Two types of endogenous cannabinoid-receptor agonists have been identified thus far. They are the ethanolamides of polyunsaturated fatty acids-arachidonoyl ethanolamide (anandamide) is the best known compound in the amide series-and 2-arachidonoyl glycerol, the only known endocannabinoid in the ester series. We report now an example of a third, ether-type endocannabinoid, 2-arachidonyl glyceryl ether (noladin ether), isolated from porcine brain. The structure of noladin ether was determined by mass spectrometry and nuclear magnetic resonance spectroscopy and was confirmed by comparison with a synthetic sample. It binds to the CB1 cannabinoid receptor (Ki ‫؍‬ 21.2 ؎ 0.5 nM) and causes sedation, hypothermia, intestinal immobility, and mild antinociception in mice. It binds weakly to the CB2 receptor (Ki > 3 M).W e have reported the isolation and identification of two types of endogenous cannabinoids that bind and activate the known cannabinoid receptors CB 1 and CB 2 . Arachidonoyl ethanolamide (anandamide; ref. 1) and later two more polyunsaturated fatty acid ethanol amides (2) were found in porcine brain. An ester, 2-arachidonoyl glycerol (2-AG) was isolated by us from canine gut (3) and by Sugiura et al. from brain (4). Anandamide and 2-AG have been the objects of numerous investigations in various areas of biology and have been found to affect processes in the nervous, cardiovascular, immune, and reproductive systems (5-8). They interact with many neurotransmitters and affect hormone levels (9-10). This ubiquity of effects led us to look for additional endocannabinoids.We report now that we have isolated from porcine brain a third endocannabinoid, 2-arachidonyl glyceryl ether, which we have named noladin ether ( Fig. 1). Materials and MethodsIsolation of Noladin Ether. Porcine brain (100 g, approximately a single brain) was added to a mixture of chloroform (200 ml) and methanol (200 ml) and mixed in a blender for 2 min. Water (100 ml) was added, and the mixing process was continued for another minute. The mixture was filtered. Two layers were formed. The water-methanol layer was separated and evaporated under reduced pressure. The residue obtained was extracted with methanol. The above isolation was repeated numerous times (from a total of 2.4 kg porcine brain). The extract obtained was chromatographed on a gravity column (i.d. 2.5 cm, height 28 cm, 82 g ICN Silica TSC, 60 A) with hexane͞acetone initially in a ratio of 10:1 (vol͞vol) (400 ml), then 9:1 (100 ml), and finally 4:1 (100 ml). The fractions eluted were monitored for binding to the CB 1 receptor from rat brain synaptosomes (prepared as described below; ref. 11) on the basis of displacement of the potent labeled agonist [ 3 H]HU-243 purchased from Tocris (Bristol, U.K.). The material eluted in fractions 45-54 (10-ml each) was found to bind to the receptor. These fractions were combined and purified further by HPLC (see below). A polar-active compound developed on a TLC plate (silica gel 60 F 254 , Merck) in a hexane͞ acetone (4:1) solvent system gave ...

The endocannabinoid N-arachidonoyl ethanolamine (anandamide), found both in the CNS and in the periphery, plays a role in numerous physiological systems. One might expect that the chemically related N-arachidonoyl-L-serine (ARA-S) could also be formed alongside anandamide. We have now isolated ARA-S from bovine brain and elucidated its structure by comparison with synthetic ARA-S. Contrary to anandamide, ARA-S binds very weakly to cannabinoid CB 1 and CB2 or vanilloid TRPV1 (transient receptor potential vanilloid 1) receptors. However, it produces endothelium-dependent vasodilation of rat isolated mesenteric arteries and abdominal aorta and stimulates phosphorylation of p44͞42 mitogen-activated protein (MAP) kinase and protein kinase B͞Akt in cultured endothelial cells. ARA-S also suppresses LPS-induced formation of TNF-␣ in a murine macrophage cell line and in wild-type mice, as well as in mice deficient in CB 1 or CB2 receptors. Many of these effects parallel those reported for abnormal cannabidiol (Abn-CBD), a synthetic agonist of a putative novel cannabinoidtype receptor. Hence, ARA-S may represent an endogenous agonist for this receptor.abnormal cannabidiol ͉ anandamide ͉ cannabinoids ͉ endothelium ͉ reactive oxygen intermediates T he identification, structural elucidation, and syntheses of the plant cannabinoids in the early 1960s led to thorough investigations of the chemistry, metabolism, and pharmacology of these compounds, in particular of the psychoactive constituent ⌬ 9 -tetrahydrocannabinol (1, 2). However, until the late 1980s and early 1990s, when specific receptors were identified and shortly thereafter cloned, the mechanism of the numerous cannabinoid actions remained an enigma (3-5). Two main receptors are now known: the CB 1 receptor, found in the CNS, as well as in some peripheral tissues, and the CB 2 receptor, found predominantly in the immune system (6-8). Additional, not yet fully identified receptors are present both in the CNS and in the periphery (6, 9, 10).Because receptors in mammals are not formed to encounter a plant constituent, research was initiated to discover endogenous ligands. In the 1990s two endogenous cannabinoids (endocannabinoids) were identified, N-arachidonoyl ethanolamine (anandamide) (11) and 2-arachidonoyl-glycerol (12, 13). Additional endocannabinoids have been reported, but their biological roles are yet obscure (6, 14). Anandamide and 2-arachidonoyl-glycerol have large spectrum of physiological actions, most of which are associated with the neural and immune systems. However, cardiovascular effects, which are in part CB 1 -mediated (15), are also well established (9,14,16).Anandamide is a product of phosphatidylethanolamine (17). Because phosphatidylserine is found alongside phosphatidylethanolamine in body tissues, one might expect that arachidonoyl-Lserine (ARA-S) is also an endogenous constituent (see Fig. 1A for the structures of anandamide and ARA-S). We report that we have isolated ARA-S from bovine brain and have evaluated some of its biological propertie...

The proposed anti-inflammatory model can be utilized for the virtual screening of large chemical databases and for indexing natural products for potential anti-inflammatory activity.

Background: Ephedra is among Palestinian medicinal plants that are traditionally used in folkloric medicine for treating many diseases. Ephedra is known to have antibacterial and antioxidant effects. The goal of this study is to evaluate the antioxidant activity of different extracts from the Ephedra alata plant growing wild in Palestine, and to analyze their phenolic and flavonoid constituents by HPLC/PDA and HPLC/MS. Materials and Methods: Samples of the Ephedra alata plant grown wild in Palestine were extracted with three different solvents namely, 100% water, 80% ethanol, and 100% ethanol. The extracts were analyzed for their total phenolic content (TPC), total flavonoid content (TFC), antioxidant activity (AA), as well as phenolic and flavonoids content by HPLC/PDA/MS. Results:The results revealed that the polarity of the extraction solvent affects the TPC, TFC, and AA of extracts. It was found that both TPC and AA are highest for plant extracted with 80% ethanol, followed by 100% ethanol, and finally with 100% water. TFC however was highest in the following order: 100% ethanol > 80% ethanol > water. Pearson correlation indicated that there is a significant correlation between AA and TPC, but there is no correlation between AA and TFC. Simultaneous HPLC-PDA and UHPLC-MS analysis of the ethanolic plant extracts revealed the presence of Luteolin-7-O-glucuronide flavone, Myricetin 3-rhamnoside and some other major polyphenolic compounds that share myricetin skeleton. Conclusion Ephedra alata extract is rich in potent falvonoid glycosidic compounds as revealed by their similar overlaid UV-Vis spectra and UHPLC-MS results. On the basis of these findings, it is concluded that Ephedra alata constitutes a natural source of potent antioxidants that may prevent many diseases and could be potentially used in food, cosmetics, and pharmaceutical products.