Determination of rifampicin in human plasma and blood spots by high performance liquid chromatography with UV detection: A potential method for therapeutic drug monitoring

A high performance liquid chromatography method has been developed that allows quantification of concentrations of rifampicin in human plasma and blood spots. Rifampicin and papaverine hydrochloride (internal standard) were extracted from plasma using a Strata-X-CW extraction cartridge. These analytes were also extracted into acetonitrile from blood spots dried onto a specimen collection card. The recovery of rifampicin from plasma and blood spots was 84.5% and 65.0%, respectively. Separation was achieved by HPLC on a Kromasil C18 column with a mobile phase composed of ammonium acetate (20 mM, pH 4.0) and acetonitrile, delivered on a gradient programme. Optimum detection was at 334 nm. The assay was linear over the concentration range of 0.5–20 μg/ml. The limit of quantification was 0.5 μg/ml in plasma; 1.5 μg/ml in blood spots. Both intraday and interday precision data showed reproducibility (R.S.D. ≤ 8.0, n = 9). Stability studies showed rifampicin was stable in plasma for up to 9 h after thawing; the samples were also stable for up to 9 h after preparation. Five patient samples were analysed using the methods described. A correlation was found between the concentrations of RIF in plasma and blood spots (r2 = 0.92). This method is proposed as a means of therapeutic drug monitoring of rifampicin in patients with tuberculosis.