Free palmitate is activated to palmitoyl-CoA in the cytosol before it can be oxidized in the mitochondria. If palmitate and [14 C]CoA are added to a liver homogenate, palmitoyl CoA isolated from cytosolic fraction is radioactive, but that isolated from the mitochondrial fraction is not. Explain.

Activation of fatty acids (palmitate) occurs in the cytoplasm where fatty acids are activated to fatty acyl CoA, reaction catalysed by an enzyme called fatty acyl CoA synthetase
A specialized carnitine carrier system catalyze transportation of activated fatty acid from cytoplasm to matrix of mitochondria, where carnitine system consists of three proteins:
Carnitine acyl transferase I located in outer membrane of mitochondria catalyze transfer of carnitine to fatty acyl CoA and produce fatty acyl carnitine
Carnitine translocase facilitate passive diffusion of fatty acyl carnitine from inter membrane space to matrix located in inner membrane
Carnitine acyl transferase III located in inner mitochondrial membrane at matrix phase catalyze transfer of CoA to fatty acyl carnitine and regenerate fatty acyl CoA

Hence, the cytosolic and mitochondrial pools of CoA are thus kept separate, and no radioactive CoA from the cytosolic pool enters the mitochondria