FIG. 1.

TFQ reduces intracellular ATP level without plasma membrane permeabilization. (A) Changes in the intracellular ATP levels were determined by variation of luminescence in L. donovani 3-Luc promastigotes treated with different TFQ concentrations: 2 (▪), 5 (○), 8 (□), and 10 μM (•). Promastigotes were preloaded with 25 μM DMNPE-luciferin, and when luminescence reached a plateau, TFQ was added (t = 0) and luminescence was monitored as described in Materials and Methods. Variation of luminiscence was normalized relative to the level in the control untreated parasites. Similar results were obtained in three independent experiments. At the concentrations tested, TFQ did not inhibit the activity of recombinant firefly luciferase. (B) The effect of TFQ on the plasma membrane permeability was determined by incubating promastigotes without (control [c]) and with 1, 5, and 10 μM TFQ for 45 min in HBS at 28°C and then treating them with 2 μM SYTOX Green for 15 min at 28°C. SYTOX Green fluorescence is represented relative to parasites treated with 0.05% Triton X-100 used as 100% permeabilization. Results are means ± standard deviations (SD) from three independent experiments.