Development, validation and comparison of three LC-MS/MS methods for determination of endogenous striatal oleoyl ethanolamine in mice
Development, validation and comparison of three LC-MS/MS methods for determination of endogenous striatal oleoyl ethanolamine in mice
复制标题
三种测定小鼠纹状体油酰乙醇胺的 LC-MS/MS 方法的开发、验证和比较
DOI：
10.1016/j.jchromb.2020.122041
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发表时间：
2020-04-01
影响因子：
3
通讯作者：
Huang, Jiangeng
中科院分区：
文献类型：
--
作者：
Guan, Yeli;Deng, Qiao;Huang, Jiangeng
肥胖已成为世界范围内严重的公共卫生问题。据报道，一种内源性脂肪酸乙醇胺油基乙醇胺（OEA）能够通过增加纹状体细胞外多巴胺浓度来减少体重和食物摄入量。然而，肥胖与纹状体OEA水平之间的关系尚不清楚。因此，有必要建立一种灵敏可靠的纹状体OEA浓度定量方法。由于没有真正的内源分析物自由空白基质，因此内源化合物的分析通常采用替代分析物、替代基质和背景减法。本研究建立了三种液相色谱-串联质谱（LC-MS/MS）测定小鼠脑匀浆中OEA浓度的方法，并进行了验证。有趣的是，稳定性结果发现，室温下OEA-d4在脑匀浆中降解，而OEA水平随着时间的推移而显著升高。由于降低温度可以明显减缓OEA的内源转化，因此样品的采集和制备在冰浴条件下进行。采用正己烷：异丙醇（9:1,v/v）作为萃取剂进行液-液萃取。方法验证后，采用三种方法定量C57B6/L小鼠正常和高脂饲料喂养4个月后纹状体匀浆OEA。三种方法的结果均显示肥胖组纹状体OEA水平显著高于对照组和肥胖抵抗组，提示肥胖可能与纹状体OEA水平升高有关。
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.