LC-Mass Spectrometer for Intramolecular Isotope Ratios at Natural Isotopic Abundance

化合物特异性同位素分析（CSIA）已经彻底改变了环境中有机污染物降解的检测和表征。在线转换的气相或液相色谱（GC， LC）序列和随后的同位素比值质谱（IRMS）允许访问以前未开发的信息来源：分子内自然存在的稳定同位素（13C/12C， 15N/14N等）的比例。同位素指纹图谱可以在地下水污染的情况下阐明有机化学物质的不同来源。污染物同位素比率随时间和空间的逐渐变化可以检测污染物的自然降解，甚至潜在的转化机制。然而，环境化学的研究已经从CSIA的成功故事（即低分子量的非极性高浓度遗留污染物）的基础上转移到探索高分子量和低浓度的更多极性化学物质（例如农药或药物）的行为。这缩小了CSIA的机会之窗。首先，高分子量极性化合物通常与气相色谱不相容，需要使用有机洗脱液进行液相色谱。这目前排除了这些目标化合物的CSIA。其次，降解的同位素效应通常只发生在一个分子位置，并在分子大小增加时被“稀释”。这给基于化合物平均同位素分馏来评估较大的化合物带来了内在的限制。第三，高分子量的分子为同位素指纹提供了更容易区分的分子位置。当只分析复合平均时，这个强大的信息来源仍然未被开发。因此，我们申请了仪器资助，通过液体注入质谱来探索极性有机分子的片段甚至位置特异性同位素分析，这将使我们能够挖掘以下潜力：(i)碎片化和高质量分辨率可以在定义的分子片段或位置中获取元素特异性同位素信息；（ii）该方法与高分子量的极性化合物兼容。在最近的工作中，该方法已被证明适用于相对简单的化合物，如氧阴离子（如硝酸盐），或利用偶然存在的已知分子内同位素分布标准（如蛋氨酸）。有了所要求的仪器，我们希望率先采用重要环境污染物和生物分子的方法：(i)合成具有已知分子内同位素组成的标准物进行校准；㈡探索同位素指纹法，以区分来源和生物合成通量；（iii）利用特定位置的同位素效应研究转化反应；（iv）探索环境样品中精确同位素分析的下限；(v)在生物学和医学中表征代谢的先锋应用。

Compound-specific isotope analysis (CSIA) has revolutionized the detection and characterization of organic contaminant degradation in the environment. Gas- or liquid-chromatography (GC, LC) in sequence with online conversion and subsequent isotope ratio mass spectrometry (IRMS) has allowed accessing a previously untapped source of information: the ratio of naturally occurring stable isotopes (13C/12C, 15N/14N, etc.) within molecules. Isotopic fingerprinting can elucidate the different origins of organic chemicals in cases of groundwater contamination. Gradual changes in contaminant isotope ratios over time and space can detect natural contaminant degradation, and even underlying transformation mechanisms. However, research in Environmental Chemistry has moved on from the foundation of CSIA’s success story (i.e., non-polar highly concentrated legacy contaminants of low molecular weight) to explore the behavior of more polar chemicals of high molecular weight and at low concentrations (e.g. pesticides or pharmaceuticals). This narrows the window of opportunity for CSIA. First, polar compounds of high molecular weight are often not compatible with GC and require liquid-chromatography involving organic eluents. This presently precludes CSIA of these target compounds. Second, isotope effects of degradation typically occur in only one molecular position and are “diluted out” when molecular size increases. This puts an intrinsic limit to assessing larger compounds based on compound-average isotope fractionation. Third, molecules of higher molecular weight offer more distinguishable molecular positions for isotopic fingerprinting. This powerful source of information remains untapped when only the compound average is analyzed. We, therefore, apply for instrument funding to explore fragment- or even position-specific isotope analysis in polar organic molecules by liquid injection MS. This will allow us to tap the potential that (i) fragmentation and high mass resolution can access element-specific isotopic information in defined molecular fragments or positions and (ii) that the approach is compatible with polar compounds of higher molecular weight. In recent work, the approach has been demonstrated for comparatively simple compounds such as oxyanions (e.g., nitrate), or taking advantage of the fortuitous existence of standards of known intramolecular isotope distribution (e.g., methionine). With the requested instrument we want to spearhead the approach for important environmental pollutants and biomolecules: (i) synthesize standards with known intramolecular isotopic composition for calibration; (ii) explore isotopic fingerprinting to distinguish sources and biosynthetic fluxes; (iii) access position-specific isotope effects to study transformation reactions; (iv) explore lower limits of precise isotope analysis in environmental samples and (v) spearhead applications to characterize metabolism in biology and medicine.