The discovery of the endogenous cannabinoid N-arachidonoylethanolamine (anandamide)1 and other N-acylethanolamines (NAEs) in chocolate2 has led to speculation that the purported rewarding properties of cocoa are due to the presence of compounds “that could act as cannabinoid mimics”2. This observation raises some important questions. First, are NAEs and anandamide, or the ‘endocannabinoid’ 2-arachidonoylglycerol (2-AG)3, present in widely consumed foods (such as milk) that are less ‘rewarding’ than chocolate? And second, to what extent do these compounds reach the bloodstream and exert pharmacological effects when consumed orally? We believe that the content of endocannabinoids in foods, and in cocoa in particular, is not sufficient to produce cannabis-like effects in mammals.
We purified NAEs and 2-AG from various foods (including mature human, bovine and goat milk, and cocoa at various stages of processing) and quantified them by gas chromatography and mass spectrometry. Milk was the only food analysed in which an endocannabinoid, 2-AG, was found in relatively high concentrations (0.33±0.11 μg ml−1 in human milk, for example). The level of NAEs varied between 0.003 and 0.024 μg ml−1, with anandamide being the least abundant compound. Oleamide, a sleep-inducing substance4 that inhibits the hydrolysis of anandamide5,6, was also present (0.055-1.27 μg ml−1).
In cocoa-derived samples, we detected NAEs (0.01-5.8 μg per g) and oleamide (0.17-6.0 μg per g), but no or very little anandamide and no 2-AG. NAE levels are much lower in unfermented cocoa beans than in cocoa powder (which contained less than 0.003 μg per g anandamide). Tiny amounts of anandamide in cocoa could therefore be explained as artefacts of processing2. Like all higher plants, cocoa plants cannot synthesize arachidonic acid or its derivatives7. Notably, in their NAE and oleamide content, cocoa and dark chocolate are similar to other plant-derived foods (including soybean, hazelnuts, oatmeal and millet), in which we detected up to 2.3, 1.1 and 2.8 μg per g oleamide, N-oleoyl-ethanolamine and N-linoleoyl-ethanol-amine, respectively.
To establish whether endocannabinoids survive their passage through the digestive system, we assayed anandamide and 2-AG after oral administration in a series of in vivo tests that are used widely to assess cannabimimetic activity8 (Table 1). The two compounds were active in four of the five behavioural tests, but only at very high concentrations relative to those in foods. In contrast, a smaller dose of the psychoactive component of marijuana, (−)-Δ9-tetra-hydrocannabinol (Δ9-THC), was sufficient to give a strong effect in all tests.
Because intraperitoneal anandamide and 2-AG are active in the same tests at doses 20- to 60-fold lower3,9, these results indicate that only 1.6-5% of the orally administered compounds enter the bloodstream, probably owing to the high levels in the gastrointestinal tract of the enzyme fatty acid amide hydrolase10, which catalyses the hydrolysis of both compounds. Therefore, it is unlikely that the amounts of anandamide and 2-AG found in food are sufficient to produce observable psychotropic effects. We did not test other unsaturated NAEs that have been suggested to enhance endogenous anandamide activity by inhibiting its inactivation2, but instead assayed oleamide — for which this kind of action is also supported by pharmacological data5,6. High oral doses are again necessary for activity to be observed in vivo (Table 1).
Our results show that the amounts of anandamide, 2-AG and oleamide in foods, including milk and cocoa, are several orders of magnitude below those required, if administered by mouth, to reach the blood and cause observable ‘central’ effects. The assays used here provide a gross evaluation of cannabimimetic activity, and tests monitoring more subtle behavioural changes that might be induced by low oral doses of NAEs/oleamide are needed before the relevance of these compounds to the purported mild rewarding and craving-inducing effects of cocoa can be dismissed. The presence in milk of 2-AG (which may be released from triglycerides during digestion) and oleamide also needs to be investigated.
Reply - Beltramo and Piomelli
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Di Marzo, V., Sepe, N., De Petrocellis, L. et al. Trick or treat from food endocannabinoids?. Nature 396, 636 (1998). https://doi.org/10.1038/25267
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