Surrogate analyte-based quantification of main endocannabinoids in whole blood using liquid chromatography-tandem...

Endocannabinoids (eCBs) are endogenous ligands of the endocannabinoid system that are known to regulate several physiological and behavioral processes. Previous studies have developed methods for the detection of main eCBs including arachidonylethanolamide (AEA) and 2-arachidonoylglycerol (2-AG), mostly in serum or plasma. Whole blood is a superior biomaterial for eCBs analysis owing to the nature of the shortened isolation procedure and decreased risk of 2-AG isomerization during preparation. In this study, a surrogate analyte-based liquid chromatography-tandem mass spectrometry assay was developed for the measurement of AEA, 2-AG and its isomer 1-arachidonoylglycerol (1-AG) using a maximum of 100 μL whole blood. Chromatographic separation was achieved using a reverse-phase column and a gradient elution. Detection was performed in selected reaction monitoring mode with an electrospray ionization source. The limits of detection of three eCBs were 0.05-0.1 ng/mL. Good linearity was observed over the concentration range. Intra- and inter-assay accuracy and precision were ≤10.9 and ≤8.7% at four quality control levels. The response factor and parallelism experiment illustrated that the surrogate analytes were suitable for accurate quantification of the main eCBs in whole blood. This surrogate analyte approach was successfully applied to authentic blood samples obtained from alcohol negative drivers and those under the influence of alcohol.