Gas chromatography-tandem mass spectrometry with electron ionization (GC-EI-MS/MS) provides rich information on stable-isotope labeling for 13C-metabolic flux analysis (13C-MFA). To pave the way for the routine application of tandem MS data for metabolic flux quantification, we aimed to compile a comprehensive library of GC-EI-MS/MS fragments of tert-butyldimethylsilyl (TBDMS) derivatized proteinogenic amino acids. First, we established an analytical workflow which combines high resolution gas chromatography-quadrupole time-of-flight mass spectrometry (GC-EI-QTOFMS) and fully 13C-labeled biomass to identify and structurally elucidate tandem MS amino acid fragments. Application of the high-mass accuracy MS procedure resulted into the identification of 129 validated precursor-product ion pairs of 13 amino acids with 30 fragments being accepted for 13C-MFA. The practical benefit of the novel tandem MS data was demonstrated by a proof-of-concept study which confirmed the importance of the compiled library for high resolution 13C-MFA.

中文翻译：

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改进大肠杆菌代谢中的 13C 代谢通量分析：应用高分辨率 MS (GC-EI-QTOF) 全面评估 MS/MS 片段

电子电离气相色谱-串联质谱 (GC-EI-MS/MS) 为 13C 代谢通量分析 (13C-MFA) 提供了有关稳定同位素标记的丰富信息。为了为代谢通量定量的串联 MS 数据的常规应用铺平道路，我们的目标是编译叔丁基二甲基甲硅烷基 (TBDMS) 衍生的蛋白氨基酸的 GC-EI-MS/MS 片段的综合文库。首先，我们建立了一个分析工作流程，将高分辨率气相色谱-四极杆飞行时间质谱 (GC-EI-QTOFMS) 和完全 13C 标记的生物质相结合，以识别和结构阐明串联 MS 氨基酸片段。应用高质量准确度 MS 程序鉴定了 129 个经过验证的 13 个氨基酸的前体-产物离子对，其中 30 个片段被 13C-MFA 接受。概念验证研究证明了新型串联 MS 数据的实际好处，该研究证实了编译的文库对于高分辨率 13C-MFA 的重要性。