Planta Med 2014; 80 - SL4
DOI: 10.1055/s-0034-1394492

Metabolism of vitetrifolin D in human liver microsomes and S9 cells: Analyses by LC/TOF-MS/MS and LC-SPE-NMR

S Sturm 1, C Högner 1, C Seger 1, 2, H Stuppner 1
  • 1Institute of Pharmacy/Pharmacognosy, Center for Molecular Biosciences, University of Innsbruck, Innrain 80 – 82, Innsbruck, Austria
  • 2Institute of Medical and Chemical Laboratory Diagnostics (ZIMCL), University Hospital/Landeskrankenhaus Innsbruck, Anichstraße 35, A-6020 Innsbruck, Austria

Fruits of Vitex agnus-castus L. (Lamiaceae), also called Monk's Pepper, are widely used for the treatment of premenstrual syndrome, menstrual disorders (amenorrhea, dysmenorrhea) and also symptoms of the menopause [1,2]. The mechanism of action is not fully understood but extracts of this herb have been shown to have dopaminergic effects resulting in changes of prolactin secretion [3,4]. Diterpenes such as rotundifuran and vitetrifolin D are considered as at least one of the active principles since these have been found to have affinity for dopamine D-2 receptors in vitro [3,4]. Only little is known about the metabolism of these constituents. Hence we decided to investigate the in vitro metabolism of vitetrifolin D qualitatively with the aim to structurally characterize as many phase I and phase II metabolites as possible. Metabolism studies were performed in human liver microsomes via cytochromes P450 (phase I) and in human S9-cells via UDP-glucuronosyltransferase (phase II) or sulfotransferase (phase II) mediated reactions. LC/TOF-MS/MS experiments allowed the identification of 20 metabolite peaks including sum formulae suggestions; high resolution MS/MS experiments led to the identification of various oxidative and reductive modifications of vitetrifolin D in phase I biotransformation and conjugation products with glucuronic acid in phase II reactions. Utilizing LC-SPE-NMR spectroscopy the structures of seven of the twenty metabolites tentatively identified by LC-MS were unambiguously confirmed. Only combining LC/TOF-MS/MS spectroscopy as metabolite profile scout experiment with LC-SPE-NMR giving access to NMR spectra as data base for a confirmative structure elucidation approach enabled us to tackle this analytical problem and to procure a feasible analytical base for envisioned in vivo experiments.

Acknowledgements: Supported by Bionorica research GmbH (Innsbruck, Austria).

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