The release of polymorphonuclear leukocyte proteinases in buffy-coat-depleted red cell concentrates was examined during a storage period of 35 days. Collagenase, gelatinase and elastase predominantly induce breakdown of connective tissue. However, when released by cell disintegration during red cell concentrate storage, the considerable proteolytic activities of these enzymes might influence the quality of the stored blood. During the observation period a considerable decrease in the polymorphonuclear leukocyte count was observed, accompanied by increases in the levels of collagenase, gelatinase and elastase. Compared with the enzyme levels on the day of red cell concentrate preparation, collagenase increased 20-fold, gelatinase 6-fold and elastase 100-fold during the storage period. When cells were treated with the chemoattractant hexapeptide, N-formyl-nle-leu-phe-nle-tyr-leu, and the degranulation promoting cytochalasin B, gelatinase exhibited the highest secreted concentration in the freshly prepared red cell concentrate, exceeding the maximum of spontaneously released elastase by 4- to 6-fold. However, these compounds stimulated enzyme release only during the first day after red cell concentrate preparation. Thereafter, no differences between stimulated and non-stimulated samples were observed. The data indicate that polymorphonuclear leukocytes contain a large storage pool of proteolytic enzymes. These enzymes together with other polymorphonuclear leukocyte enzymes, e.g. hydrolases and oxidoreductases, might alter the erythrocyte membrane surface and thus influence the storage quality of the prepared red cell concentrate.