylacetone to all reaction mixtures; their conversions to 9-anthracenemethanol trimethylsilyl ether and the oxime derivative of 1,3-diphenylacetone were used as measures of the completion of these derivatization reactions. Any reactions with completions less than 99% were repeated.</p> <p>GC-MS analysis of blood plasma samples detected 100 peaks; 44 were positively identified by comparing retention indices and mass spectra with those of authentic standards. LC-MS analyses were conducted on a HILIC column (aminopropyl phase) with MS detection in both negative ion and positive ion modes and resulted in the identification of 97 peaks; 47 were observed in the positive ion mode, 58 in the negative ion mode with 8 peaks observed in both modes.</p> <p>The multi-dimensional LC-GC approach was not designed as a routine analytical method; rather the purpose of this approach was to see how many compounds could be observed in the sample and to obtain better quality mass spectra and retention index values. The LC separation afforded 16 fractions which upon derivatization GC-MS analysis gave an additional 176 peaks from a total of 276 peaks. The MS data from these additional spectra can be used to develop selected ion monitoring GC-MS or tandem mass spectrometry analytical methods. This thesis has demonstrated the power of off-line comprehensive methods to identify compounds that neither the GC nor the LC methods detected.</p> / Master of Science (MSc)
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A Novel Miniaturised Dynamic Hollow-Fibre Liquid-Phase Micro-Extraction Method for Xenobiotics in Human Plasma SamplesHansson, Helena January 2010 (has links)
Bioanalytical chemistry is a challenging field, often involving complex samples, such as blood, plasma, serum or urine. In many applications, sample cleanup is the most demanding and time-consuming step. In the work underlying this thesis a novel dynamic miniature extractor, known as a hollow-fibre liquid-phase microextractor (HF-LPME), was designed, evaluated and studied closely when used to clean plasma samples. Aqueous-organic-aqueous liquid extraction, in which the organic liquid is immobilised in a porous polypropylene membrane, was the principle upon which the extractor was based, and this is discussed in all the papers associated with this thesis. This type of extraction is known as supported-liquid membrane extraction (SLM). The aim of this work was the development of a dynamic system for SLM. It was essential that the system could handle small sample volumes and had the potential for hyphenations and on-line connections to, for instance, LC/electrospray-MS. The design of a miniaturised HF-LPME device is presented in Paper I. The extraction method was developed for some weakly acidic pesticides and these were also used for evaluation. In the work described in Paper II, the method was optimised on the basis of an experimental design using spiked human plasma samples. Paper III presents a detailed study of the mass-transfer over the liquid membrane. The diffusion through the membrane pores was illustrated by a computer-simulation. Not surprisingly, the more lipophilic, the greater the retention of the compounds, as a result of dispersive forces. The main focus of the work described in Paper IV was to make the HF/LPME system more versatile and user-friendly; therefore, the extractor was automated by hyphenation to a SIA system. / At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 4: Manuscript.
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Core–shell bioprinting of vascularized in vitro liver sinusoid modelsTaymour, Rania, Chicaiza-Cabezas, Nathaly Alejandra, Gelinsky, Michael, Lode, Anja 18 April 2024 (has links)
In vitro liver models allow the investigation of the cell behavior in disease conditions or in response to changes in the microenvironment. A major challenge in liver tissue engineering is to mimic the tissue-level complexity: besides the selection of suitable biomaterial(s) replacing the extracellular matrix (ECM) and cell sources, the three-dimensional (3D) microarchitecture defined by the fabrication method is a critical factor to achieve functional constructs. In this study, coaxial extrusion-based 3D bioprinting has been applied to develop a liver sinusoid-like model that consists of a core compartment containing pre-vascular structures and a shell compartment containing hepatocytes. The shell ink was composed of alginate and methylcellulose (algMC), dissolved in human fresh frozen plasma. The algMC blend conferred high printing fidelity and stability to the core–shell constructs and the plasma as biologically active component enhanced viability and supported cluster formation and biomarker expression of HepG2 embedded in the shell. For the core, a natural ECM-like ink based on angiogenesis-supporting collagen-fibrin (CF) matrices was developed; the addition of gelatin (G) enabled 3D printing in combination with the plasma-algMC shell ink. Human endothelial cells, laden in the CFG core ink together with human fibroblasts as supportive cells, formed a pre-vascular network in the core in the absence and presence of HepG2 in the shell. The cellular interactions occurring in the triple culture model enhanced the albumin secretion. In conclusion, core–shell bioprinting was shown to be a valuable tool to study cell–cell-interactions and to develop complex tissue-like models.
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