Lipidomic Analysis of Human Meibum Using HPLC-MSn
Lipidomic Analysis of Human Meibum Using HPLC-MSn
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DOI：
10.1007/978-1-60761-322-0_11
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发表时间：
2009-01-01
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影响因子：
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通讯作者：
Butovich, Igor A.
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文献类型：
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作者：
Butovich, Igor A.
高压液相色谱-质谱（HPLC-MS）已成为复杂生物样品脂质组学分析中事实上的标准分析工具。该技术在目前可用的分析方法中提供了选择性和灵敏度的最佳组合，并且不仅提供了分析物的保留时间，还提供了它们的m/x值，由此可以推断化合物的分子量。该技术的进一步增强来自这样一个事实，即一些MS仪器（也称为离子阱或MSn仪器）能够对分析物进行多级碎片化，从而使研究人员能够进行结构解析。这些功能使HPLC-MSn成为分析小型复杂脂质样品（如人睑脂）的理想工具。睑板脂是一种非常复杂的脂质混合物，由睑板腺分泌到眼表面。睑脂在人眼表的生物化学和生理学中起关键作用。然而，尽管做出了所有努力，其（生物）化学成分仍然难以捉摸。在本章中，几个HPLC-MSn方法开发的脂质组学分析人类睑脂将进行讨论。考虑到分析物的性质（所有这些都是难溶于水的疏水化合物，其中大部分是电中性的），本研究中使用的唯一MS技术将是大气压化学电离（APCI）MSn的正离子和负离子模式。电喷雾离子化技术虽然在磷脂分析中有用，但被发现不足以分析极性较低的化合物，如蜡酯。由于本章提供的数据将表明，睑脂主要由非极性脂质组成，包括脂肪酯、胆固醇酯和三酰甘油家族，没有明显数量的极性脂质存在，因此APCI MSn似乎是睑脂和类似脂质混合物的脂质组学分析的首选方法。
High-pressure liquid chromatography-mass spectrometry (HPLC-MS) has become a de facto standard analytical tool in lipidomic analyses of complex biological samples. This technique offers that best combination of selectivity and sensitivity among the currently available analytical methods, and provides not only the retention times of analytes, but also their m/x values, from which molecular masses of the compounds can be deduced. Further enhancement of the technique comes from the fact that some of the MS instruments (also known as ion traps, or MSn instruments) are capable of multistage fragmenting of the analytes, thus enabling the researcher to perform their structural elucidation. These capabilities make HPLC-MSn an ideal tool for analyzing small, complex lipid sample such as human meibum. Meibum is a very complex lipid mixture which is secreted onto the ocular surface by Meibomian glands. Meibum plays a critical role in the biochemistry and physiology of the human ocular surface. However, despite all efforts, its (bio)chemical composition remains elusive. In this chapter, several HPLC-MSn methods developed for lipidomic analysis of human meibum will be discussed. Considering the nature of analytes (all of which are hydrophobic compounds poorly soluble in water, and most of which are electroneutral), the only MS technique used in the study will be atmospheric pressure chemical ionization (APCI) MSn in both the positive and the negative ion modes. Electrospray ionization technique, though useful in phopholipid analyses, was found to be inadequate for analyzing less polar compounds, such as wax esers. As the data provided in this chapter will show that meibum is composed predominantly of nonpolar lipids of was ester, cholesteryl ester, and triacylglycerol families with no appreciable amounts of more polar lipids present, APCI MSn seems to be a method of choice for lipidomic analysis of meibum and similar lipid mixtures.