本文關(guān)鍵詞： 先天性畸形 代謝組學(xué) 超高效液相色譜聯(lián)合質(zhì)譜 氣相色譜串聯(lián)質(zhì)譜 模式識(shí)別　出處：《重慶醫(yī)科大學(xué)》2014年碩士論文　論文類(lèi)型：學(xué)位論文

【摘要】：代謝組學(xué)(Metabolomics或Metabonomics)是近年來(lái)在系統(tǒng)生物學(xué)蓬勃開(kāi)展的背景下發(fā)展起來(lái)的一門(mén)新興學(xué)科。目前國(guó)內(nèi)外將代謝組學(xué)分為4個(gè)層次：代謝物靶標(biāo)分析、代謝輪廓(譜)分析、代謝組學(xué)和代謝指紋分析。其中，代謝指紋分析(metabolic finger printing)是以整個(gè)圖譜(指紋圖譜)代表特定細(xì)胞或組織的特定代謝模式,不分離鑒定具體單一組分,是一種快速、高通量、全局分析手段,因此樣本制備相對(duì)簡(jiǎn)單。代謝指紋分析通常結(jié)合模式識(shí)別和判別技術(shù)用來(lái)分類(lèi)代謝指紋和識(shí)別不同模式指紋的某些特征,是在不同狀態(tài)判別篩選、疾病診斷及其特定代謝模式的發(fā)現(xiàn)中最有用的方法。代謝指紋分析能通過(guò)反映整體的代謝指紋圖譜監(jiān)測(cè)體內(nèi)代謝網(wǎng)絡(luò)的運(yùn)行狀態(tài)、診斷疾病、評(píng)價(jià)療效、預(yù)測(cè)人體未來(lái)的健康發(fā)展趨勢(shì)。因此，代謝組學(xué)研究應(yīng)用于產(chǎn)前診斷具有十分重要意義，本文建立了兩種方法（超高效液相色譜聯(lián)合質(zhì)譜以及氣相色譜質(zhì)譜）對(duì)先天性畸形羊水代謝進(jìn)行詳細(xì)研究，內(nèi)容如下。 1.基于超高效液相色譜聯(lián)合質(zhì)譜對(duì)先天性畸形羊水代謝組學(xué)輪廓研究 先天性畸形是嬰兒死亡主要原因之一，它可能與基因、環(huán)境等因素有關(guān)，但通過(guò)傳統(tǒng)檢查方法很難解釋其發(fā)病機(jī)制。代謝組學(xué)研究生物系統(tǒng)變化，能夠發(fā)現(xiàn)與疾病相關(guān)的小分子。因此本文建立基于超高效液相色譜聯(lián)合質(zhì)譜（UPLC-MS）聯(lián)合多種識(shí)別模式對(duì)先天性畸形羊水和正常羊水代謝組學(xué)輪廓研究，鑒定出14種物質(zhì)，它們都涉及重要的代謝途徑，包括亮氨酸代謝、谷氨酸代謝、絲氨酸代謝、組氨酸、精氨酸、苯丙氨酸代謝以及能量代謝。這些發(fā)現(xiàn)可能對(duì)于研究先天性畸形發(fā)病機(jī)制有一定價(jià)值，，同時(shí)可以為產(chǎn)前診斷提供新的代謝標(biāo)志物。 2.基于氣相色譜串聯(lián)質(zhì)譜對(duì)先天性畸形羊水中氨基酸定量分析的研究 氨基酸在許多代謝通路中有重要作用，如核酸、維生素、蛋白質(zhì)的生物合成。本文通過(guò)運(yùn)用氣相色譜聯(lián)合質(zhì)譜（GC-MS）及柱前三甲基硅烷衍生化檢測(cè)先天性畸形羊水中的氨基酸，最終檢測(cè)出8種氨基酸，其中亮氨酸、異亮氨酸、絲氨酸及蘇氨酸差異性較大，可能與先天性畸形關(guān)系密切。同時(shí)對(duì)衍生化時(shí)間和溫度進(jìn)行優(yōu)化，在最優(yōu)條件下（70℃以及60min），氨基酸在0.5μg/ml-10μg/ml濃度范圍內(nèi)呈線(xiàn)性關(guān)系，相關(guān)系數(shù)大于0.99，該方法的精密度0.84%-9.33%，回收率91.12%-104.41%，該方法靈敏性、精密度較好。研究結(jié)果表明先天性畸形羊水中氨基酸改變可能是其致病發(fā)展的原因之一，該研究結(jié)果可以讓我們多了解先天性畸形發(fā)病機(jī)制。
[Abstract]:Metabolomics or Metabonomicsis a new subject developed in the background of the vigorous development of systems biology in recent years. At present, the credit of metabolic group is divided into four levels at home and abroad: metabolite target analysis, metabolic profile (spectrum) analysis. Metabonomics and fingerprinting. Metabolic finger printings represent a specific metabolic pattern of a particular cell or tissue, and do not separate and identify a single component, which is a fast, high-throughput. Metabolic fingerprint analysis is usually combined with pattern recognition and discriminant technology to classify metabolic fingerprint and identify some characteristics of different pattern fingerprints. The most useful method in disease diagnosis and the discovery of specific metabolic patterns. Metabolic fingerprinting can monitor the operating status of metabolic networks in the body, diagnose diseases, and evaluate the efficacy by reflecting the overall metabolic fingerprint. Therefore, the application of metabonomics in prenatal diagnosis is of great significance. In this paper, two methods (ultra-high performance liquid chromatography combined mass spectrometry and gas chromatography-mass spectrometry) were established to study the amniotic fluid metabolism in congenital malformation. 1. To study the metabolic profile of amniotic fluid in congenital malformation based on ultra-high performance liquid chromatography and mass spectrometry. Congenital malformation is one of the main causes of infant death, it may be related to genes, environment and other factors, but it is difficult to explain its pathogenesis by traditional methods. Therefore, based on UPLC-MS) and multiple recognition patterns, the metabolic profiles of congenital malformed amniotic fluid and normal amniotic fluid were studied, and 14 substances were identified. They all involve important metabolic pathways, including leucine metabolism, glutamate metabolism, serine metabolism, histidine, arginine, These findings may be valuable in studying the pathogenesis of congenital malformation and may provide a new metabolic marker for prenatal diagnosis. 2. Quantitative analysis of amino acids in congenital amniotic fluid based on gas chromatography tandem mass spectrometry. Amino acids play an important role in many metabolic pathways, such as the biosynthesis of nucleic acids, vitamins and proteins. In this paper, amino acids in congenital malformed amniotic fluid were detected by gas chromatography-mass spectrometry (GC-MS) and pre-column trimethylsilane derivation. Finally, eight kinds of amino acids were detected, including leucine, isoleucine, serine and threonine, which may be closely related to congenital malformation. At the same time, the derivation time and temperature were optimized. Under the optimum conditions of 70 鈩

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