Sickle hemoglobin is relatively unstable upon oxidation or mechanical shaking. During denaturation, it generates oxygen radicals and hemichromes and ultimately precipitates in the form of micro-Heinz bodies. It is not clear, however, whether the degradation product hemin, which is a potent hemolytic agent and a potential perturbant to protein-protein interactions in the red cell membrane skeleton, is also generated during sickle hemoglobin denaturation. By specific absorption of hemin with Dowex AG 1-X8 anion-exchange resin at high-ionic strength conditions, we now separate hemin for quantitation from the bulk hemoglobin and its derivatives. We demonstrate that upon mechanical shaking oxyhemoglobin S denatures much faster than oxyhemoglobin A and that a considerably higher level of hemin is detected in the shaken hemoglobin S as compared with hemoglobin A. By using the same method to measure the hemin content in the hemolysate of fresh red cells from patients with sickle cell disease, we detect a three- to fivefold increase in the hemin content in these patients (0.4 to 0.75 mumol/L) as compared with normal individuals (0.1 to 0.15 mumol/L). These data suggest that the instability of sickle oxyhemoglobin leads to increased intracellular precipitation of hemoglobin and the release of hemin, which may play a role in the membrane lesion of sickle red cells.