RESUMO
The conversion of cyclohexanecarboxylic acid to hippuric acid in subcellular fractions from guinea pig liver was studied using a gas chromatographic-mass spectrometric method employing selected ion monitoring. Comparison of the activities of the cyclohexanecarboxylic acid to hippuric acid converting system (CHC-system) and marker enzymes in the various subcellular fractions showed that the CHC-system is localized exclusively in the mitochondria. No contribution to the total activity of the system was made by microsomal enzymes. The activity of the CHC-system in whole liver homogenate and in isolated mitochondria was similar when the latter were supplemented with ATP, alpha-ketoglutaric acid, glycine, ethylenediaminetetraacetate, PO4(3-) and Mg2+. The formation of hippuric acid in these mitochondrial preparations was linear with respect to time over a period of at least 60 min. Studies designed to optimize the incubation conditions showed that the activity of the CHC-system was reduced by PO4(3-) concentrations greater than approximately 70 mM. Conversely, both ATP and alpha-ketoglutaric acid stimulated the system. It is possible that two different types of acyl-CoA synthetases, one which is ATP-specific and one which is GTP-specific, may operate in the activation of cyclohexanecarboxylic acid.