Both enzymes, CPT 1 and CPT 2, play an important role in metabolization of fatty acids. They are different proteins, localized on different genes. While CPT 1 is integrated in the mitochondrial membrane by transmembranal domains, CPT 2 is only in lose association. Disorders of the CPT enzyme system are diseases, able to vary in clinical symptoms. While some patients only develope mild symptoms, for example stress induced muscle pain, also hypocetotic hypoglycaemia, rhabdomyolysis, cardiac myopathy and other symptoms, leading to death, are possible. Previous studies of the CPT enzyme system rarely examined its function in human skeletal muscle and often only some of the possible substrates were used. Aim of this study was the determination of maximal enzymatic activity, affinity and sensitivity to malonyl-CoA of CPT 1 and CPT 2 for fatty acids with chain length of 8 till 18 carbon atoms. First both proteins were seperated from other transferases. In the next step the isoenzymes CPT 1 and 2 were seperated from each other by ultrazentrifugation and detergent treatment. Both, pellet (CPT 1) and superatant (CPT2) fraction, under in vitro conditiones showing highest enzymatic activities for short and medium/long chain fatty acids. Partially, unphysiological high substrate concentrations were used. Highest affinities were determined for long chain fatty acids, but in the supernatant (CPT 2) the Km constant showed a remarkable deviation with substrate lauroyl-CoA (C:12-CoA) from this tendency. Is this, may be, a new transferase activity with specificity for C-12, activated by detergent treatment and ultrazentrifugation? With regard to substrate inhibiton of the enzymes by fatty acids, used in concentrations over substrate optimum, there were clear differences between the fractions on the one, and the substrates on the other hand. Long chain fatty acids are inhibiting transferase activity at lower concentrations in comparison with medium/long chain fatty acids.