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Determination of mitotane and its metabolites in biological fluids by SPE-HPLC method

Mitotane is the drug of choice for the treatment of adrenal cortical carcinoma and is used as a chemotherapeutic agent after resection. Its mechanism of action and pharmacokinetics are not fully known. To achieve optimal therapeutic concentration and to prevent dangerous side effects, it is very important to determinate the concentrations levels of mitotane and its metabolites in the body after administration. The aim of this work was to develop and validate a SPE-HPLC method for determination of mitotane and its metabolites in plasma, erythrocytes and urine of a patient with adrenal cortical carcinoma. Blood and urine samples were collected during 8 hours after administration of Lysodrena® (500 mg) pill. SPE conditions: Discovery DSC column, 500 mg, 3 mL (Supelco), preconditioned with 1 mL of methanol and 2 mL of acetonitrile. HPLC conditions: Symmetry C18 column (Waters), 150 × 4.6 mm ; 3.5 μm. Mobile phase: acetonitrile : MiliQ water : formic acid = 90 : 10 : 0.1 ; flow rate 1 mL/min ; DAD detector λmax 230 nm. Method was validated according to ICH guidelines and following parameters were examined: selectivity, linearity, limit of detection and determination, accuracy, precision and robustness. The proposed method was successfully applied for determination of mitotane and its metabolites in plasma, erythrocytes and urine.

mitotane; metabolites; Solid phase extraction; HPLC; biological fluids

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