Assay for mitolactol and its bifunctional alkylating metabolites in plasma

A method involving precolumn derivatization and high-performance liquid chromatography is described for the measurement of mitolactol levels in plasma. The basis of the assay is the reaction at pH 7.4 and 50 degrees C of mitolactol with diethyldithiocarbamate to form 1,6-bis(diethyldithiocarbamoyl)-2,3,4,5-tetrahydroxyhexane. This derivative is then extracted into chloroform, resolved by normal-phase chromatography, and detected by UV (254 nm) absorbance. The method quantitates the sum of mitolactol and its active bifunctional metabolites, bromoepoxydulcitol and dianhydrogalactitol, in plasma down to concentrations of 0.5 microM. The pharmacokinetic parameters of the drug in mice have been determined following the intraperitoneal injection of either 20 or 100 mg/kg of body weight. Absorption from the peritoneal cavity was largely complete by 5 min. Parameters obtained include a first-order elimination constant, k = 0.92 X 10(-2) min-1 and an apparent volume of distribution, Vd = 0.78 L/kg. For a 100-mg/kg dose, the area under the concentration-time curve was 49 mM X min, and the mean peak drug concentration was reached at 40 min following intraperitoneal injection. Concentrations of mitolactol in total plasma and in plasma ultrafiltrates were identical, indicating that the drug is not (less than 4%) reversibly bound to plasma proteins.