MASS DETERMINATION OF THE FATTY-ACIDS RELEASED FROM TANNIN-STIMULATEDRABBIT ALVEOLAR MACROPHAGES

Previous studies with macrophages that had been prelabeled with [C-14] arachidonic acid (20:4) have shown that condensed tannin is a potent a gonist for the release of arachidonic acid. However, it has not been d emonstrated that the percentage release of [C-14]20:4 accurately refle cts the metabolic activity of the endogenous 20:4 pool. In order to me asure the 20:4 mass release relative to the total cellular 20:4 pool, the free fatty acids of freshly isolated alveolar macrophages were der ivatized with a fluorescent reagent, and then separated and quantified by high-performance liquid chromatography. The amounts of esterified fatty acids were measured by gas chromatography of the methyl esters. Free fatty acid levels were compared to those of the total esterified plus unesterified fatty acids to determine the actual percentage relea sed of each fatty acid. Tannin-stimulated release of 20:4 mass reflect ed that previously reported for the release of [C-14]20:4 label but at a slower rate and at a much lower percentage indicating that [C-14]20 :4 had been incorporated into part of a more reactive pool. The specif icity of the fatty acid release induced by tannin and beta-1,3-glucan, a known agonist for 20:4 release, was also examined. Both agonists pr omoted an increase in the levels of free 20:4 and of other fatty acids . A comparison of the absolute increases of each of the fatty acids in dicated that tannin caused a preferential increase in the mass of free 20:4, whereas beta-1,3-glucan evoked a selective increase in the mass of 16:0.