Ceramide profiling of complex lipid mixtures by electrospray ionization mass spectrometry
Ceramide profiling of complex lipid mixtures by electrospray ionization mass spectrometry
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DOI：
10.1006/abio.1996.9915
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发表时间：
1997-01-15
影响因子：
2.9
通讯作者：
Aebersold, R
中科院分区：
文献类型：
--
作者：
Gu, M;Kerwin, JL;Aebersold, R
神经酰胺和鞘氨醇碱是重要的细胞内第二信使，在细胞生长、分化和程序性细胞死亡的调节中发挥作用。到目前为止，神经酰胺第二信使的定量和定性分析由于缺乏能够检测各个神经酰胺种类的内源水平和区分各个神经酰胺种类的分析方法而受到限制。在这里，我们报告使用电喷雾电离串联质谱法对神经酰胺进行定性和定量分析。碰撞诱导的碎裂导致鞘氨醇和二氢鞘氨醇 (sphinganine) 头部基团的特征产物离子分别为 m/z 264 和 282 以及 m/z 266 和 284，无论脂肪酰链的长度如何，光谱在低至 25 nM (25 fmol/μl) 的浓度下可重现。这些报告离子用于通过前体离子扫描分析检测复杂混合物中的鞘氨醇和二氢鞘氨醇基神经酰胺。我们展示了该方法在牛脑神经酰胺商业制剂中的神经酰胺组成分析的应用，以及在最小限度的色谱分离后，培养的 T 细胞的脂质提取物。此外，通过比较三种相关淋巴细胞系 Jurkat、U937 和 WEHI 231 的概况，我们可以轻松检测到各个神经酰胺种类水平的相对差异。最后，通过添加非天然存在的内标，我们表明该技术可用于测量此类生物来源的脂质制剂中神经酰胺水平的定量变化。 (C) 1997 学术出版社
Ceramides and sphingoid bases are important intracellular second messengers that play a role in the regulation of cell growth, differentiation, and programmed cell death. Until now, quantitative and qualitative analysis of ceramide second messengers has been limited by a lack of analytical methods capable of detecting endogenous levels of, and differentiating between, individual ceramide species. Here we report the use of electrospray ionization tandem mass spectrometry for the qualitative and quantitative analysis of ceramides. Collision-induced fragmentation resulted in characteristic product ions for the sphingosine and dihydrosphingosine (sphinganine) head groups at m/z 264 and 282 and m/z 266 and 284, respectively, regardless of the length of the fatty acyl chains, with spectra being reproducible at concentrations as low as 25 nM (25 fmol/mu l). These reporter ions were used to detect both sphingosine- and sphinganine-based ceramides in complex mixtures using precursor ion scan analysis. We demonstrated the application of this method for profiling the composition of ceramides in a commercial preparation of bovine brain ceramides and, following minimal chromatographic separation, a lipid extract of cultured T cells. Furthermore, we easily detected relative differences in individual ceramide species levels by comparing the profiles of three related lymphocyte cell lines, Jurkat, U937, and WEHI 231. Finally, by the addition of a nonnaturally occurring internal standard, we show that the technique can be used to measure quantitative changes in ceramide levels in such biologically derived lipid preparations. (C) 1997 Academic Press