HeLa cells exposed to chloramphenicol for approximately one cell generation were found to contain a mitochondria-associated DNA polymerase with a significantly lower specific activity than that of control cells. This observation was not due to the presence of inhibitors in mitochondrial DNA polymerase preparations of chloramphenicol-treated cell cultures. In addition, there was no accumulation of a typical mitochondrial DNA polymerase in the post-mitochondrial supernatant of drug-treated cells.