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  • Title: Determination of 21-hydroxycorticosteroids in human urine by high-performance liquid chromatography with fluorescence detection.
    Author: Yamaguchi M, Yoshitake T, Ishida J, Nakamura M.
    Journal: Chem Pharm Bull (Tokyo); 1989 Nov; 37(11):3022-5. PubMed ID: 2632047.
    Abstract:
    A simple and sensitive high-performance liquid chromatographic method with fluorescence detection for the determination of nineteen 21-hydroxycorticosteroids is described. The corticosteroids are oxidized by cupric acetate to form the corresponding glyoxal derivatives. The derivatives are converted into fluorescent quinoxalines by reaction with 1,2-diamino-4,5-methylenedioxybenzene, a fluorogenic reagent for alpha-dicarbonyl compounds. The quinoxalines are separated within 70 min on a reversed-phase column (TSK gel ODS-120T) by stepwise elution with mixtures of methanol, acetonitrile, and 1.0 M ammonium acetate. The detection limits are 0.14-29.4 pmol at a signal-to-noise ratio of 3 in a 50-microliter injection volume. This sensitivity permits precise determination of hydrocortisone, cortisone, corticosterone, and their tetrahydro derivatives in 500 microliters of normal human urine.
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