Development and validation of a liquid chromatography-tandem mass spectrometry method for the quantitation of synthetic cannabinoids of the aminoalkylindole type and methanandamide in serum and its application to forensic samples

After the discovery of synthetic cannabimimetic substances in 'Spice'-like herbal mixtures marketed as 'incense' or 'plant fertilizer' the active compounds have been declared as controlled substances in several European countries. As expected, a monitoring of new herbal mixtures which continue to appear on the market revealed that shortly after control measures have been taken by legal authorities, other compounds were added to existing mixtures and to new products. Several compounds of the aminoalkylindole type have been detected so far in herbal mixtures but still their consumption cannot be detected by commonly used drug-screening procedures, encouraging drug users to substitute cannabis with those products. There is a increasing demand on the part of police authorities, hospitals and psychiatrists for detection and quantification of synthetic cannabinoids in biological samples originating from psychiatric inpatients, emergency units or assessment of fitness to drive. Therefore, a liquid chromatography-tandem mass spectrometry method after liquid-liquid extraction for the quantitation of JWH-015, JWH-018, JWH-073, JWH-081, JWH 200, JWH-250, WIN 55,212-2 and methanandamide and the detection of JWH-019 and JWH-020 in human serum has been developed and fully validated according to guidelines for forensic toxicological analyses. The method was successfully applied to 101 serum samples from 80 subjects provided by hospitals, detoxification and therapy centers, forensic psychiatric centers and police authorities. Fifty-seven samples or 56.4% were found positive for at least one aminoalkylindole. JWH-019, JWH-020, JWH-200, WIN 55,212-2 and methanandamide were not detected in any of the analyzed samples.

β-Caryophyllene ameliorates cisplatin-induced nephrotoxicity in a cannabinoid 2 receptor-dependent manner

(E)-β-caryophyllene (BCP) is a natural sesquiterpene found in many essential oils of spice (best known for contributing to the spiciness of black pepper) and food plants with recognized anti-inflammatory properties. Recently it was shown that BCP is a natural agonist of endogenous cannabinoid 2 (CB(2)) receptors, which are expressed in immune cells and mediate anti-inflammatory effects. In this study we aimed to test the effects of BCP in a clinically relevant murine model of nephropathy (induced by the widely used antineoplastic drug cisplatin) in which the tubular injury is largely dependent on inflammation and oxidative/nitrative stress. β-caryophyllene dose-dependently ameliorated cisplatin-induced kidney dysfunction, morphological damage, and renal inflammatory response (chemokines MCP-1 and MIP-2, cytokines TNF-α and IL-1β, adhesion molecule ICAM-1, and neutrophil and macrophage infiltration). It also markedly mitigated oxidative/nitrative stress (NOX-2 and NOX-4 expression, 4-HNE and 3-NT content) and cell death. The protective effects of BCP against biochemical and histological markers of nephropathy were absent in CB(2) knockout mice. Thus, BCP may be an excellent therapeutic agent to prevent cisplatin-induced nephrotoxicity through a CB(2) receptor-dependent pathway. Given the excellent safety profile of BCP in humans it has tremendous therapeutic potential in a multitude of diseases associated with inflammation and oxidative stress.

Beta-caryophyllene is a dietary cannabinoid

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.

Synthetic versus natural curcumin: bioequivalence in an in vitro oral mucositis model

BACKGROUND Curcumin (CUR) is a dietary spice and food colorant (E100). Its potent anti-inflammatory activity by inhibiting the activation of Nuclear Factor-kappaB is well established. METHODS The aim of this study was to compare natural purified CUR (nCUR) with synthetically manufactured CUR (sCUR) with respect to their capacity to inhibit detrimental effects in an in vitro model of oral mucositis. The hypothesis was to demonstrate bioequivalence of nCUR and sCUR. RESULTS The purity of sCUR was HPLC-confirmed. Adherence and invasion assays for bacteria to human pharyngeal epithelial cells demonstrated equivalence of nCUR and sCUR. Standard assays also demonstrated an identical inhibitory effect on pro-inflammatory cytokine/chemokine secretion (e.g., interleukin-8, interleukin-6) by Detroit pharyngeal cells exposed to bacterial stimuli. There was bioequivalence of sCUR and nCUR with respect to their antibacterial effects against various pharyngeal species. CONCLUSION nCUR and sCUR are equipotent in in vitro assays mimicking aspects of oral mucositis. The advantages of sCUR include that it is odorless and tasteless, more easily soluble in DMSO, and that it is a single, highly purified molecule, lacking the batch-to-batch variation of CUR content in nCUR. sCUR is a promising agent for the development of an oral anti-mucositis agent.