A number of publications recently provided measurements of plasma ECs by liquid chromatographytandem mass spectrometry (LCMS/MS), however the reported circulating levels in normal subjects are scattered in a very wide range of concentrations. A quantitative LCMS/MS method for anandamide (AEA), palmitoylethanolamide (PEA), oleoylethanolamyde (OEA) and 2arachidonoyl-glycerol (2AG), chromatographically resolved by its inactive isomer 1AG, was validated in house. We firstly evaluated the stability of ECs during the early stages of the blood processing. Five blood aliquots were withdrawn in EDTA-containing tubes from 20 subjects, and blood was kept at 4°C until centrifugation after 0, 30, 60, 120 and 240 min, followed by storage of plasma at −80°C. Probably due to a blood cells spill-over, we found AEA, PEA and OEA dramatically increase to about 150% of the t0 concentration in the first 30 min, reaching 250% after 240 min, while 2AG level slowly decreases to 80% of t0 concentration after 240 min, maybe due to isomerisation. Once equipped for the proper storage of plasma within 1520 min from the withdrawal, samples from 215 healthy and completely drug free volunteers were taken. Normal values (pmol/ml) in 112 normal weight subjects were: AEA: 1.16±0.31; PEA: 12.25±2.71; OEA: 4.22±1.44; 2AG: 0.88±0.34; no gender differences were observed. In females increased AEA and PEA levels were shown both in 34 overweight (22 and 11%, respectively) and 13 obese subjects (21 and 7%, respectively); in male obese group (n=13) AEA, PEA and 2AG levels increased of 36, 31 and 42%, respectively, but no differences were observed in overweight group (n=43). No changes were observed in OEA levels. A gender specific association between circulating ECs and overweight/obesity was revealed: AEA and PEA in females, AEA, PEA and, to a greater extent, 2AG, in male obesity.
30 Apr - 04 May 2011
European Society of Endocrinology