Coumarins isolated from Calophyllum brasiliense produce ultrastructural alterations and affect in vitro infectivity of Trypanosoma cruzi

Abstract

Background: The current drugs for Chagas Disease caused by the protozoan Trypanosoma cruzi have limited therapeutic potential and are associated with serious side effects. Natural products can aid to develop new chemotherapeutic agents. Several natural coumarins, especially Mammea A/BA, have shown significant activity against T. cruzi and low toxicity on human lymphocytes, but its effectivity on a wide range of strains need to be tested, as well as to deepen in their mode of action and safety.

Hypothesis/purpose: To discern the effects and explore the action mechanisms of mammea A/BA and a mixture of mammea coumarins isolated from Calophyllum brasiliense on Mexican strains of T. cruzi belonging to different genotypes and compare its effectivity with the drug benznidazole.

Study design: We evaluated the trypanocidal activity in vitro of mammea A/BA (93.6%), and a mixture of coumarins, mammea A/BA + A/BB + A/BD (86:10:1%) on Mexican T. cruzi strains belonging to different genotypes Ninoa, Querétaro (TcI) and Ver6 (TcVI).

Material and methods: Mammea A/BA and the mixture of coumarins, were isolated from Calophyllum brasiliense, identified by proton NMR and purity determined by HPLC. The in vitro trypanocidal activity was evaluated on mobility, growth recovery, morphology and infectivity of T. cruzi. The cytotoxicity on mammalian cells was compared with benznidazole. The ultrastructure of the treated epimastigotes was analyzed by transmission electron microscopy (TEM).

Results: Mammea A/BA and the mixture of coumarins showed high trypanocidal activity, affecting the mobility, growth recovery, morphology, ultrastructure of epimastigotes, and drastically reduce trypomastigotes infectivity on Vero cells. These substances were four times more potent than benznidazole and showed low cytotoxicity and high selectivity index. The TEM showed severe alterations on the plasmatic membrane, nuclear envelope, as well as, mitochondrial swelling, that leads to the death of parasites.

Conclusion: Mammea A/BA (93.6%) and a mixture of mammea A/BA + A/BB and A/BD (86: 10: 1%) isolated from the tropical tree C. brasiliense showed higher trypanocidal activity than the current drug benznidazole on three Mexican strains of T. cruzi. These compounds induced severe physiological and morphological alterations. These results suggest their possible use in preclinical studies.

Keywords: Calophyllum brasiliense; Mammea type coumarins; Trypanocidal activity; Trypanosoma cruzi.