This study investigates the "external" NADH-cytochrome c reductase system found in rat-liver mitochondria, which is insensitive to respiratory chain inhibitors and not coupled to phosphorylation. The authors demonstrate that this system resembles the microsomal NADH-cytochrome c reductase system in stereochemical properties but differs from the respiratory chain-linked NADH dehydrogenase. They show that microsomal contamination cannot account for the rotenone-insensitive activity observed in mitochondria. A method is developed to separate two particulate subfractions: one containing the respiratory chain components and the other containing the rotenone-insensitive NADH-cytochrome c reductase system. Morphological evidence supports the conclusion that the former subfraction consists of mitochondria devoid of the outer membrane, while the latter represents derivatives of the outer membrane. The data suggest that the electron-transport system associated with the mitochondrial outer membrane involves catalytic components similar to or identical with the microsomal NADH-cytochrome b6 reductase and cytochrome b6.This study investigates the "external" NADH-cytochrome c reductase system found in rat-liver mitochondria, which is insensitive to respiratory chain inhibitors and not coupled to phosphorylation. The authors demonstrate that this system resembles the microsomal NADH-cytochrome c reductase system in stereochemical properties but differs from the respiratory chain-linked NADH dehydrogenase. They show that microsomal contamination cannot account for the rotenone-insensitive activity observed in mitochondria. A method is developed to separate two particulate subfractions: one containing the respiratory chain components and the other containing the rotenone-insensitive NADH-cytochrome c reductase system. Morphological evidence supports the conclusion that the former subfraction consists of mitochondria devoid of the outer membrane, while the latter represents derivatives of the outer membrane. The data suggest that the electron-transport system associated with the mitochondrial outer membrane involves catalytic components similar to or identical with the microsomal NADH-cytochrome b6 reductase and cytochrome b6.