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  • Title: Separation of the major adrenal steroids by reversed-phase high-performance liquid chromatography.
    Author: Capp MW, Simonian MH.
    Journal: Anal Biochem; 1985 Jun; 147(2):374-81. PubMed ID: 2990253.
    Abstract:
    The major adrenal steroids were separated by multistep gradient elution with a reversed-phase high-performance liquid chromatography system, employing water and 1-propanol as solvents. With this solvent system, a wide range of 21 5-ene-3 beta-ol and 4-ene-3-one steroids can be resolved in a single chromatogram, which was not possible with previously published gradient solvent systems. In particular, intermediate steroids of the biosynthetic pathway, 17 alpha-hydroxyprogesterone, 17 alpha-hydroxypregnenolone, and dehydroepiandrosterone, were separated with baseline or sufficient resolution to allow accurate quantitation. Using the 1-propanol-water gradient, the separations of 5-ene and 4-ene steroids were compared on different octadecylsilyl packings. Optimum resolution was obtained with a fully covered, spherical particle. The 1-propanol-water gradient was compared to a previously published methanol-water gradient in the analysis of steroidogenesis by adrenocortical cell cultures. HPLC analysis of the steroid production was quantitatively the same with both gradient solvent systems. However, qualitatively, the methanol-water gradient system did not resolve the above-mentioned intermediate steroids.
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