Corona Giuseppe, Elia Caterina, Casetta Bruno, Diana Crivellari, Rosalen Sara, Bari Mario, Toffoli Giuseppe
Division of Experimental and Clinical Pharmacology, Department ofMolecular Biology and Translational Research, National Cancer Institute and Center for Molecular Biomedicine, Aviano (PN), Italy.
J Mass Spectrom. 2009 Jun;44(6):920-8. doi: 10.1002/jms.1566.
A simple and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has been developed and validated for the quantitation of exemestane (Exe) and its main metabolite 17-dihydroexemestane (DhExe) in human plasma. The analytes were extracted by protein precipitation with acetonitrile, containing stable 13C-labelled Exe (13C3-Exe) as internal standard, and measured by LC-MS/MS. The best chromatographic separation of the analytes from the interferences was achieved by using a Phenyl column operating under isocratic regime conditions. The total chromatographic runtime was 5.0 min and the elution of Exe and DhExe occurred at 2.5 min and 2.9 min, respectively. Quantitation was performed by employing the positive electrospray ionization (ESI) technique and multiple reaction monitoring mode (MRM). The monitored precursor to product-ion transitions for Exe, DhExe and 13C3-Exe internal standard were m/z 297.0 --> 120.8, m/z 299.1 --> 134.9 and m/z 300.0 --> 123.2, respectively. The lower limit of quantitation (LLOQ) was 0.1 ng/ml for DhExe and 0.2 ng/ml for Exe. The method was linear up to 36-51 ng/ml with r2 > or = 0.998. The intra- and inter-assay precision were < or = 7.7% and 5.1% for Exe and < or = 8.1 and 4.9% for DhExe while deviations from nominal values were in the 1.5-13.2% and - 9.0-5.8% ranges for Exe and DhExe, respectively. The analytical method resulted robust and suitable for pharmacokinetic monitoring of Exe and its main metabolite during adjuvant therapy in patients with breast cancer.
已开发并验证了一种简单且灵敏的液相色谱 - 串联质谱（LC - MS/MS）方法，用于定量测定人血浆中的依西美坦（Exe）及其主要代谢物17 - 二氢依西美坦（DhExe）。采用含稳定的13C标记依西美坦（13C3 - Exe）作为内标的乙腈进行蛋白沉淀提取分析物，然后通过LC - MS/MS进行测定。通过使用在等度洗脱条件下运行的苯基柱，实现了分析物与干扰物的最佳色谱分离。总色谱运行时间为5.0分钟，依西美坦和17 - 二氢依西美坦的洗脱时间分别为2.5分钟和2.9分钟。采用正电喷雾电离（ESI）技术和多反应监测模式（MRM）进行定量。依西美坦、17 - 二氢依西美坦和13C3 - Exe内标的监测前体离子到产物离子的跃迁分别为m/z 297.0 --> 120.8、m/z 299.1 --> 134.9和m/z 300.0 --> 123.2。17 - 二氢依西美坦的定量下限（LLOQ）为0.1 ng/ml，依西美坦为0.2 ng/ml。该方法在36 - 51 ng/ml范围内呈线性，r2≥0.998。依西美坦的批内和批间精密度分别≤7.7%和5.1%，17 - 二氢依西美坦分别≤8.1%和4.9%，而依西美坦和17 - 二氢依西美坦与标称值的偏差分别在1.5 - 13.2%和 - 9.0 - 5.8%范围内。该分析方法结果可靠，适用于乳腺癌患者辅助治疗期间依西美坦及其主要代谢物的药代动力学监测。
