A phagocytic vesicle fraction with high NADPH-dependent O-2-forming activity was obtained in large quantity from pig blood polymorphonuclear leukocytes, phagocytosing oil droplets in the presence of cyanide. The activity of NADPH-dependent 2, 6-dichlorophenolindophenol (DCIP) or ferricyanide reduction was also observed in the vesicles. Increasing the concentration of DCIP, the O-2-forming activity decreased and the DCIP-reducing activity enhanced, whereas NADPH oxidation was constant, indicating that electrons from NADPH flow to DCIP on the way to oxygen in the O-2-forming system. The O-2 formation was completely inhibited by 25μM p-chloromercuriben-zoate, whereas the DCIP reduction was only slightly affected. This suggests that the O-2-forming activity may consist of, at least, two components. The O-2-forming activity and the DCIP-reducing activity were effectively extracted with a mixture containing deoxycholate and Tween 20 and the specific activiities increased about 10 times by the extraction. Both activities in extract were enhanced 2-hold by the addition of 1μM FAD, suggesting that the flavin is a component of the enzyme system. A mechanism by which the O-2-forming system is stimulated was also investigated and results which indicate possible involvement of calmodulin as well as protein-phosphorylating and -dephosphorylating reactions in the mechanism were obtained.