Examination of a long-term clozapine administration by high resolution segmental hair analysis

Abstract

The long-term administration of clozapine could be verified by fine segmentation and analysis of single hairs of one person to examine the history of a multiple poisoning case.Segments of 1–2.5mm length were extracted by ultrasonification in 30μl of the mobile phase (mixture of methanol+water, 50+50). By application of isocratic liquid chromatography and using narrow bore columns (Synergy Polar-RP, Phenomenex), an acceleration and miniaturization of the HPLC–MS–MS assay could be achieved.Total amounts of clozapine down to 30fg (on column) and its desmethyl metabolite could be analysed in multiple reaction monitoring mode. According to typical sample amounts of ∼16μg, relevant hair concentrations higher than 1pg/mg were detected.Significant and reproducible concentration profiles along the hair fibres revealed characteristic administration cycles. The administration time course -in particular the time of its termination—could be verified with a precision of a few days. The accuracy and reproducibility of the concentration profile was proven based on multiple investigations of single hairs. An individual hair growth rate of 0.55mm/day was determined with a relative standard deviation of 8% by comparison of concentration profiles in hairs collected after a time span of 165 days.