Simple and cost-effective liquid chromatographic method for determination of pyrimethamine in whole blood samples dried on filter paper

A cost-effective HPLC method for determination of pyrimethamine (PYR) in human whole blood samples dried on filter paper (Whatman®) is reported. Trimethoprim (TMP) was used as an internal standard. Whole blood spiked with PYR was transferred (100 μl) onto filter paper and dried at room temperature. Capillary blood samples (100 μl) after ingestion of three tablets of sulfadoxine-pyrimethamine (SP) by one subject were also tested. PYR and an internal standard (IS) TMP were extracted into di-isopropyl ether as bases and then re-extracted with 150 μl mobile phase. A C-18 column was used and the mobile phase consisted of phosphate buffer (0.05 M, pH 5):acetonitrile:concentrated perchloric acid (750:300:2.5, v/v/v). sorbances of PYR and IS were monitored at 270 nm. The limit of quantification was 40 ng/ml. The within- and between-assay coefficient of variations were <10% at the limit of quantification.