The effect of oxygenated sterols on platelet aggregation induced by thrombin and ADP has been studied. All oxygenated sterols tested with a hydroxyl group on the side chain enhanced thrombin-induced aggregation at 25 microM. In the case of ADP-induced aggregation, however, only 22S-hydroxycholesterol[(22S)-5-cholestene-3 beta,22-diol] enhanced the aggregation, and 22R-hydroxycholesterol[(22R)-5-cholestene-3 beta,22-diol], 24S-hydroxycholesterol[(24S)-5-cholestene-3 beta,24-diol], and 25-hydroxycholesterol[5-cholestene-3 beta,25-diol] inhibited it. These effects were observed within 10 min after the addition of oxygenated sterols to platelet suspensions in plasma.