Department of Ophthalmology and Visual Sciences, Moran Eye Center, University of Utah School of Medicine, Salt Lake City, UT 84132, USA.
J Chromatogr A. 2013 Sep 13;1307:191-200. doi: 10.1016/j.chroma.2013.07.103. Epub 2013 Aug 1.
Fatty acids (FAs), including long-chain and very long-chain polyunsaturated fatty acids (LC-PUFAs, C12-22; VLC-PUFAs, C24-38), play an important role in retinal function and health. Deficiencies in LC-PUFAs and VLC-PUFAs, as well as mutations in the enzyme responsible for elongation of very long-chain fatty acids (ELOVL4), have been associated with macular dystrophies and degenerations. Published analytical methods, including high-performance liquid chromatography coupled with mass spectrometry (HPLC-MS) and gas chromatography-MS (GC-MS), can quantify VLC-PUFAs but require at least an entire human retina which limits the ability to understand physiologically relevant variations in lipids that can occur at a regional level within the retina. Until now, quantification of VLC-PUFAs in just the human macula, the cone-rich region of the central retina responsible for high acuity vision, has not been feasible due to its small size (4-5mm in diameter). In this study, we have developed a sensitive GC-MS method using newer generation enhanced GC-MS detector sensitivity which for the first time quantifies not only 14 VLC-PUFAs and 26 LC-FAs but also n-3/n-6 ratios of PUFAs in 4mm punches of human retina or a single pair of mouse retinas. Our results showed that saturated LC-FAs are higher in the human peripheral retina than in the macula, while unsaturated LC-FAs are higher in the macula than in the peripheral retina. On the other hand, the VLC-PUFAs are higher in the peripheral retina compared to macula. There is no difference in n-3/n-6 ratios of PUFAs observed between human macula and peripheral retina, while mouse retina has almost ten times more VLC-PUFAs than human macula and peripheral retina (2.27% versus 0.25% and 0.32%, respectively) and much higher n-3/n-6 ratios compared to human retina (9:1 versus ∼0.9:1). This high sensitivity analytical technique provides a valuable new tool for studies on the role of FAs in the pathological processes of macular degenerations and dystrophies.
脂肪酸（FAs），包括长链和超长链多不饱和脂肪酸（LC-PUFAs，C12-22；VLC-PUFAs，C24-38），在视网膜功能和健康中发挥着重要作用。LC-PUFAs 和 VLC-PUFAs 的缺乏以及负责延长超长链脂肪酸的酶（ELOVL4）的突变，与黄斑营养不良和变性有关。已发表的分析方法，包括高效液相色谱-质谱联用（HPLC-MS）和气相色谱-质谱联用（GC-MS），可以定量 VLC-PUFAs，但至少需要整个人类视网膜，这限制了在视网膜的区域水平上理解与生理相关的脂质变化的能力。到目前为止，由于人类黄斑（中央视网膜中富含视锥细胞的区域，负责高敏锐度视觉）的尺寸较小（直径为 4-5mm），因此无法对其进行 VLC-PUFAs 的定量分析。在这项研究中，我们开发了一种使用新一代增强型 GC-MS 检测器灵敏度的敏感 GC-MS 方法，该方法首次不仅可以定量分析 14 种 VLC-PUFAs 和 26 种 LC-FAs，还可以定量分析人类视网膜 4mm 打孔或一对老鼠视网膜中的 n-3/n-6 比值。我们的结果表明，饱和 LC-FAs 在人类周边视网膜中的含量高于黄斑，而不饱和 LC-FAs 在黄斑中的含量高于周边视网膜。另一方面，VLC-PUFAs 在周边视网膜中的含量高于黄斑。在人类黄斑和周边视网膜之间没有观察到 n-3/n-6 比值的 PUFA 差异，而老鼠视网膜中的 VLC-PUFAs 是人类黄斑和周边视网膜的近十倍（分别为 2.27%、0.25%和 0.32%），且 n-3/n-6 比值远高于人类视网膜（9:1 比约 0.9:1）。这种高灵敏度的分析技术为研究 FAs 在黄斑变性和营养不良的病理过程中的作用提供了一种有价值的新工具。
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