Partially Purified Leaf Fractions of Azadirachta indica Inhibit Trypanosome Alternative Oxidase and Exert Antitrypanosomal Effects on Trypanosoma congolense

INTRODUCTION: Trypanosomiasis is a neglected disease of humans and livestock caused by single-celled flagellated haemo-protozoan parasites belonging to the genus Trypanosoma. PURPOSE: Widespread resistance to trypanocidal drugs creates urgent need for new, more effective drugs with potential to inhibit important trypanosome molecular targets. METHODS: Nine column chromatographic, partially purified leaf fractions of Azadirachta indica (AIF) were subjected to trypanosome alternative oxidase (TAO) inhibition assay using ubiquinol oxidase assay. The potent TAO inhibitors were evaluated for trypanocidal activities against T. congolense in rat model using in vitro, ex vivo, and in vivo assays. Complete cessation or reduction in parasite motility was scored from 0 (no parasite) to 6 (greater than or equal to 6 × 10(7) trypanosomes/milliliter of blood), and was used to evaluate the efficacy of in vitro treatments. RESULTS: Only AIF1, AIF2, and AIF5 significantly inhibited TAO. AIF1 and AIF5 produced significant, dose-dependent suppression of parasite motility reaching score zero within 1 h with EC(50) of 0.005 and 0.004 µg/µL, respectively, while trypanosome-laden blood was still at score six with an EC(50) of 44,086 µg/µL. Mice inoculated with the concentrations at scores 0 and 1 (1–2 moribund parasites) at the end of the experiment did not develop parasitaemia. The two fractions significantly (p < 0.05) lowered parasite burden, with the AIF5 exhibiting highest in vivo trypanocidal effects. Packed cell volume was significantly higher in AIF1 (p < 0.05) and AIF5 (p < 0.001) groups compared to DMSO-treated group. Only AIF5 significantly (p < 0.05) lowered malondialdehyde. CONCLUSION: AIF1 and AIF5 offer prospects for the discovery of TAO inhibitor(s).