Synthesis of acetyl,docosahexaenoyl-glycerophosphocholine and its characterization using nuclear magnetic resonance

Docosahexaenoic acid (DHA) circulates in mammals in lipoproteins and bound to serum albumin as a nonesterified fatty acid as well as esterified in lys ophosphatidylcholine (lysoPC). 1-Lyso,2-DHA-glycerophosphocholine (GPC) is an unstable isomer because of a primary alcohol at the sn-l position. To ke ep DHA at the sn-2 position of lysoPC, its usual position for the correspon ding lysoPC to be acylated into PC in tissues, we synthesized 1-acetyl,2-DH A-GPC and confirmed its structure by use of nuclear magnetic resonance (NMR ) spectroscopy in comparison with its positional isomer, 1-DHA,2-acetyl-GPC . l-Lyso,2-DHA-GPC was prepared from 1-stearoyl,2-DHA-GPC by enzymatic hydr olysis and purified by high-performance liquid chromatography. The isomeriz ation of 1-lyso,2-DHA-GPC into l-DHA,2-lyso-GPC was obtained by keeping the former overnight at room temperature under nitrogen. Both lysoPC isomers w ere acetylated by acetic anhydride into 1-acetyl,2-DHA-GPC and l-DHA,2-acet yl-GPC, respectively, and the resulting phospholipids were fully characteri zed by NMR. In particular, the 1,2 substitution pattern of the acetyl and D HA chains could be easily detected by 2D heteronuclear multibond correlatio n. We conclude that 1 -acetyl,2-DHA-GPC might be considered as a stable for m of l-lyso,2-DHA-GPC for its delivery to tissues, if the latter exhibits a cetyl hydrolase activity.