Fluorescence analysis of the interaction of isometamidium with Trypanosoma congolense

Isometamidium chloride (Samorin) is the only compound recommended for prophylaxis against bovine trypanosomiasis in sub-Saharan Africa. The fluorescence property of this compound was used to investigate the interaction of the molecule with in vitro-derived bloodstream forms of Trypanosoma congolense IL 1180. Incubation of isometamidium with trypanosomes at 37 degrees C for 180 min resulted in a gradual alteration of the lambda max. with time (from 600 to 584 nm) and an increase in the intensity of trypanosome-associated fluorescence of approx. 2-fold. The alteration in fluorescence was temperature-dependent and inhibited by the addition of N-ethylmaleimide. In contrast, with intact cells addition of digitonin caused a rapid increase in fluorescence intensity to approximately four times that observed with intact cells. Uptake of isometamidium was also determined using radiolabelled drug; the results indicated that the time course of the uptake process resembled the fluorescence profile and was temperature-dependent. The results therefore indicate that the alteration of fluorescence is due to interaction of isometamidium with an intracellular component(s) and that isometamidium is transported across the plasma membrane via a protein carrier. The data also indicate that the described fluorescence technique can be used to investigate the role of membrane transport in resistance to isometamidium.