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Updated: Jul 3, 2026
Multi-step Preparation Technique to Recover Multiple Metabolite Compound Classes for In-depth and Informative Metabolomic Analysis
Published on: July 11, 2014
Comprehensive profiling of the human circulating endocannabinoid metabolome: clinical sampling and sample storage
JodiAnne T Wood1, John S Williams, Lakshmipathi Pandarinathan
1Center for Drug Discovery, Northeastern University, Boston, MA 02115, USA. j.wood@neu.edu
Background:
Endogenous cannabinoid-receptor ligands (endocannabinoids) and over a dozen related metabolites now comprise the "endocannabinoid metabolome". The diverse (patho)physiological roles of endocannabinoids, the predictive/diagnostic utility of systemic endocannabinoid levels, and the growing interest in endocannabinoid-related pharmacotherapeutics mandate a valid clinical protocol for processing human blood that does not jeopardize profiling of the circulating endocannabinoid metabolome.
Methods:
We systematically evaluated the potential effect of pre-analytical variables associated with phlebotomy and sample handling/work-up on the human-blood endocannabinoid metabolome as quantified by state-of-the-art liquid chromatography-mass spectrometry.
Results:
Neither subject posture during phlebotomy nor moderate activity beforehand influenced the blood levels of the 15 endocannabinoid-system lipids quantified. Storage of fresh blood at 4 degrees C selectively enhanced ethanolamide concentrations artifactually without affecting monoglycerides and nonesterified fatty acids, such as arachidonic acid. In marked contrast, ethanolamides and monoglycerides remained stable through three plasma freeze/thaw cycles, whereas plasma arachidonic acid content increased, probably a reflection of ongoing metabolism.
Conclusions:
Class- and compound-selective pre-analytical influences on circulating human endocannabinoid levels necessitate immediate plasma preparation from fresh blood and prompt plasma apportioning and snap-freezing. Repeated plasma thawing and refreezing should be avoided. This protocol ensures sample integrity for evaluating the circulating endocannabinoid metabolome in the clinical setting.
