Liquid chromatography-tandem mass spectrometric quantitation of cyclophosphamide and its hydroxy metabolite in plasma and tissue for determination oftissue distribution

Cyclophosphamide (CP) and its metabolite, hydroxycyclophosphamide (OH-CP) h ave been quantitated in mouse plasma and tissue by derivatization combined with liquid chromatography-tandem mass spectrometry (LC-MS-MS). The derivat ization was conducted immediately upon sample collection, to trap the OH-CP metabolite intermediate prior to further conversion to phosphoramide musta rd or other reaction products. This simple and straightforward derivatizati on procedure, combined with sample extraction via protein precipitation, al lowed quantitation of CP and the oxime derivative of OH-CP in plasma for co ncentrations ranging from approximately 12.5-3333 ng/ml, and in spleen tiss ue for concentrations of 1250-50 000 ng/g. The short cycle time (2.5 min) o f the LC-MS-MS method allowed high throughput analysis with minimal matrix interference. Mouse plasma levels were quantitated for doses of 40, 65 and 120 mg/kg; spleen concentrations were determined for mice dosed at 120 mg/k g. The CP acid oxime plasma levels correlated well with dose amounts. The C P levels in the spleen and plasma were similar while the oxime levels in th e spleen were significantly lower than the plasma. (C) 2001 Elsevier Scienc e B.V. All rights reserved.