Chemical stability of 2-arachidonylglycerol under biological conditions.
Recent evidence indicates that 2-arachidonylglycerol (2-AG) is a potent and specific ligand for the central and peripheral cannabinoid receptors. Therefore, the chemical stability of this molecule under biological conditions is of interest. A method for the isolation and detection of 2-AG using HPLC with evaporative light scattering detection is described. The method provides an extraction recovery from aqueous media of 78%, and a limit of detection of 60 ng on column. Incubation of 2-AG in culture medium or biological buffers indicated that it is stable to oxidation and ester hydrolysis for up to 6 h at 37 degrees C. However, gradual disappearance of the compound was noted due to adherence to glass and plastic surfaces. During incubation in RPMI culture medium, 2-AG rearranged to 1(3)-arachidonylglycerol (1(3)-AG) in a first order process with a half-life of 10 min in the absence of serum and 2.3 min in the presence of 10% fetal calf serum. Further studies indicated that the acyl migration reaction is base catalyzed (k(cat)=78,000/min M), and that the reaction is affected slightly by changes in buffer (Tris) concentration and not at all by changes in ionic strength. The results indicate that 2-AG is readily converted to 1(3)-AG under conditions commonly used to study receptor-ligand interactions, findings that have significant implications for the interpretation of relative ligand potency between the two isomers.
Rouzer
Carol A
CA
Department of Biochemistry, Vanderbilt-Ingram Cancer Center, Vanderbilt University School of Medicine, Nashville, TN 37232-0146, USA. rouzer@toxicology.mc.vanderbilt.edu
Ghebreselasie
Kebreab
K
Marnett
Lawrence J
LJ
D001095:Arachidonic Acids; D002021:Buffers; D002851:Chromatography, High Pressure Liquid; D003470:Culture Media; D004355:Drug Stability; D063388:Endocannabinoids; D005989:Glycerides / Q000737:chemistry*; D008024:Ligands; D010084:Oxidation-Reduction; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug / Q000819:agonists
D001095:Arachidonic Acids; D002021:Buffers; D003470:Culture Media; D063388:Endocannabinoids; D005989:Glycerides; D008024:Ligands; D043882:Receptors, Cannabinoid; D011955:Receptors, Drug; C094503:glyceryl 2-arachidonate