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Testing time for testosterone -
07-01-2008
Testing time for testosterone A new analytical approach to testing for testosterone and related steroids in body fluids using non-specific micellar electrokinetic chromatography (MEKC) has been developed. The partial filling (PF) technique was employed, which means the approach is suitable for detection by either ultraviolet spectrophotometry or electrospray ionization mass spectrometry. With sporting events continuing to push the boundaries of human capabilities in terms of achievements, gambling and financial rewards, it is perhaps inevitable that some participants at the highest level will be tempted to chemically enhance those capabilities. There is thus an ongoing battle between athletes using illicit steroids to pump up their muscles and the sports authorities using dope testing science to identify and quantifytestosterone and related compounds in body fluids. "Testosterone is the principal androgen, produced in men mainly by the testes and in women by the ovaries and the adrenal glands, writes Lotta Amundsen, "it has both anabolic and androgenic effects in the human body." Amundsen working at the VTT Technical Research Centre of Finland and the Laboratory of Analytical Chemistry, Department of Chemistry, Faculty of Science, University of Helsinki, Finland, explains that capillary electromigration techniques are a relatively new approach forsteroid research. Capillary electrophoresis was introduced by Jorgenson and Lukacs in 1981 and rapidly became the separation tool of choice in clinical, pharmaceutical, and bioanalytical studies across the globe. Today it is a routine laboratory technique based on separating materials based on charge, mass, and three-dimensional structure properties. However, such capillary electromigration techniques suffer from a significant drawback in that steroids are electrically neutral and so their separation by capillary techniques requires the use of charged electrolyte additives that interact with the steroids either specifically or non-specifically. Working under Heli Sirén at VTT, Amundsen, has developed an efficient, quantitative PF-MEKC-UV method for steroid standards. An optimized pseudostationary phase comprising surfactants and cyclodextrins, macrocyclic starch molecules resembling a doughnut in shape. She has found that PF-MEKC-UV is a more sensitive, efficient and repeatable method for testing steroids than the mass spectrometric equivalent, PF-MEKC-ESI-MS. Indeed, ESI-MS interfacing sets significant limitations not only on the chemistry affecting the ionization and detection processes of neutral steroids, but also on their separation. "The new PF-MEKC-UV method was successfully employed in the determination of testosterone in male urine samples after microscale immunoaffinity solid-phase extraction (IA-SPE)," explains Amundsen, "The IA-SPE method, relying on specific interactions betweentestosterone and a recombinant anti-testosterone Fab fragment, is the first such method described for testosterone." As part of the study, the researchers have obtained new data for the interactions of steroids and human, or bovine serum albumins, using affinity capillary electrophoresis. This data was then used to devised a new algorithm for the calculation of association constants between proteins and neutral ligands, such as steroids. "The new capillary electromigration methods provide versatile tools for steroid analysis. The use of non-specific interactions between analytes and electrolyte additives makes possible an efficient separation and quantification of steroids. Finally, the use of specific interactions between steroids and biomolecules may provide interesting and accurate data difficult to obtain by any other technique," concludes Admundsen. The new technique could find application not only in sports doping control but in medical diagnostics where steroid assays are just as important. source: spectroscopynow.com 
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