Department of Pharmaceutics, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 13 Hangkong Road, Wuhan 430030, China.
Department of Pharmacology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, 13 Hangkong Road, Wuhan 430030, China.
J Chromatogr B Analyt Technol Biomed Life Sci. 2020 Apr 1;1142:122041. doi: 10.1016/j.jchromb.2020.122041. Epub 2020 Feb 24.
Obesity has become a severe public health problem worldwide. An endogenous fatty acid ethanolamine oleoyl ethanolamine (OEA) is reported to be capable of reducing body weight and food intake by increasing striatal extracellular dopamine concentration. However, association between obesity and striatal OEA level remains unknown. As such, it is necessary to develop a sensitive and reliable method to quantitate OEA concentration in striatum. Because true endogenous analytes free blank matrix is not available, surrogate analyte, surrogate matrix and background subtraction methods are often employed for the analysis of endogenous compounds. In this study, three liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods were developed and validated for the determination of OEA concentration in mouse brain homogenate. Interestingly, stability results found that OEA-d4 degraded in brain homogenate under room temperature, while OEA level remarkably increased with time. Since lowering temperature could observably decelerate the endogenous transformation of OEA, sample collection and preparation were carried out under ice-bath condition. Hexane: isopropanol (9:1, v/v) was employed as an extractant for liquid-liquid extraction. After method validation, three methods were applied to quantify OEA in striatum homogenate from C57B6/L mice following normal and high fat diet feeding for 4 months. Results from three methods all showed the striatal OEA level in obesity group was significantly higher than control group and obesity-resist group, which indicated that obesity might be associated with elevated striatal OEA level.
肥胖已成为全球严重的公共卫生问题。据报道，内源性脂肪酸乙醇胺油酰乙醇胺 (OEA) 能够通过增加纹状体细胞外多巴胺浓度来降低体重和食物摄入量。然而，肥胖与纹状体 OEA 水平之间的关联尚不清楚。因此，有必要开发一种灵敏、可靠的方法来定量测定纹状体中的 OEA 浓度。由于真正的内源性分析物没有空白基质，因此通常采用替代分析物、替代基质和背景扣除方法来分析内源性化合物。在这项研究中，开发并验证了三种液相色谱-串联质谱 (LC-MS/MS) 方法来测定小鼠脑匀浆中的 OEA 浓度。有趣的是，稳定性结果发现 OEA-d4 在室温下脑匀浆中降解，而 OEA 水平随时间显著增加。由于降低温度可以明显减缓 OEA 的内源性转化，因此在冰浴条件下进行样品采集和制备。正己烷：异丙醇 (9:1，v/v) 用作液液萃取的提取剂。方法验证后，三种方法均用于定量测定正常和高脂肪饮食喂养 4 个月后的 C57B6/L 小鼠纹状体匀浆中的 OEA。三种方法的结果均表明肥胖组纹状体 OEA 水平明显高于对照组和肥胖抵抗组，这表明肥胖可能与纹状体 OEA 水平升高有关。
