Department of Biochemistry, Graduate School of Medicine, Juntendo University, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421, Japan.
Faculty of Pharma-Science, Teikyo University, Tokyo 173-8605, Japan.
J Biochem. 2021 Oct 12;170(3):389-397. doi: 10.1093/jb/mvab054.
Fatty acids (FAs) play important roles in several physiological and pathophysiological processes, functioning as both nonesterified free FAs (FFAs) and components of other lipid classes. Although many lipid classes are readily measured using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS), the measurement of FFAs by this method is not straightforward because of inconsistent fragmentation behaviours. In this study, we describe a strategy to measure FFAs using conventional reverse-phase LC-ESI-MS/MS, without derivatization. The strategy combines three key methods: (i) an isocratic LC separation with a high organic solvent ratio, (ii) postcolumn base addition, and (iii) pseudo-multiple reaction monitoring. The method facilitates the measurement of ultra-long-chain FAs, the accumulation of which is a common biochemical abnormality in peroxisomal disorders. This study delivers a broad strategy that measures a wide spectrum of FFA species in complex biological samples.
脂肪酸（FAs）在许多生理和病理生理过程中发挥着重要作用，既可以作为非酯化游离脂肪酸（FFAs），也可以作为其他脂质类别的组成部分。尽管许多脂质类别可以使用液相色谱-电喷雾串联质谱（LC-ESI-MS/MS）进行准确测量，但由于碎片行为不一致，该方法测量 FFAs 并不简单。在这项研究中，我们描述了一种无需衍生化即可使用常规反相 LC-ESI-MS/MS 测量 FFAs 的策略。该策略结合了三种关键方法：（i）高有机溶剂比例的等度 LC 分离，（ii）柱后碱添加，和（iii）拟多重反应监测。该方法促进了超长链 FAs 的测量，超长链 FAs 的积累是过氧化物酶体疾病中常见的生化异常。本研究提供了一种广泛的策略，可以测量复杂生物样品中广泛的 FFA 种类。
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