Isotopically labeling a metabolite and tracing its metabolic fate has provided invaluable insights about the role of metabolism in human diseases in addition to a variety of other issues. ¹³C-labeled metabolite tracers or unlabeled ¹H-based NMR experiments are currently the most common application of NMR to metabolomics studies. Unfortunately, the coverage of the metabolome has been consequently limited to the most abundant carbon-containing metabolites. To expand the coverage of the metabolome and enhance the impact of metabolomics studies, we present a protocol for ¹⁵N-labeled metabolite tracer experiments that may also be combined with routine ¹³C tracer experiments to simultaneously detect both ¹⁵N- and ¹³C-labeled metabolites in metabolic samples. A database consisting of 2D ¹H–¹⁵N HSQC natural-abundance spectra of 50 nitrogen-containing metabolites are also presented to facilitate the assignment of ¹⁵N-labeled metabolites. The methodology is demonstrated by labeling Escherichia coli and Staphylococcus aureus metabolomes with ¹⁵N₁-ammonium chloride, ¹⁵N₄-arginine, and ¹³C₂-acetate. Efficient ¹⁵N and ¹³C metabolite labeling and identification were achieved utilizing standard cell culture and sample preparation protocols.

中文翻译：

---

基于氮的NMR扩展了代谢组的覆盖范围

同位素标记代谢物并追踪其代谢命运，除了各种其他问题外，还提供了关于代谢在人类疾病中的作用的宝贵见解。¹³ C标记的代谢物示踪剂或未标记的基于¹ H的NMR实验是NMR在代谢组学研究中最常见的应用。不幸的是，代谢组的覆盖范围因此仅限于最丰富的含碳代谢产物。为了扩大代谢组学的覆盖面并增强代谢组学研究的影响，我们提出了一种用于¹⁵ N标记的代谢物示踪剂实验的方案，该协议还可以与常规的¹³ C示踪剂实验结合使用，以同时检测¹⁵ N和代谢样品中¹³ C标记的代谢物。还提供了一个数据库，该数据库由50种含氮代谢物的2D ¹ H - ¹⁵ N HSQC自然丰度光谱组成，以促进^15种N标记代谢物的分配。该方法通过用¹⁵ N _1-氯化铵，¹⁵ N _4-精氨酸和¹³ C _2-乙酸酯标记大肠杆菌和金黄色葡萄球菌代谢组来证明。高效¹⁵ N和¹³利用标准细胞培养和样品制备方案可实现C代谢物的标记和鉴定。