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Clinical Chemistry 43: 786-793, 1997;
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(Clinical Chemistry. 1997;43:786-793.)
© 1997 American Association for Clinical Chemistry, Inc.

Articles

Convenient chromatographic prepurification step before measurement of urinary cortisol by radioimmunoassay

Gilles Morineau1,a, James Gosling2, Marie-Claude Patricot3, Hany Soliman1, Philippe Boudou1, Akram Al Halnak1, Gaëlle Le Brun1, Jean-Louis Brérault1, René Julien1, Jean-Marie Villette1 and Jean Fiet1,4

1 Laboratoire de Biologie Hormonale, Hôpital Saint-Louis, 75010 Paris, France.

2 Department of Biochemistry and National Diagnostics Centre, University College, Galway, Ireland.

3 Laboratoire d'Hormonologie, bâtiment 3B, Centre hospitalier Lyon-Sud, 69495 Pierre-Benite cedex, France.

4 Laboratoire de Biochimie, Faculté de pharmacie, 75006 Paris, France.
a Address correspondence to this author at: Laboratoire de Biologie Hormonale, Hôpital Saint-Louis, 1 ave. Claude Vellefaux, 75475 Paris cedex 10, France. Fax Int + 33 1 42 49 42 80; e-mail bio.horm.fiet{at}chu-stlouis.fr

We applied various prepurification protocols (extraction with different solvents, liquid/solid separation on bonded silica media, Celite, and Sephadex LH20 chromatography) with a range of commercially available RIA kits to measure cortisol in urine samples. We then compared the results with the concentrations measured by a HPLC method validated with reference to isotope dilution gas chromatography–mass spectrometry. We conclude that chromatography on a commercial, prepacked diol minicolumn (WatersTM Sep-Pak Vac RC) in combination with dichloromethane extraction is a convenient and very effective purification step before RIA of urinary cortisol in patients not receiving corticoid medication. We tested numerous steroids for interference and found that free polar cortisol derivatives (hydroxylated or hydrogenated) could only partially account for the overestimations routinely encountered when free urinary cortisol concentrations are measured by direct RIA.


Key Words: indexing terms: chromatography • Celite • Sephadex LH 20 • HPLC • ID GC-MS • diol minicolumns.




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Hypothalamic-Pituitary-Adrenal Axis Function after Inhaled Corticosteroids: Unreliability of Urinary Free Cortisol Estimation
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R J Meijer, D S Postma, and H A M Kerstjens
Conventional RIA underestimates cortisol suppression in the presence of prednisolone
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