J. Neurochem. (2012) 120, 842–849. Abstract Endocannabinoids are neuromodulatory lipids that mediate the central and peripheral neural functions. Endocannabinoids have demonstrated their anti‐proliferative, anti‐angiogenic and pro‐apoptotic properties in a series of studies. In the present study, we investigated the levels of two major endocannabinoids, anandamide and 2‐arachidonylglycerol (2‐AG), and their receptors, CB1 and CB2, in human low grade glioma (WHO grade I‐II) tissues, high grade glioma (WHO grade III‐IV) tissues, and non‐tumor brain tissue controls. We also measured the expressions and activities of the enzymes responsible for anandamide and 2‐AG biosynthesis and degradation, that is, N‐acylphosphatidylethanolamine‐hydrolysing phospholipase D (NAPE‐PLD), fatty acid amide hydrolase (FAAH), monoacylglycerol lipase (MGL), and diacylglycerol lipase‐alpha (DGL), in the same samples. Liquid chromatography–mass spectometry analysis showed that the levels of anandamide decreased, whereas the levels of 2‐AG increased in glioma tissues, comparing to the non‐tumor controls. The expression levels and activities of NAPE‐PLD, FAAH and MGL also decreased in glioma tissues. Furthermore, quantitative‐PCR analysis and western‐blot analysis revealed that the expression levels of cananbinoid receptors, CB1 and CB2, were elevated in human glioma tissues. The changes of anandamide and 2‐AG contents in different stages of gliomas may qualify them as the potential endogenous biomarkers for glial tumor malignancy.