DETERMINATION OF THE NEW AROMATASE INHIBITOR EXEMESTANE IN BIOLOGICAL-FLUIDS BY AUTOMATED HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY FOLLOWED BY RADIOIMMUNOASSAY

A procedure for the determination of exemestane, a new aromatase inhib itor, in biological fluids is described in this paper. Exemestane is e xtracted from human plasma and urine by solid-phase and liquid-liquid extraction, respectively. The test compound is then isolated from endo genous steroids, its metabolites and/or degradation products by HPLC. The exemestane-containing fraction is collected and its exemestane con tent measured by radioimmunoassay (RIA). The automated HPLC system all owed a high specificity and reproducibility of retention times, and el iminated almost all manual operations. The RIA allowed the accurate an d precise measurement of 12 pg of exemestane/ml in plasma (inter- and intra-assay RSD=17.7 and 13.4%, respectively) and 25 pg/ml in urine (i nter- and intra-assay RSD=14.5% and 8.7%, respectively). The recovery of the whole procedure was evaluated by comparison of the RIA calibrat ion curve obtained in plasma or urine (after extraction and HPLC) with that obtained directly in RIA buffer (without extraction and HPLC). T he calibration curves were practically superimposable, indicating that the recovery of the whole procedure was excellent. The method was val idated in terms of reproducibility, recovery and precision in the rang e 10-500 pg of exemestane/ml of plasma and 20-1000 pg/ml of urine. Fin ally the plasma levels of exemestane in a postmenopausal healthy volun teer treated daily for 7 days with oral exemestane at a dose of 1 mg ( the lowest dose administered in clinical trials)ere monitored using th e method here described. Exemestane was detectable in all plasma sampl es collected (up to 24 h after drug intake). Therefore the analytical method described here should be sufficiently sensitive and specific fo r the determination of exemestane in plasma and urine from clinical tr ials in which therapeutic doses of the drug (10-25 mg/day) are adminis tered.