Comprehensive Quantitation Using Two Stable Isotopically Labeled Species and Direct Detection of N-Acyl Moiety of Sphingomyelin.
Comprehensive Quantitation Using Two Stable Isotopically Labeled Species and Direct Detection of N-Acyl Moiety of Sphingomyelin.
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DOI：
10.1007/s11745-017-4279-5
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发表时间：
2017-09
期刊：
影响因子：
1.9
通讯作者：
Yokoyama K
中科院分区：
文献类型：
--
作者：
Hama K;Fujiwara Y;Tabata H;Takahashi H;Yokoyama K
鞘磷脂（神经酰胺-磷酸胆碱，CerPCho）是哺乳动物细胞中常见的鞘脂，由磷酸胆碱和神经酰胺分别作为极性和疏水组分组成。在这项研究中，定性液相色谱-电喷雾电离串联质谱（LC-ESI-MS/MS/MS）分析，提出了CerPCho结构分配的基础上对应于鞘氨醇磷酸胆碱和N-酰基部分的产品离子光谱。从CerPCho的MS/MS/MS分析中，我们观察到N-酰基脂肪酸作为[RCO 2]−离子以及鞘氨醇磷酸胆碱的产物离子光谱。使用两种稳定的同位素标记的CerPCho物质构建CerPCho的校准曲线，然后用于定量HeLa细胞中的CerPCho物质，作为原理验证研究。本研究提出了一种准确的方法，通过LC-ESI-MS/MS/MS分析定量和分配结构的每一个CerPCho物种在粗生物样品。
Sphingomyelin (ceramide-phosphocholine, CerPCho) is a common sphingolipid in mammalian cells and is composed of phosphorylcholine and ceramide as polar and hydrophobic components, respectively. In this study, a qualitative liquid chromatography-electrospray ionization tandem mass spectrometry (LC–ESI–MS/MS/MS) analysis is proposed in which CerPCho structures were assigned based on product ion spectra corresponding to sphingosylphosphorylcholine and N-acyl moieties. From MS/MS/MS analysis of CerPCho, we observed product ion spectra of the N-acyl fatty acids as [RCO2]− ions as well as sphingosylphosphorylcholine. A calibration curve for CerPCho was constructed using two stable isotopically labeled CerPCho species and then used to quantify the CerPCho species in HeLa cells as a proof-of-principle study. The present study proposes an accurate method for quantifying and assigning structures to each CerPCho species in crude biologic samples by LC–ESI–MS/MS/MS analysis.
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