Analysis of products derived from 5,8,11-eicosatrienoic acid via the 5-lipoxygenase-leukotriene pathway showed that this fatty acid is readily converted to leukotriene (LT)A3. When 10,000 X g supernatant from rat basophilic leukemia cell homogenates was incubated with 30 microM fatty acid, 5,8,11-eicosatrienoic acid produced 6.2 +/- 1.1 nmol of LTA3 and arachidonic acid 15.5 +/- 1.9 nmol of LTA4 (n = 4). However, only insignificant amounts of LTB3 were formed (0.15 +/- 0.04 nmol of LTB3 and 4.2 +/- 0.4 nmol of LTB4, n = 4). These data indicate that the LTA-hydrolase requires not only the three double bonds of the triene but also the double bond at C-14 to efficiently convert LTA to LTB. These findings have significant implications for essential fatty acid deficiency.