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  • Title: Analysis of steroid conjugates in sewage influent and effluent by liquid chromatography-tandem mass spectrometry.
    Author: Reddy S, Iden CR, Brownawell BJ.
    Journal: Anal Chem; 2005 Nov 01; 77(21):7032-8. PubMed ID: 16255606.
    Abstract:
    Environmental endocrine disruptors such as estrone (E1) and beta-estradiol (E2) are excreted in human urine primarily as water-soluble glucuronides and sulfates that can dissociate in wastewater treatment systems to the more active free estrogens. Measurement of the distribution and fate of the steroid conjugates and the corresponding free estrogens in treatment plants and receiving waters is critical for understanding the reproductive and developmental effects of these substances on aquatic organisms. A sensitive method to measure steroid estrogen conjugates in matrix-rich sewage influents and effluents (method detection limits ranged from 0.04 to 0.28 ng/L) has been developed using HPLC tandem mass spectrometry with electrospray ionization. The method employs extensive sample purification by selective extraction from an Oasis HLB solid-phase cartridge followed by separation by anion exchange chromatography. This purification scheme, combined with a stable isotope dilution approach, was used to overcome problems of matrix suppression of ionization and permitted selective and sensitive detection of six target conjugates of E1 and E2. Accurate quantitation was highly dependent on the method of sample preservation. Acidification of each sample (pH 2.0) was effective in preventing enzymatic or chemical decomposition of steroid conjugates in all sample types, whereas glucuronide conjugates were hydrolyzed in the presence of mercury and formalin preservatives. Measured concentrations of steroid sulfates in the influent to a sewage treatment plant were approximately 100 times greater than that of the respective steroid glucuronides, suggesting that the preponderance of glucuronides had dissociated prior to reaching the treatment plant. A small percentage of the steroid sulfates persisted through biological treatment of sewage and was measured in the effluent. Steroid conjugates that survive decomposition or bypass biological treatment of municipal wastewater are released into surface waters and may serve as a source of free steroids.
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