A bioanalytical method using liquid chromatography–tandem mass spectrometry (LC/MS/MS) was developed and validated for the determination of clonidine in human serum. Clonidine and internal standard clonidine-d4 were extracted from alkalized human serum using liquid/liquid extraction. The extract was evaporated to dryness, reconstituted, and injected onto a Betasil silica column (50 mm × 3 mm) using an aqueous-organic mobile phase consisting of acetonitrile–water–formic acid (80 : 20 : 1, v/v/v). The flow rate was 0.7 mL/min. The chromatographic run time was 2.5 min per injection, with retention time of 1.7 min for clonidine. The detection was by monitoring clonidine at m/z 230→213 and IS at m/z 236→219, respectively. The standard curve range was 10–1000 pg/mL. The low limit of quantitation (LLOQ) is 10 pg/mL. The inter-day precision and accuracy of the quality control (QC) samples were <6.4% relative standard deviation (RSD) and <7.9% relative error (RE). The recovery of extraction was 95% for clonidine and 90% for the internal standard.
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Pelzer et al. (2002) studied this question.
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