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OBJECTIVES To evaluate the population pharmacokinetics of different benznidazole treatment regimens and the drug's biodistribution in mice. METHODS Two hundred mice were divided into five groups according to benznidazole dosing regimens (1) 100 mg/kg/day for 20 days; (2) 100 mg/kg/day for 40 days; (3) 200 mg/kg/day for 20 days; (4) 40 mg/kg/day for 20 days; or (5) 40 mg/kg/day for 40 days. The mice were euthanized and blood, heart, liver, colon and brain were collected. Samples were prepared by liquid-liquid extraction and analysed by H