Adenosine diphosphate (ADP) released into blood induces platelet aggregation and contributes to hemostasis and thrombosis. Released ATP can also induce platelet aggregation and there is evidence that blood leukocytes and also erythrocytes play important roles in this. Rapid metabolism of ADP and ATP by endothelial cells is important in protecting platelets from their effects. Here we have performed a systematic investigation of adenine nucleotide metabolism in human blood and the involvement of blood cells. Conversion of ATP to ADP in blood was due almost exclusively to the presence of leukocytes; plasma, platelets and erythrocytes made little or no contribution. Mononuclear leukocytes (MNLs) and polymorphonuclear leukocytes (PMNLs) were equally effective. Conversion of ADP to AMP was also promoted by leukocytes, with no involvement of platelets or erythrocytes. Some ADP was also converted to ATP in blood, apparently via an enzyme present in plasma, but ATP was then rapidly removed by the leukocytes. Conversion of AMP to adenosine occurred via a plasma enzyme with little or no contribution from any cellular element. As expected, in blood the adenosine produced was removed very rapidly by erythrocytes and then converted to inosine and then hypoxanthine. In the absence of erythrocytes plasma supported only a slow conversion of adenosine to inosine and hypoxanthine, which was not influenced by platelets or leukocytes. This study has demonstrated that leukocytes and erythrocytes play a major role in adenine nucleotide metabolism in blood and that these cells, as well as endothelial cells, may be important determinants of the effects of ATP and ADP on platelets.