Minced tissues from both infarcted and apparently normal areas of canine myocardium 6 h after ligation of the left descending branch of the coronary artery were incubated with [1,2-(14)C]ethanolamine. In each case substantial amounts of radioactivity were incorporated into both phosphatidylethanolamine and N-acylphosphatidylethanolamine. Incubation of homogenates from the same tissues with N-[1-(14)C]palmitoyl phosphatidylethanolamine yielded labeled N-palmitoyl ethanolamine. The data support the concept that N-acylethanolamines, pharmacologically active compounds which accumulate in infarcted myocardium, are produced by N-acylation of ethanolamine phospholipids followed by phosphodiesterase action.