Mitochondria of Plasmodium falciparum (K1 strain) were isolated by differential centrifugation. Mitochondrial DNA topoisomerase II from P. falciparum was partially purified using fast protein liquid chromatography(FPLC). Parasite mitochondria contained approximately 8% of DNA topoisomerase II activity compared with its nuclear fraction. The effects of fluoroquinolones, inhibitors of bacterial DNA topoisomerase II or DNA gyrase, against partially purified P. falciparum mitochondrial DNA topoisomerase II were investigated using a decatenation assay. Minimum inhibitory concentrations (MIC) of ofloxacin, ciprofloxacin and norfloxacin were > 1, 10 and 100 mM, compared with that of >0.5 and 10 mM for eukaryotic DNA topoisomerase II inhibitor etoposide (VP-16) and amsacrine, respectively. The results indicate that partially purified mitochondrial DNA topoisomerase II was insensitive to fluoroquinolones and it is suggested that their inhibitory effects on P. falciparum growth may be directed against plastid DNA topoisomerase II.