Zacharia Lefteris C, Jackson Edwin K, Kloosterboer Helenius J, Imthurn Bruno, Dubey Raghvendra K
Center for Clinical Pharmacology, Department of Medicine, University of Pittsburgh Medical Center, Pittsburgh, PA, USA.
Menopause. 2006 Nov-Dec;13(6):926-34. doi: 10.1097/01.gme.0000227331.49081.d7.
Tibolone, a hormone therapy drug, is used to treat climacteric symptoms. This drug is rapidly metabolized into three major metabolites (3alpha-hydroxytibolone, 3beta-hydroxytibolone, and Delta4-tibolone). One clinical study provided evidence of conversion of tibolone to another estrogenic metabolite, 7alpha-methyl-ethinyl estradiol (MEE). However, no evidence of MEE formation was found in another study using the human aromatase enzyme. Because MEE was analyzed by gas chromatography-mass spectrometry (GC-MS), which requires derivatization, together with the fact that derivatization of some steroids may lead to aromatization, it is feasible that the MEE detected resulted from an artifact generated during the derivatization process. Hence, our objective was to assess whether tibolone is converted to MEE.
We assayed MEE formation in a nonbiological system using GC-MS after derivatization and by analyzing MEE formation using liquid chromatography-mass spectrometry (LC-MS) in nonderivatized samples.
MEE formation was evident in tibolone samples derivatized with either pentafluoropropionic anhydride or trimethylsilyl and analyzed by GC-MS. The amount of MEE formed increased with increasing amounts of tibolone (0.5, 1, 2.5, and 5 microg) derivatized; however, relative to tibolone, the percentage of MEE formed remained constant and ranged between 0.22% and 0.29% of tibolone. In contrast to GC-MS, no MEE formation was seen when tibolone was analyzed by liquid chromatography-mass spectrometry without derivatization.
Our findings prove that conversion of tibolone to MEE is an artifact that is generated in a GC-MS system and is largely due to the intense heating step involved in GC-MS. Caution should be exercised to extrapolate clinical implications from existing data on MEE formation using a GC-MS system.
