Determination of 17-oxosteroid glucuronides and sulfates in urine and serum by fluorescence high-performance liquid chromatography using dansyl hydrazine as a prelabeling reagent.
A fluorescence high-performance liquid chromatographic method is described for the direct determination of conjugated 17-oxosteroids in biological fluids without hydrolysis. Conjugated 17-oxosteroids are extracted with Sep-Pak C18 cartridge, labeled with dansyl hydrazine in trichloroacetic acid--benzene solution and then separated by high-performance liquid chromatography on reversed-phase muBondapak C18 column using 0.01 M sodium acetate in methanol-water-acetic acid (65:35:1, v/v) as the mobile phase. The eluate is monitored by a fluorophotometer at 365 nm (excitation) and 520 nm (emission). Linearities of fluorescence intensities (peak heights) with the amounts of various conjugated 17-oxosteroids were obtained between 10 pmol and 100 pmol. This method is sensitive, reliable and useful for the simultaneous determination of conjugated 17-oxosteroids in urine and serum.
D015068:17-Ketosteroids / Q000032:analysis* / Q000097:blood / Q000652:urine; D000328:Adult; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003619:Dansyl Compounds; D005260:Female; D005453:Fluorescence; D006801:Humans; D006834:Hydrazines; D006868:Hydrolysis; D008297:Male; D008875:Middle Aged
D015068:17-Ketosteroids; D003619:Dansyl Compounds; D006834:Hydrazines; C027913:dansyl hydrazine