Quantitation of catechol estrogens and their N-acetylcysteine conjugates in urine of rats and hamsters

A method for the analysis of N-acetylcysteine conjugates of catechol estrogens [catechol estrogen mercapturates (CE SRs)], which are likely to be urinary markers of estrogen-induced tumors, was established in this study. The characteristics of the method that was established were ( I) cleanup of urine using the immunoaffinity column of CE SRs, (2) detection of catechol estrogens (CEs) and CE SRs by electrochemical detection, which provided the high specificity, and (3) stability of CE SRs through the cleanup. Using this method, the simultaneous quantitation of 2-hydroxy-17 beta -estradiol (2-OHE2), 4-hydroxy-17 beta -estradiol (4-OHE2), 2-hydroxyestrone (2-OKE1), 4-hydroxyestrone (4-OHE1), 2-hydroxyestrone 1-N-acetylcysteine thioether (2-OKE1 ISR), 2-hydroxyestrone 4-N-acetylcysteine thioether (2-OHE1 4SR), and 4-hydroxyestrone 2-N-acetylcysteine thioether (4-OHE1 2SR) in the range of 1-15 ng was performed. We first demonstrated the presence of CE SRs, 2-OHE1 1SR and 2-OHE1 4SR, in urine from rats treated intraperitoneally with 17 beta -estradiol (E-2) at a dose of 5 mg/kg. In female rats, the amount of 2-OHE1 ISR was several-fold greater than that of 2-OHE1 4SR, while the presence of 4-OHE1 2SR was not confirmed. The level of CEs and CE SRs in male rats was approximately 1/2-1/20 of that in female rats. The excretion rate following administration of 2-OHE1 at 2 mg/kg and that following the administration of 4-OHE1 at 2 mg/kg were different in female rats. In addition, 4-OHE1 2SR was present in the urine of male Syrian hamsters treated intraperitoneally with E-2, whereas it was absent in rats.