Leibniz Institute of Plant Genetics and Crop Plant Research, 06466 Gatersleben, Germany
Leibniz Institute of Plant Genetics and Crop Plant Research, 06466 Gatersleben, Germany.
Plant Physiol. 2017 Oct;175(2):600-618. doi: 10.1104/pp.17.00470. Epub 2017 Aug 11.
Lipid oxidation is a process ubiquitous in life, but the direct and comprehensive analysis of oxidized lipids has been limited by available analytical methods. We applied high-resolution liquid chromatography-mass spectrometry (LC-MS) and tandem mass spectrometry (MS/MS) to quantify oxidized lipids (glycerides, fatty acids, phospholipids, lysophospholipids, and galactolipids) and implemented a platform-independent high-throughput-amenable analysis pipeline for the high-confidence annotation and acyl composition analysis of oxidized lipids. Lipid contents of 90 different naturally aged wheat () seed stocks were quantified in an untargeted high-resolution LC-MS experiment, resulting in 18,556 quantitative mass-to-charge ratio features. In a posthoc liquid chromatography-tandem mass spectrometry experiment, high-resolution MS/MS spectra (5 mD accuracy) were recorded for 8,957 out of 12,080 putatively monoisotopic features of the LC-MS data set. A total of 353 nonoxidized and 559 oxidized lipids with up to four additional oxygen atoms were annotated based on the accurate mass recordings (1.5 ppm tolerance) of the LC-MS data set and filtering procedures. MS/MS spectra available for 828 of these annotations were analyzed by translating experimentally known fragmentation rules of lipids into the fragmentation of oxidized lipids. This led to the identification of 259 nonoxidized and 365 oxidized lipids by both accurate mass and MS/MS spectra and to the determination of acyl compositions for 221 nonoxidized and 295 oxidized lipids. Analysis of 15-year aged wheat seeds revealed increased lipid oxidation and hydrolysis in seeds stored in ambient versus cold conditions.
脂质氧化是生命中普遍存在的过程，但由于可用的分析方法的限制，直接和全面地分析氧化脂质一直具有挑战性。我们应用高分辨率液相色谱-质谱（LC-MS）和串联质谱（MS/MS）来定量氧化脂质（甘油酯、脂肪酸、磷脂、溶血磷脂和半乳糖脂），并实现了一个独立于平台的高通量分析管道，用于氧化脂质的高置信度注释和酰基组成分析。在一个非靶向高分辨率 LC-MS 实验中，对 90 种不同自然老化的小麦（）种子进行了脂质含量的定量分析，共得到 18556 个定量质荷比特征。在随后的液相色谱-串联质谱实验中，对 LC-MS 数据集的 12080 个假定单同位素特征中的 8957 个记录了高分辨率 MS/MS 谱（5 mD 精度）。基于 LC-MS 数据集的精确质量记录（1.5 ppm 公差）和过滤程序，共注释了 353 种非氧化和 559 种氧化脂质，这些脂质最多带有四个额外的氧原子。对这些注释中的 828 种进行了 MS/MS 谱分析，通过将脂质的实验已知的碎裂规则转化为氧化脂质的碎裂来进行分析。这导致通过精确质量和 MS/MS 谱鉴定了 259 种非氧化和 365 种氧化脂质，并确定了 221 种非氧化和 295 种氧化脂质的酰基组成。对 15 年老化的小麦种子的分析表明，在常温而非冷藏条件下储存的种子中，脂质氧化和水解增加。
