Studies on steroids. CCXXIX. Separation and characterization of catechol oestrogen glucuronides in urine of pregnant women by high-performance liquid chromatography.
The separation and characterization of catechol oestrogen glucuronides in the urine of pregnant women by high-performance liquid chromatography with electrochemical detection is described. The urine samples from pregnant women were passed through Amberlite XAD-2 and Sep-Pak C18 columns and subsequented to ion-exchange chromatography on piperidinohydroxypropyl Sephadex LH-20 to isolate the oestrogen glucuronide fraction. The subsequent resolution into individual oestrogen glucuronides was achieved by high-performance liquid chromatography on a reversed-phase column. 2-Hydroxyoestrone 2-glucuronide and 4-hydroxyoestrone 4-glucuronide were identified on the basis of their behaviour in high-performance liquid chromatography with three different mobile phases. Derivatization of two glucuronides with 2-ferrocenylethylamine, followed by chromatographic separation and measurement of hydrodynamic voltammograms with an electrochemical detector was carried out for unequivocal identification. Enzymatic cleavage of the glucuronoside linkage followed by the identification of deconjugated catechol oestrogens by high-performance liquid chromatography with electrochemical detection further supported the structural assignment of these metabolites. The ratio between the amounts of 2-hydroxyoestrone 2-glucuronide and 4-hydroxyoestrone 4-glucuronide excreted in the urine of pregnant women was found to be ca. 5:1.
Shimada
K
K
Pharmaceutical Institute, Tohoku University, Sendai, Japan.
Xie
F M
FM
Niwa
T
T
Wakasawa
T
T
Nambara
T
T
D002851:Chromatography, High Pressure Liquid; D004563:Electrochemistry; D004964:Estriol / Q000652:urine; D002393:Estrogens, Catechol / Q000652:urine*; D005260:Female; D005296:Ferrous Compounds; D005965:Glucuronates / Q000652:urine*; D005966:Glucuronidase; D006801:Humans; D006868:Hydrolysis; D009942:Organometallic Compounds; D011247:Pregnancy; D013256:Steroids / Q000652:urine*
D002393:Estrogens, Catechol; D005296:Ferrous Compounds; D005965:Glucuronates; D009942:Organometallic Compounds; D013256:Steroids; C054010:2-ferrocenylethylamine; D005966:Glucuronidase; D004964:Estriol