Therapeutic doses of glucocorticoids are thought to inhibit prostaglandin and leukotriene formation in humans. Several studies in animals, however, have failed to demonstrate modulation of eicosanoid biosynthesis by steroids in vivo. We administered prednisone (60 mg/day) to eight healthy volunteers and measured eicosanoid formation by a variety of cell types in vivo and ex vivo, using sensitive and specific physicochemical assays. We found that the in vivo course of prednisone failed to inhibit the synthesis of thromboxane A2, prostaglandin I2 (prostacyclin), prostaglandin E2, and leukotriene E4 in vivo and of leukotriene B4 ex vivo. Biosynthesis of leukotriene B4, thromboxane B2, and prostaglandins F2 and E2 by macrophage-rich bronchoalveolar lavage cells was strongly suppressed. These findings indicate that therapeutic regimens of glucocorticoids suppress eicosanoid biosynthesis in human macrophages but not in a number of other cell types with steroid receptors, the capacity for eicosanoid formation, and lipocortin-like material.