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Feasibility of a liquid-phase microextraction sample clean-up and liquid chromatographic/mass spectrometric screening method for selected anabolic steroid glucuronides in biological samples

Authors

  • Tiia Kuuranne,

    1. Division of Pharmaceutical Chemistry, Department of Pharmacy, P.O. Box 56, 00014 University of Helsinki, Finland
    2. Doping Control Laboratory, United Laboratories Ltd, Höyläämötie 14, 00380 Helsinki, Finland
    3. Viikki Drug Discovery and Technology Center, Department of Pharmacy, P.O. Box 56, 00014 University of Helsinki, Finland
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  • Tapio Kotiaho,

    1. Viikki Drug Discovery and Technology Center, Department of Pharmacy, P.O. Box 56, 00014 University of Helsinki, Finland
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  • Stig Pedersen-Bjergaard,

    1. School of Pharmacy, University of Oslo, P.O. Box 1068 Blindern, 0316 Oslo, Norway
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  • Knut Einar Rasmussen,

    1. School of Pharmacy, University of Oslo, P.O. Box 1068 Blindern, 0316 Oslo, Norway
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  • Antti Leinonen,

    1. Doping Control Laboratory, United Laboratories Ltd, Höyläämötie 14, 00380 Helsinki, Finland
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  • Steven Westwood,

    1. National Analytical Reference Laboratory, Australian Government Analytical Laboratories, P.O. Box 385, Pymble, N.S.W. 2073, Australia
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  • Risto Kostiainen

    Corresponding author
    1. Division of Pharmaceutical Chemistry, Department of Pharmacy, P.O. Box 56, 00014 University of Helsinki, Finland
    2. Viikki Drug Discovery and Technology Center, Department of Pharmacy, P.O. Box 56, 00014 University of Helsinki, Finland
    • Division of Pharmaceutical Chemistry, Department of Pharmacy, P.O. Box 56, 00014 University of Helsinki, Finland.
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Abstract

Anabolic androgenic steroids (AAS) are metabolized extensively in the human body, resulting mainly in the formation of glucuronide conjugates. Current detection methods for AAS are based on gas chromatographic/mass spectrometric (GC/MS) analysis of the hydrolyzed steroid aglycones. These analyses require laborious sample preparation steps and are therefore time consuming. Our interest was to develop a rapid and straightforward method for intact steroid glucuronides in biological samples, using liquid-phase microextraction (LPME) sample clean-up and concentration method combined with liquid chromatographic/tandem mass spectrometric (LC/MS/MS) analysis. The applicability of LPME was optimized for 13 steroid glucuronides, and compared with conventional liquid–liquid extraction (LLE) and solid-phase extraction (SPE) procedures. An LC/MS/MS method was developed for the quantitative detection of AAS glucuronides, using a deuterium-labeled steroid glucuronide as the internal standard. LPME, owing to its high specificity, was shown to be better suited than conventional LLE and SPE for the clean-up of urinary AAS glucuronides. The LPME/LC/MS/MS method was fast and reliable, offering acceptable reproducibility and linearity with detection limits in the range 2–20 ng ml−1 for most of the selected AAS glucuronides. The method was successfully applied to in vitro metabolic studies, and also tested with an authentic forensic urine sample. For a urine matrix the method still has some unsolved problems with specificity, which should be overcome before the method can be reliably used for doping analysis, but still offering additional and complementary data for current GC/MS analyses. Copyright © 2003 John Wiley & Sons, Ltd.

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