NMR Spectroscopy-Based Metabolic Profiling of Biospecimens.
NMR Spectroscopy-Based Metabolic Profiling of Biospecimens.
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DOI：
10.1002/cpps.98
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发表时间：
2019-12-01
影响因子：
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通讯作者：
Weljie, Aalim M
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文献类型：
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作者：
Sengupta, Arjun;Weljie, Aalim M
代谢组学是指对血清、组织和尿液等生物样本中代谢物的研究。核磁共振 (NMR) 光谱和超高效液相色谱-串联质谱（UPLC-MS/MS；质谱与液相色谱联用）最常用于分析复杂的生物/临床样品。与 UPLC-MS/MS 相比，NMR 是一种相对不敏感的工具，但提供简单的定量和鉴定以及简单的样品处理。一维 1 H NMR 光谱本质上是定量的，可以轻松用于代谢物定量，无需单独的代谢物标准品。二维光谱最常用于代谢物的鉴定，但也可用于定量。尽管 NMR 实验对于分析物的化学性质没有偏见，但遵守正确的代谢物提取方案以获得最佳结果至关重要。选择和实施适当的 NMR 脉冲程序也很重要。最后，采用正确的代谢物定量策略也至关重要。在本单元中，介绍了从典型生物样本运行 NMR 代谢组学实验的分步指南。该单元描述了优化的代谢物提取方案，然后使用所谓的“目标分析”技术实施 NMR 实验和量化策略。该方法依赖于在类似条件下获得的代谢物光谱的基础基础集。还提出了一些数据统计分析策略。总的来说，这套协议应该为任何希望进入基于 NMR 的代谢组学分析领域的人提供指导。 © 2019 John Wiley & Sons, Inc. 基本方案 1：从不同生物样本中提取代谢物 基本方案 2：制备用于 NMR 分析的干燥上级分 备用方案：制备用于 NMR 分析的尿液样品 基本方案 3：NMR 实验 基本方案 4：代谢物的光谱处理和定量 基本方案 5：数据的统计分析。
Metabolomics refers to study of metabolites in biospecimens such as blood serum, tissues, and urine. Nuclear magnetic resonance (NMR) spectroscopy and ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS; mass spectrometry coupled with liquid chromatography) are most frequently employed to analyze complex biological/clinical samples. NMR is a relatively insensitive tool compared to UPLC-MS/MS but offers straightforward quantification and identification and easy sample processing. One-dimensional 1 H NMR spectroscopy is inherently quantitative and can be readily used for metabolite quantification without individual metabolite standards. Two-dimensional spectroscopy is most commonly used for identification of metabolites but can also be used quantitatively. Although NMR experiments are unbiased regarding the chemical nature of the analyte, it is crucial to adhere to the proper metabolite extraction protocol for optimum results. Selection and implementation of appropriate NMR pulse programs are also important. Finally, employment of the correct metabolite quantification strategy is crucial as well. In this unit, step-by-step guidance for running an NMR metabolomics experiment from typical biospecimens is presented. The unit describes an optimized metabolite extraction protocol, followed by implementation of NMR experiments and quantification strategies using the so-called "targeted profiling" technique. This approach relies on an underlying basis set of metabolite spectra acquired under similar conditions. Some strategies for statistical analysis of the data are also presented. Overall, this set of protocols should serve as a guide for anyone who wishes to enter the world of NMR-based metabolomics analysis. © 2019 by John Wiley & Sons, Inc. Basic Protocol 1: Metabolite extraction from different biospecimens Basic Protocol 2: Preparation of dried upper fraction for NMR analysis Alternate Protocol: Preparation of urine samples for NMR analysis Basic Protocol 3: NMR experiments Basic Protocol 4: Spectral processing and quantification of metabolites Basic Protocol 5: Statistical analysis of the data.