Tracing Nitrogen Metabolism in Mouse Tissues with Gas Chromatography-Mass Spectrometry
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Author:
Date:
2021-02-20
[Abstract] Nitrogen-containing metabolites including ammonia, amino acids, and nucleotides, are essential for cell metabolism, growth, and neural transmission. Nitrogen metabolism is tightly coordinated with carbon metabolism in the breakdown and biosynthesis of amino acids and nucleotides. Both nuclear magnetic resonance spectroscopy and mass spectrometry including gas chromatography-mass spectrometry (GC MS) and liquid chromatography (LC MS) have been used to measure nitrogen metabolism. Here we describe a protocol to trace nitrogen metabolism in multiple mouse tissues using 15N-ammonia coupled with ...
[摘要] [摘要]含氮代谢物,包括氨,氨基酸和核苷酸,对于细胞代谢,生长和神经传递至关重要。在氨基酸和核苷酸的分解和生物合成中,氮代谢与碳代谢紧密相关。核磁共振光谱法和质谱法(包括气相色谱-质谱法(GC MS)和液相色谱法(LC MS))均已用于测量氮代谢。在这里,我们描述了使用15 N氨气与GC MS结合追踪多种小鼠组织中氮代谢的方案。该协议在 包括示踪剂注射,组织制备,代谢物提取,GC MS分析和自然丰度校正的详细程序。该协议将为研究代谢活性组织(例如视网膜,脑,肝和肿瘤)中组织特异性氮提供有用的工具。
[背景]核磁共振波谱法(NMR)和质谱法(包括质谱法(GC MS)和液相色谱法(LC MS))已成功用于各种生物体的全系统代谢物测量(Fiehn,2002和20 1 6; Chokkathukalam等,2014)。但是,每种方法都有其局限性,取决于研究类型,包括绝对定量,代谢物性质,灵敏度,稳健性,同位素分析和成本效益(Chokkathukalam等人,2014)。与NMR相比,基于MS的方法具有更高的灵敏度和代谢物覆盖率,因此更加常用。稳定的同位素标记加上LC MS或GC MS可以灵敏地定量健康和患病组织或细胞中的动态代谢变化(Jang等人,2018)。LC MS与稳定的氮同位素结合揭示了谷氨酰胺氮代谢如何与癌细胞中的碳代谢协调(Wang等人,2019)。GC ...
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Pipecolic Acid Quantification Using Gas Chromatography-coupled Mass Spectrometry
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Author:
Date:
2020-12-05
[Abstract] Pipecolic acid (Pip), a non-proteinacious product of lysine catabolism, is an important regulator of immunity in plants and humans alike. For instance, Pip accumulation is associated with the genetic disorder Zellweger syndrome, chronic liver diseases, and pyridoxine-dependent epilepsy in humans. In plants, Pip accumulates upon pathogen infection and is required for plant defense. The aminotransferase ALD1 catalyzes biosynthesis of Pip precursor piperideine-2-carboxylic acid, which is converted to Pip via ornithine cyclodeaminase. A variety of methods are used to quantify Pip, and some of ...
[摘要] [摘要]胡椒酸(Pip)是赖氨酸分解代谢的非蛋白质产物,是植物和人类免疫力的重要调节剂。例如,Pip积累与人类遗传病Zellweger综合征,慢性肝病和吡ido醇依赖性癫痫有关。在植物中,Pip会在病原体感染时积聚,是植物防御所必需的。氨基转移酶ALD1催化Pip前体哌啶-2-羧酸的生物合成,后者通过鸟氨酸环脱氨酶转化为Pip。多种方法可用于定量Pip,其中一些方法涉及使用昂贵的氨基酸分析试剂盒。在这里,我们描述了从植物组织中定量定量分析的简化程序。P ipecolic酸是从叶组织中提取与内标正缬氨酸一起衍生与丙基氯甲酸酯和分析通过气相色谱-质谱联用选择性离子模式。此过程简单,经济且有效,并且不涉及同位素内标或多步衍生化。
关键词:胡椒酸GC-MS拟南芥植物防御氯甲酸丙酯
[背景]匹,非蛋白氨基酸赖氨酸的氨基酸衍生物,积聚在病原体感染的植物的本地和远端的组织和患者的体液与过氧化物酶体疾病(Schutgens等人,1986;尹和An,2010;Návarová等人。,2012;王等人,2018) ...
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Gas Chromatography Detection Protocol of Short-chain Fatty Acids in Mice Feces
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Author:
Date:
2020-07-05
[Abstract] Short-chain fatty acids (SCFAs), which are formed mainly by bacteria fermenting undigested carbohydrates in the colon, they are based on the number of carbon atoms in the carbon chain. Organic fatty acids with less than 6 carbon atoms are called short-chain fatty acids. SCFAs are closely related to various aspects of the human body, so more and more researchers concentrate on SCFAs. This protocol describes, a direct injection gas chromatography detection method with a pretreatment method for extracting SCFA from mice feces by combining acidification. The corresponding sample limit of ...
[摘要] [摘要] 短链脂肪酸(SCFAs)主要由细菌在结肠中发酵未消化的碳水化合物而形成,它们的基础是碳链中碳原子的数量。碳原子少于6个的有机脂肪酸称为短链脂肪酸。SCFAs与人体的各个方面密切相关,因此SCFAs的研究越来越多。本方案描述了一种直接进样气相色谱检测法和预处理法相结合从小鼠粪便中提取SCFA的方法。相应的定量限(LOQ)和检测限(LOD)分别为0.8-1.0 mg/L和0.5-0.8 mg/L。校准曲线的相关系数大于0.999。加标回收率为80%~102%。该方法可用于小鼠粪便中SCFAs的分析测定。因此,这是一种经济、有效、重复性好的小鼠SCFAs测定方法。
[背景]短链脂肪酸(SCFA)是调节和控制生物功能的重要分子(Siri Tarino et al.,2015)。短链脂肪酸通常指含有少于六个碳原子的挥发性有机脂肪酸(Wong等人,2006年;Marette和Jobin,2015年)。主要包括异丁酸、异丁酸、异丁酸。SCFAs主要由肠道微生物群产生,在结肠中发酵未消化的碳水化合物和膳食纤维。(Henningsson等人,2001年)。许多研究表明,支链脂肪酸来自支链氨基酸和亮氨酸(Koh等人,2016)。少量支链脂肪酸是通过发酵未消化的蛋白质产生的(Cummings and ...
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