The effect of digitonin on the release from rat-liver mitochondria of adenylate kinase (EC 2.7.4.3) and malate dehydrogenase (EC 1.1.1.37) and on the unmasking of activities of cytochrome oxidase (EC 1.9.3.1) and succinate-cytochrome c reductase with external cytochrome c was studied. Electron micrographs of digiton-in-treated mitochondria were also taken. The results indicate that the unmasking of mitochondrial enzyme activities with exogenous cytochrome c goes parallely with the release of adenylate kinase and the detachment of the outer mitochondrial membrane but does not require the opening of the inner membrane. © 1969.