Extraction and analysis of sphingolipids from biological samples is a critical step in lipidomics, especially for minor species such as sphingoid bases and sphingosine-1-phosphate. Although several liquid chromatography-mass spectrometry methods enabling the determination of sphingolipid molecular species have been reported, they were limited in analytical sensitivity and reproducibility by causing significant peak tailing, especially by the presence of phosphate groups, and most of the extraction techniques are laborious and do not cover a broad range of sphingolipid metabolites. In this study, we developed a rapid single-phase extraction and highly sensitive analytical method for the detection and quantification of sphingolipids (including phosphates) comprehensively using liquid chromatography-triple quadruple mass spectrometry. After validating the reliability of the method, we analyzed the intestinal tissue sphingolipids of germ-free (GF) and specific pathogen-free (SPF) mice and found significantly higher levels of free sphingoid bases and sphingosine-1-phosphate in the GF condition as compared to the SPF condition. This method enables a rapid extraction and highly sensitive determination of sphingolipids comprehensively at low femtomolar ranges.

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从生物样品中提取和分析鞘脂是脂质组学中的关键步骤，特别是对于小物种，如鞘氨醇碱基和鞘氨醇-1-磷酸酯而言。尽管已经报道了几种能够确定鞘脂分子种类的液相色谱-质谱联用方法，但是由于引起显着的峰拖尾，特别是由于存在磷酸根基团，它们的分析灵敏度和重现性受到限制，并且大多数萃取技术费力且费时。不包括广泛的鞘脂代谢产物。在这项研究中，我们开发了一种快速单相萃取和高灵敏度分析方法，用于使用液相色谱-三重四极杆质谱仪全面检测和定量鞘脂（包括磷酸盐）。在验证了方法的可靠性之后，我们分析了无菌（GF）和无特定病原体（SPF）小鼠的肠道组织鞘脂，发现在GF条件下，游离鞘氨醇碱和鞘氨醇-1-磷酸水平显着较高，因为与SPF条件相比。该方法能够在低飞摩尔范围内快速提取并高灵敏度地全面测定鞘脂。

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