Analytical Chemistry, Department of Chemistry--Biomedical Center and Science for Life Laboratory, Uppsala University, Box 599, 751 24 Uppsala, Sweden.
Anal Bioanal Chem. 2013 Feb;405(6):1929-35. doi: 10.1007/s00216-012-6658-3. Epub 2013 Jan 11.
Circulating free fatty acids (FFAs) play important physiological roles as contributing components in cellular structure as well as energy utilization. Elevated levels of circulating FFAs are associated with metabolic aberrations in humans. FFAs differ in chain length and saturation and may be altered in metabolically dysregulated conditions, such as type 2 diabetes mellitus. Potentially, alterations in circulating levels of specific FFAs could also be important in terms of prognostic value. Various methods have been used to analyze FFAs. In this study, a straightforward and accurate method for the determination of FFAs in plasma has been established and evaluated, through conversion of plasma FFAs into acid fluorides followed by conversion to Weinreb amides (dimethylamide). The method is mild, efficient, selective, and quantitative for FFAs, when analyzed with capillary gas chromatography tandem mass spectrometry. Standard curves were linear over the range of 1,000-20,000 ng/mL with a correlation coefficient (r(2)) of 0.998, and coefficient of variation of triplicate analysis was <10 %. The gas chromatography-mass spectrometry (GC-MS) technique was reproducible and repeatable, and recoveries were above 90 %. From the generated MS spectra, five specific FFAs were identified. An explicit interest was the quantification of palmitate (C16:0) and palmitoleate (C16:1), which have been connected with detrimental and positive effects on the insulin-producing beta cells, respectively. The results demonstrate the suitability of Weinreb amides for efficient and rapid isolation of FFAs in plasma, prior to quantitative GC-MS analysis. We suggest that the method can be used as a routine standardized way of quantifying FFAs.
循环游离脂肪酸 (FFAs) 作为细胞结构的组成成分和能量利用的贡献者，发挥着重要的生理作用。循环 FFAs 水平升高与人类代谢异常有关。FFAs 在链长和饱和度上存在差异，在代谢失调的情况下（如 2 型糖尿病）可能会发生改变。潜在地，特定 FFAs 循环水平的改变也可能具有重要的预后价值。已经使用了各种方法来分析 FFAs。在这项研究中，建立并评估了一种直接、准确的血浆 FFAs 测定方法，即将血浆 FFAs 转化为酸氟化物，然后转化为 Weinreb 酰胺（二甲基酰胺）。该方法温和、高效、选择性强，定量分析毛细管气相色谱串联质谱法分析 FFAs 时，标准曲线在 1000-20000ng/ml 范围内呈线性，相关系数（r²）为 0.998，重复分析的变异系数 <10%。气相色谱-质谱 (GC-MS) 技术重现性和可重复性好，回收率高于 90%。从生成的 MS 谱中鉴定出了五种特定的 FFAs。特别关注的是棕榈酸 (C16:0) 和棕榈油酸 (C16:1) 的定量，它们分别与胰岛素分泌β细胞的有害和有益作用有关。结果表明，Weinreb 酰胺适合于在定量 GC-MS 分析之前，高效快速地从血浆中分离 FFAs。我们建议该方法可作为定量 FFAs 的常规标准化方法。
