A simple, rapid, and accurate reversed phase ultra-performance liquid chromatographic (RP-UPLC) method has been developed and subsequently validated for the simultaneous determination of proguanil (PRG) and Atovaquone (ATV) in combination. The separation is carried out using a mobile phase consisting of 0.05% perchloric acid and Acetonitrile. The column used is Extended C18(50 × 3.0 mm, 1.8 µm). With flow rate of 0.8 mL/min using UV detection at 257nm. The total run time is 6 min and the

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... on time of PRG and ATV is 1.6 minand 3.0 min respectively. The described method is linear for the assay of PRG and ATV over a concentration range of 2.5-20 µg/Ml and 6.25-50 µg/mL respectively. Results of the analysis have been validated statistically and by recovery studies. The limit of quantitation for PRG and ATV has been found to be 0.0357 µg/mL and 0.126 µg/mL respectively. The results of the studies showed that the proposed RP-UPLC method is simple, rapid, precise, and accurate, which is useful for the routine determination of PRG and ATV in bulk drug and its pharmaceutical dosage form.