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. 2008 Jul 1;105(26):9099-104.
doi: 10.1073/pnas.0803601105. Epub 2008 Jun 23.

Beta-caryophyllene Is a Dietary Cannabinoid

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Free PMC article

Beta-caryophyllene Is a Dietary Cannabinoid

Jürg Gertsch et al. Proc Natl Acad Sci U S A. .
Free PMC article

Abstract

The psychoactive cannabinoids from Cannabis sativa L. and the arachidonic acid-derived endocannabinoids are nonselective natural ligands for cannabinoid receptor type 1 (CB(1)) and CB(2) receptors. Although the CB(1) receptor is responsible for the psychomodulatory effects, activation of the CB(2) receptor is a potential therapeutic strategy for the treatment of inflammation, pain, atherosclerosis, and osteoporosis. Here, we report that the widespread plant volatile (E)-beta-caryophyllene [(E)-BCP] selectively binds to the CB(2) receptor (K(i) = 155 +/- 4 nM) and that it is a functional CB(2) agonist. Intriguingly, (E)-BCP is a common constituent of the essential oils of numerous spice and food plants and a major component in Cannabis. Molecular docking simulations have identified a putative binding site of (E)-BCP in the CB(2) receptor, showing ligand pi-pi stacking interactions with residues F117 and W258. Upon binding to the CB(2) receptor, (E)-BCP inhibits adenylate cylcase, leads to intracellular calcium transients and weakly activates the mitogen-activated kinases Erk1/2 and p38 in primary human monocytes. (E)-BCP (500 nM) inhibits lipopolysaccharide (LPS)-induced proinflammatory cytokine expression in peripheral blood and attenuates LPS-stimulated Erk1/2 and JNK1/2 phosphorylation in monocytes. Furthermore, peroral (E)-BCP at 5 mg/kg strongly reduces the carrageenan-induced inflammatory response in wild-type mice but not in mice lacking CB(2) receptors, providing evidence that this natural product exerts cannabimimetic effects in vivo. These results identify (E)-BCP as a functional nonpsychoactive CB(2) receptor ligand in foodstuff and as a macrocyclic antiinflammatory cannabinoid in Cannabis.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.
Caryophyllane- and humulane-type sesquiterpenes found in C. sativa and numerous other plants. Shown are the chemical structures of the bicyclic sesquiterpenes (E)-β-caryophyllene, (Z)-β-caryophyllene, caryophyllene oxide, and the ring-opened isomer α-humulene (α-caryophyllene).
Fig. 2.
Fig. 2.
(E)-BCP and (Z)-BCP displace [3H]CP-55,940 from hCB2 receptors expressed in HEK293 cells. (A) Sigmoidal displacement curves (R2 = 0.93 for (E)-BCP and 0.98 for (Z)-BCP) show overall displacement (71% and 84%, respectively) with Ki values of 155 and 485 nM, respectively. Data show mean values of nine measurements ± SEM. (B) Hill plot showing linearized data and the corresponding Ki values. (C) Dixon plot of the competitive binding interaction of (E)-BCP with the CP-55,940 receptor binding site in the CB2 receptor. Radioligand assays were performed by using 84 pM, 168 pM, and 252 pM [3H]CP-55,940.
Fig. 3.
Fig. 3.
Model of the putative interaction of (E)-BCP with the CB2 receptor determined by Surflex–Dock and MD/MM calculations. (E)-BCP is located in the hydrophobic region of the amphipathic CB2 receptor binding pocket where it closely interacts with hydrophobic residues F117, I198, W258, V113, and M265. In this model, significant π–π stacking interactions between the (E)-BCP double bonds and F117 and W258, respectively, facilitate binding.
Fig. 4.
Fig. 4.
G protein triggered effects of (E)-BCP upon CB2 receptor binding. (A) The high-affinity CB ligand WIN55,212–2 and (E)-BCP dose-dependently inhibit forskolin-stimulated cAMP production in CB2 receptor-transfected CHO-K1 cells. Data are mean values of three independent experiments measured in triplicates ± SEM in a nonlinear dose-response curve (R2 > 0.97). (B) Like 2-AG, which was used as positive control, (E)-BCP dose-dependently triggers [Ca2+]i transients in CB2 expressing HL60 cells. The FACS histogram shows CB2 immunofluorescence of HL60 cells. Data are mean values of three independent experiments ± SEM shown in a nonlinear dose-response curve (R2 = 0.99). (C) No [Ca2+]i transients were induced in HL60 cells devoid of CB2 surface expression. The FACS histogram shows the lack of CB2 Ab immunofluorescence in these HL60 cells. (D) Addition of the CB2-selective antagonist SR144528 (1 μM) inhibited the [Ca2+]i release triggered by (E)-BCP (10 μM) in CB2-positive HL60 cells. Data are mean values from three independent experiments ± SEM (paired t test **, P < 0.01).
Fig. 5.
Fig. 5.
Anti-inflammatory effects of (E)-BCP in vitro and in vivo. (A) (E)-BCP (1 h incubation before stimulation) inhibits LPS-stimulated (313 ng/ml) IL-1β and TNF-α protein expression in human peripheral whole blood (determined after 18 h). Prior incubation with the CB2 receptor antagonist AM630 (5 μM for 1 h) blocked the effect of (E)-BCP. Data show values from three independent experiments ± SEM (paired t test *, P < 0.05; **, P < 0.01; ***, P < 0.001). (B) Intraplantar carrageenan (30 μl)-induced edema formation is inhibited by orally administered (E)-BCP (5 and 10 mg/kg) in C57BL/6J wild-type (Cnr2+/+) mice but not in CB2 (Cnr2−/−) knockout mice. Shown is the mean increase in paw volume over time of at least nine mice per group ± SEM (ANOVA, *, P < 0.05; **, P < 0.01; ***, P < 0.001). (C) Comparison of the effects of oral JWH133 (CB2 selective agonist, Ki = 3.4 nM) and (E)-BCP on carrageenan (30 μl)-induced edema formation in C57BL/6J wild-type (Cnr2+/+) mice after 240 min. Shown is the percentage of increase in paw volume (relative to control) of at least nine mice per group ± SEM (paired t test, *, P < 0.05; **, P < 0.01; ***, P < 0.001).

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