Infrared laser spectroscopy of mass-separated metabolites

质量分离代谢物的红外激光光谱

• 批准号: 8829876
• 负责人: Nicolas C Polfer
• 金额: $ 17.34万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2018-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8829876
• 关键词: Address; Algorithms; Base Sequence; Benchmarking; Binding; Biochemical; Bioinformatics; Biological; Biological Markers; Chemicals; Colon Carcinoma; Complex; Complex Mixtures; Computational algorithm; Coupling; Custom; Data; Databases; Development; Diagnostic; Disease; Fingerprint; Freezing; Goals; Gold; Health; Ions; Lasers; Link; Liquid Chromatography; Mass Spectrum Analysis; Measurement; Methodology; Methods; Molecular; Molecular Conformation; Pattern Recognition; Prevention; Process; Proteomics; Research; Resolution; Sampling; Spectrum Analysis; Structure; Techniques; Technology; Training; absorption; base; complex biological systems; cryogenics; infrared spectroscopy; innovation; insight; instrumentation; metabolomics; molecular dynamics; new technology; novel; quantum; research study; tandem mass spectrometry; tool; vibration

DESCRIPTION (provided by applicant): Challenge: Developments of enabling technologies underpin continuing advances in biomolecular research. For instance, mass spectrometry (MS)-based sequencing techniques have spurned proteomics research in the past decade. Currently, there is no "gold standard" technique in metabolomics that allows a routine characterization of the thousands of constituents contained in these samples. NMR is limited to the more abundant analytes due to sensitivity issues. On the other hand, MS is typically capable of detecting many more features, but is often not able to structurally characterize these molecules. Rationale: By coupling tunable infrared (IR) lasers to mass spectrometry instrumentation, the IR spectra of mass- separated ions can be recorded. IR laser spectroscopy of ions combines the high sensitivity and ability to analyze complex mixtures of MS with the enhanced structural information from vibrational spectroscopy. The technique hence allows a chemical elucidation of many unknowns based on diagnostic vibrations and IR spectral fingerprints. Aim 1: Development of cryogenic mass spectrometry and multiplexed IR spectroscopy. In order to make IR spectroscopy a useful bioanalytical tool for biomolecular ions, it is essential that the IR spectra of analytes are well- resolved, and thus distinguishable, and that multiple analytes in mixtures can be probed simultaneously in a multiplexed fashion. We propose to develop a custom-built, cryogenic linear ion trap, where the ions are tagged with weakly-bound molecules (e.g. N2), which are selectively detached upon resonant IR absorption. Aim 2: IR spectroscopy of mass-separated metabolites. Our application of IR spectroscopy of biomolecules focuses on metabolites, where we expect the technique to have most potential. Control experiments on standard metabolites will establish how many analytes can be successfully probed in a multiplexed approach. The methodology will then be applied to selected metabolite samples from colon cancer studies, which have previously been analyzed by high- throughput liquid chromatography and high-resolution mass spectrometry. Aim 3: Structural elucidation of unknown metabolites by comparison to computed IR spectra and bioinformatics approaches. The ultimate goal of this proposal is to chemically characterize unknown biomarkers that cannot be identified by current MS approaches. This requires a comparison of the experimental data for each analyte, namely its mass and its IR spectrum, to putative matches from metabolite databases. The IR spectra of known standards (from aim 2) will serve as a training set and as a benchmark for implementing this identification methodology. Innovation and Impact: The techniques developed here are expected to have the largest impact in global metabolomics, where current tandem mass spectrometry methodologies limit the number of constituents that can be identified in these mixtures. We expect the enhanced structural information from vibrational spectroscopy to yield many new insights in biomarker discovery.

描述（由申请人提供）： 挑战：使能技术的发展支撑着生物分子研究的不断进步。例如，在过去十年中，基于质谱 (MS) 的测序技术已经抛弃了蛋白质组学研究。目前，代谢组学中还没有“金标准”技术可以对这些样品中包含的数千种成分进行常规表征。由于灵敏度问题，NMR 仅限于更丰富的分析物。另一方面，MS 通常能够检测更多特征，但通常无法表征这些分子的结构。原理：通过将可调谐红外 (IR) 激光器与质谱仪器耦合，可以记录质量分离离子的红外光谱。离子红外激光光谱结合了 MS 的高灵敏度和分析复杂混合物的能力以及振动光谱的增强结构信息。因此，该技术可以根据诊断振动和红外光谱指纹对许多未知物进行化学阐明。目标 1：开发低温质谱和多重红外光谱。为了使红外光谱成为生物分子离子的有用生物分析工具，红外光谱至关重要 分析物的光谱分辨率良好，因此可区分，并且可以以多重方式同时探测混合物中的多种分析物。我们建议开发一种定制的低温线性离子阱，其中离子被弱结合分子（例如 N2）标记，这些分子在共振红外吸收时选择性分离。目标 2：质量分离的代谢物的红外光谱。我们对生物分子红外光谱的应用主要集中在代谢物上，我们预计该技术在代谢物方面最具潜力。标准代谢物的对照实验将确定在多重方法中可以成功探测多少分析物。然后，该方法将应用于结肠癌研究中选定的代谢物样本，这些样本之前已通过高通量液相色谱法和高分辨率质谱法进行了分析。目标 3：通过与计算的红外光谱和生物信息学方法进行比较，阐明未知代谢物的结构。该提案的最终目标是对当前 MS 方法无法识别的未知生物标志物进行化学表征。这需要将每种分析物的实验数据（即其质量和红外光谱）与代谢物数据库中的假定匹配进行比较。已知标准的红外光谱（来自目标 2）将作为训练集和实施该识别方法的基准。创新和影响：这里开发的技术预计将对全球代谢组学产生最大的影响，目前的串联质谱方法限制了这些混合物中可以识别的成分数量。我们期望振动光谱中增强的结构信息能够在生物标志物发现方面产生许多新的见解。