Development and Validation of a Sensitive UPLC-ESI-MS/MS
Method for the Simultaneous Quantification of 15 Endocannabinoids
and Related Compounds in Milk and Other Biofluids
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Abstract
The
endocannabinoid (eCB) system has gained an increasing interest
over the past decades since the discovery of anandamide and 2-arachidonoyl
glycerol (2-AG). These, and structurally related compounds, are associated
with a wide variety of physiological processes. For instance, eCB
levels in milk have been associated with infants’ feeding and
sleeping behavior. A method based on ultraperformance liquid chromatography
coupled with electrospray ionization tandem mass spectrometry (UPLC–ESI-MS/MS)
was developed and validated for the simultaneous quantification of
15 eCBs and related compounds, including both fatty acid amides and
glycerols. Linearity (0.9845 < <i>R</i><sup>2</sup> <
1), limit of detection and quantification (0.52–293 pg on column),
inter- and intraday accuracy (>70%) and precision (CV < 15%),
stability,
and recovery (in milk and plasma) were established in accordance to
the U.S. Food and Drug Administration guidelines. The method was successfully
applied to bovine and elk milk revealing species-specific eCB profiles,
with significant different levels of 2-AG, 2-linoleoyl glycerol, docosahexaenoyl
ethanolamide, palmitoyl ethanolamide, and oleoyl ethanolamide. Furthermore,
stearoyl ethanolamide and docosatetraenoyl ethanolamide were only
detected in elk milk. In summary, our UPLC–ESI-MS/MS method
may be used for quantification of eCBs and related compounds in different
biofluids and applied to investigations of the role of these emerging
compounds in various physiological processes