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Metabolipidomická analýza bílé tukové tkáně pomocí UPLC-MS/MS / Metabolipidomic profiling of white adipose tissue by UPLC-MS/MSRombaldová, Martina January 2019 (has links)
Obesity is a serious problem in society today [1,2]. It might seems to have been caused simply by excess consumption of food compared to energy expenditure but obesity is actually a complex metabolic disorder centred on adipose lipid metabolism and cellular signalling systems linked to it [3]. Understanding the biology of adipose tissue (AT) is very important for the identification of novel and potential therapeutic targets in order to prevent and treat obesity-related disorders [4]. We utilized analytical approach liquid chromatography coupled to mass spectrometry (LC-MS) to study adipose tissue metabolism. Also, we were especially interested in the effect of omega-3 polyunsaturated fatty acids (PUFA) on that metabolism. Rodent and cell line experiments were performed and analyses were done of white adipose tissue (WAT), serum/plasma samples or cells as well as milk samples from mothers. At first, we established several ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) methods for analysis of acylcarnitines (AC), amino acids (AmA) and other metabolites. Importantly, these methods were able to distinguish isobaric species of AC which is not usually possible. Using these approaches we uncovered several acylcarnitines, i.e. long chain AC, carnitine, acylcarnitine C4 and...
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Metabolomic Assessment of Dietary Interventions in Obesity by Capillary Electrophoresis Mass SpectrometryLam, Karen Phoebe January 2018 (has links)
Capillary electrophoresis mass spectrometry (CE-MS) is a versatile instrumental
method for metabolomics, which allows for comprehensive metabolite profiling of
volume-limited biological specimens in order to better understand the molecular
mechanisms associated with chronic diseases, including an alarming epidemic of
obesity worldwide. Multiplexed CE separations enable high-throughput metabolite
screening with quality assurance to prevent false discoveries when combined with
rigorous method validation, robust experimental designs, complementary statistical
methods, and high-resolution tandem mass spectrometry (MS/MS) for unknown
metabolite identification. In this thesis, multiplexed CE-MS technology is applied for
both targeted and untargeted metabolite profiling of various biological fluids, including
covalently bound thiol-protein conjugates, as well as free circulating metabolites in
serum and plasma, and excreted/bio-transformed compounds in urine due to complex
host-gut microflora co-metabolism. This work was applied to characterize aberrant
metabolic responses of obese subjects in response to dietary challenges, and measure
the benefits of dietary interventions that reduce adiposity without deleterious muscle
loss. Chapter 2 presents, a simple, sensitive yet robust analytical protocol to expand
metabolome coverage in CE-MS for the discovery of labile protein thiols in human
plasma using a rapid chemical derivatization method based on N-tert-butylmaleimide
(NTBM). Chapter 3 describes targeted metabolite profiling of serum and plasma
to investigate the differential metabolic responses between healthy and unhealthy
obese individuals before and after consumption of a standardized high-caloric meal,
respectively. Chapter 4 of this thesis describes an untargeted metabolite profiling
strategy for urine using multisegment-injection (MSI)-CE-MS for elucidating the effects of protein supplementation following a short-term dietary weight-loss intervention
study. This work revealed six urinary metabolites that were classified as top-ranking
treatment response biomarkers useful for discriminating between subjects consuming
carbohydrate (control), soy, and whey supplemented diets. In summary, this thesis
demonstrated the successful implementation of multiplexed CE-MS technology for
biomarker discovery in nutritional-based metabolomic studies as required for more
effective treatment and prevention of obesity for innovations in public health. / Thesis / Doctor of Philosophy (PhD)
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