Determination of Mitotane (o,p`-DDD) and its Metabolites o,p`-DDA and o,p`-DDE in Plasma by High-Performance Liquid Chromatography

Summary: The adrenolytic agent mitotane (o,p`-DDD or 1,1-(o,p`-dichlorodiphenyl)-2,2-dichoroethane) is believed to be metabolically activated, resulting in the generation of reactive intermediates or the formation of o,p`-DDA. A new high-performance liquid chromatographic (HPLC) method for measurement of mitotane and two of its metabolites, o,p`-DDA (1,1-(o,p`-dichlorodiphenyl) acetic acid) and o,p`-DDE (1,1-(o,p`-dichlorodiphenyl)-2,2 dichloroethene), in plasma has been developed. Sample preparation involves precipitation of plasma proteins with acetone before chromatographic separation under isocratic conditions on a silica-based diphenyl column. Mitotane and metabolites are quantified by ultraviolet detection at 218 nm. Recoveries for the three compounds range between 99% and 110%, with between-day and within-day coefficients of variation <6% within the therapeutic range. Limits of detection are 0.5 μM and the assay is linear up to at least 800 μM for o,p`-DDA and 100 μM for mitotane and o,p`-DDE. The method has been evaluated using samples from three patients on mitotane therapy, revealing o,p`-DDA levels in plasma 3 to 10 times higher than the levels of the parent compound.