Assessment and validation of the MS/MS fragmentation patterns of the macrolide immunosuppressant everolimus

被引:29
作者
Boernsen, K. Olaf
Egge-Jacobsen, Wolfgang
Inverardi, Bruno
Strom, Tobin
Streit, Frank
Schiebel, Hans-Martin
Benet, Leslie Z.
Christians, Uwe [1 ]
机构
[1] Univ Colorado, Hlth Sci Ctr, Dept Anesthesiol, Clin Res & Dev, Denver, CO 80202 USA
[2] Novartis Pharma AG, Basel, Switzerland
[3] Univ Oslo, Dept Mol Biosci, Oslo, Norway
[4] Novartis Inst Biomed Res, Basel, Switzerland
[5] Univ Gottingen, Klin Chem Abt, D-3400 Gottingen, Germany
[6] Tech Univ Carolo Wilhelmina Braunschweig, Inst Chem, D-3300 Braunschweig, Germany
[7] Univ Calif San Francisco, Dept Biopharmaceut Sci, San Francisco, CA 94143 USA
来源
JOURNAL OF MASS SPECTROMETRY | 2007年 / 42卷 / 06期
关键词
everolimus; LC-MS/MS; fragmentation; H/D exchange; certican; hydroxyethyl rapamycin; structure; RAD;
D O I
10.1002/jms.1215
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Everollimus (40-O-(2-hydroxyethyl)rapamycin, Certican) is a 31-membered macrolide lactone. In lymphocytes, it inhibits the mammalian target of rapamycin (mTOR) and is used as an immunosuppressant after organ transplantation. Due to its instability in pure organic solvents and insufficient HPLC separation, NMR spectroscopy analysis of its metabolite structures is nearly impossible. Therefore, structural identification based on tandem mass spectrometry (MS/MS) and MSn fragmentation patterns is critical. Here, we have systematically assessed the fragmentation pattern of everolimus during liquid chromatography (LC)-electrospray ionization (ESI)-MS/MS and validated the fragment structures by (1) comparison with structurally identified derivatives (sirolimus), (2) high-resolution mass spectrometry, (3) elucidation of fragmentation pathways using ion trap mass spectrometry (up to MS') and (4) H/D exchange. In comparison with the structurally related immunosuppressants tacrolimus and sirolimus, our study was complicated by the low ionization efficiency of everolimus. Detection of positive ions gave the best sensitivity, and everolimus and its fragments were mainly detected as sodium adducts. LC-ESI-MS/MS of everollimus in combination with collision-induced dissociation (CID) resulted in a complex fragmentation pattern and the structures of 53 fragments were identified. These detailed fragmentation pathways of everolimus provided the basis for structural elucidation of all everolimus metabolites generated in vivo und in vitro. Copyright (c) 2007 John Wiley & Sons, Ltd.
引用
收藏
页码:793 / 802
页数:10
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