In-tube solid-phase microextraction directly coupled to tandem mass spectrometry for anandamide and 2-arachidonoylglycerol determination in rat brain samples from an animal model of Parkinson's disease

To evaluate the endocannabinoid system in an animal model of Parkinson's disease, in-tube solid-phase microextraction (in-tube SPME) was directly coupled to a tandem mass spectrometry (MS/MS) system for determination of the endocannabinoids anandamide (AEA) and 2-arachidonoylglycerol (2-AG) in rat brain samples. In-tube SPME-which consisted of a microtube of restricted access material (RAM) with a hydrophilic diol external surface and a hydrophobic octyl inner surface-efficiently excluded (up to 95%) macromolecules from the biological samples and selectively pre-concentrated the analytes. In-tube SPME parameters, such as sample volume, mobile phases, flow rate, and pre-concentration time, were evaluated to improve the extraction efficiency and throughput performance. The selectivity of the in-tube SPME and MS/MS (MRM mode) techniques allowed them to be directly coupled online, which dismissed the need for the chromatographic separation step. The in-tube SPME-MS/MS method was validated and shown to be linear from 6.0 to 30.0 ng mL(-1) for AEA and from 10.0 to 100.0 ng mL(-1) for 2-AG; the intra-and inter-assay accuracy and precision were lower than 15%. Parallelism between the calibration curves constructed in the matrix and aqueous solution confirmed that there was no matrix effect. The method allowed endogenous concentrations of AEA and 2-AG to be determined in rat brain striatum from unilaterally 6-hydroxydopamine-lesioned animals. The concentrations of these endocannabinoids in striatum ipsilateral and contralateral to the lesion differed significantly (p<0.001). (C) 2020 Elsevier B.V. All rights reserved.

Automated summary

The study successfully evaluated the endocannabinoid system in an animal model of Parkinson's disease using in-tube solid-phase microextraction coupled to tandem mass spectrometry. By optimizing the in-tube SPME parameters and selective methods, the concentrations of AEA and 2-AG in rat brain samples were determined, providing data support for further research.