Oxidation of fatty acids and tricarboxylic acid cycle intermediates by isolated rat liver mitochondria.

Beginning with the fundamental observation of Warburg in 1913 (1) it has been a general finding that the more highly organized enzyme systems of animal tissues responsible for oxidation of metabolites by molecular oxygen are associated with the insoluble particulate portion of the cell. Among the several approaches which have been used to study the morphology and composition of such catalytically active particulate material, the most fruitful has been the differential centrifugation technique for separation of nuclei, mitochondria or “large granules,” and other substructures developed by Bensley and his school (2) and refined by Claude (3,4) and Hogeboom, Schneider, and Pallade (5, 6). Considerable work on the composition and enzymatic activity of the various particulate fractions has been described by the Rockefeller group (7, 8), Schneider (9), and other investigators. For instance, quantitative assays have revealed that most of the succinoxidase and cytochrome oxidase activity is present in the mitochondria or “large granules” (7). In this laboratory studies have been made on the enzymatic oxidation of fatty acids to acetoacetate and also via the Krebs tricarboxylic acid cycle. These complex and highly organized reactions take place in suspensions of particulate material separated from rat liver homogenates by centrifugation (10, 11). Certain observations on the properties of this enzyme system (11) suggested that the activity was to some extent dependent on osmotic factors, and Potter, on the basis of measurements of “cytolysis quotients,” suggested that the activity was present only in intact cells (12). With the publication of what appears to be a definitive method for the isolation of mitochondria or “large granules” by Hogeboom, Schneider, and Pallade (5,6), it was possible to demonstrate that mitochondria isolated by this method bear all the demonstrable fatty acid oxidase activity of whole rat liver. The particulate material isolated by this method is stated to be homogeneous and identical in morphology and vital staining characteristics