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A simple LC-MS/MS method for the simultaneous quantification of endocannabinoids in biological samples
Journal of Chromatography B ( IF 3 ) Pub Date : 2020-09-06 , DOI: 10.1016/j.jchromb.2020.122371
Maria Bobrich , Rico Schwarz , Robert Ramer , Philipp Borchert , Burkhard Hinz

The arachidonic acid derivatives N-arachidonoylethanolamine (anandamide; AEA), 2-arachidonoylglycerol (2-AG), N-arachidonoyldopamine (NADA), 2-arachidonoylglycerol ether (noladin ether; 2-AGE) and O-arachidonoylethanolamine (virodhamine; VA) were identified as physiological components of the endocannabinoid (EC) system. In order to gain further profound knowledge about the different EC-induced physiological and pathophysiological effects, appropriate analytical methods are required. The method described here uses liquid chromatography in combination with positive electrospray ionization mass spectrometry (LC-MS/MS) to quantify the concentrations of the above-mentioned EC compounds in cells. Sample preparation prior to LC-MS/MS analysis was performed by means of two liquid extractions with ethyl acetate. The method has been validated according to the bioanalytical guidelines of the Food and Drug Administration (FDA). The lower limits of quantification were 0.03 ng/mL for AEA, 2 ng/mL for 2-AG, 0.03 ng/mL for NADA, 0.3 ng/mL for 2-AGE and 0.15 ng/mL for VA. Linearity was demonstrated up to 10 ng/mL (AEA, NADA, 2-AGE and VA) and 50 ng/mL (2-AG). The values for intra- and inter-day precision and accuracy were within the guideline recommended acceptance criteria for assay validation. Low matrix effects and good recovery were found for AEA, 2-AG and 2-AGE, while a higher matrix effect was observed for NADA and VA. Extraction yields were lowest for VA. The method was used for EC measurement in different cell lines and in mouse brains.



中文翻译:

一种同时定量生物样品中内源性大麻素的简单LC-MS / MS方法

花生四烯酸衍生物N-花生四烯酸乙醇胺(anandamide; AEA),2-花生四烯酸甘油酯(2-AG),N-花生四烯酸多巴胺(NADA),2-花生四烯酸甘油醚(noladin ether; 2-AGE)和O花生四烯酸乙醇胺(virodhamine; VA)被鉴定为内源性大麻素(EC)系统的生理成分。为了获得有关EC诱导的不同生理和病理生理效应的更深入的知识,需要适当的分析方法。此处描述的方法将液相色谱法与正电喷雾电离质谱法(LC-MS / MS)结合使用,以量化细胞中上述EC化合物的浓度。LC-MS / MS分析之前的样品制备是通过两次用乙酸乙酯进行的液体萃取进行的。该方法已根据食品药品管理局(FDA)的生物分析指南进行了验证。定量下限为AEA为0.03 ng / mL,2-AG为2 ng / mL,NADA为0.03 ng / mL,2-AGE为0.3 ng / mL和0。VA为15 ng / mL。线性高达10 ng / mL(AEA,NADA,2-AGE和VA)和50 ng / mL(2-AG)。日内和日间精度和准确性的值均在用于检验验证的准则建议的接受标准之内。AEA,2-AG和2-AGE的基质效应低,回收率高,而NADA和VA的基质效应更高。VA的提取率最低。该方法用于不同细胞系和小鼠大脑中的EC测量。而NADA和VA的基质效应更高。VA的提取率最低。该方法用于不同细胞系和小鼠大脑中的EC测量。而NADA和VA的基质效应更高。VA的提取率最低。该方法用于不同细胞系和小鼠大脑中的EC测量。

更新日期:2020-09-06
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