The effects of salicylic acid on palmitic acid oxidation were studied using rat skeletal muscle mitochondria. Salicylic acid, in concentrations that exerted no effect on mitochondrial coupling (0.1 mM), significantly stimulated mitochondrial palmitic acid oxidation, with maximal stimulation occurring at subsaturating concentrations of substrate. In the same preparation, salicylate had no effect on the oxidation of palmitoylcarnitine or palmitoyl-CoA. Salicylate appears to augment the initial step of palmitic acid oxidation by lowering the apparent Michaelis constant (Km) of long chain fatty acid: CoASH ligase (AMP) (EC 6.2.1.3) for palmitic acid.