Microbial Degradation of Free and Halogenated Estrogens in River Water-Sediment Microcosms.

Halogenated estrogens are formed during chlorine-based wastewater disinfection and have been detected in wastewater treatment plant effluent; however, very little is known about their susceptibility to biodegradation in natural waters. To better understand the biodegradation of free and halogenated estrogens in a large river under environmentally relevant conditions, we measured estrogen kinetics in aerobic microcosms containing water and sediment from the Willamette River (OR, USA) at two concentrations (50 and 1250 ng L-1). Control microcosms were used to characterize losses due to sorption and other abiotic processes, and microbial dynamics were monitored using 16S rRNA gene sequencing and ATP. We found that estrogen biodegradation occurred on timescales of hours to days and that in river water spiked at 50 ng L-1 half-lives were significantly shorter for 17ß-estradiol (t1/2,bio = 42 ± 3 h) compared to its monobromo (t1/2,bio = 49 ± 5 h), dibromo (t1/2,bio = 88 ± 12 h), and dichloro (t1/2,bio = 98 ± 16 h) forms. Biodegradation was also faster in microcosms with high initial estrogen concentrations as well as those containing sediment. Free and halogenated estrone were important transformation products in both abiotic and biotic microcosms. Taken together, our findings suggest that biodegradation is a key process for removing free estrogens from surface waters but likely plays a much smaller role for the more highly photolabile halogenated forms.

Photochemical degradation of halogenated estrogens under natural solar irradiance.

Halogenated estrogens are thought to be moderately potent endocrine-disrupting compounds that are formed during chlorine-based wastewater disinfection processes and may represent a significant fraction of the total amount of estrogen delivered from wastewater treatment plants to receiving waters. Yet we lack key information about the photochemical degradation of halogenated estrogens, a process that has important implications for UV-based wastewater treatment and environmental fate modeling. To better understand halogenated estrogen degradation in aquatic environments, we studied the direct photolysis of 17ß-estradiol (E2), 2-bromo-17ß-estradiol (monoBrE2), 2,4-dibromo-17ß-estradiol (diBrE2), and 2,4-dichloro-17ß-estradiol (diClE2) as well as the indirect photolysis of diBrE2 under natural solar irradiance. We found that direct photolysis rate constants increased with halogenation as pKa values decreased and molar absorptivity spectra shifted toward higher wavelengths. Compared to E2, quantum yields were threefold larger for monoBrE2, but 15-32% smaller for the dihalogenated forms. The rate of diBrE2 (pKa ∼ 7.5) photolysis was strongly influenced by pH. At pH 7, diBrE2 degraded on minute time scales due to the large red-shifted molar absorptivity values and greater quantum yields of the phenolate form. Degradation rates were only slightly different in the presence of Suwannee River Humic Acid (5 mg L-1), and quenching experiments pointed to excited triplet state dissolved organic matter (3DOM*) as the dominant reactive intermediate responsible for the indirect photolysis of diBrE2. Overall, our data suggest that halogenated estrogens are particularly susceptible to photochemical degradation at environmentally relevant pH values.

Steroidal estrogen sources in a sewage-impacted coastal ocean.

Estrogens are known to be potent endocrine disrupting chemicals that are commonly found in wastewater effluents at ng L(-1) levels. Yet, we know very little about the distribution and fate of estrogens in coastal oceans that receive wastewater inputs. This study measured a wide range of steroidal estrogens in sewage-impacted seawater using ultra high performance liquid chromatography (UHPLC) tandem mass spectrometry (MS/MS) together with the method of standard addition. In Massachusetts Bay, we find conjugated, free, and halogenated estrogens at concentrations that are consistent with dilution at sites close to the sewage source. At a site 6 miles down current of the sewage source, we observe estrone (E1) concentrations (520 ± 180 pg L(-1)) that are nearly double the nearfield concentrations (320 ± 60 pg L(-1)) despite 9-fold dilution of carbamazepine, which was used as a conservative sewage tracer. Our results suggest that background E1 concentrations in Massachusetts Bay (∼270 ± 50 pg L(-1)) are derived largely from sources unrelated to wastewater effluent such as marine vertebrates.

Measuring free, conjugated, and halogenated estrogens in secondary treated wastewater effluent.

Steroidal estrogens are potent endocrine-disrupting chemicals that enter natural waters through the discharge of treated and raw sewage. Because estrogens are detrimental to aquatic organisms at sub-nanogram per liter concentrations, many studies have measured so-called "free" estrogen concentrations in wastewater effluents, rivers, and lakes. Other forms of estrogens are also of potential concern because conjugated estrogens can be easily converted to potent free estrogens by bacteria in wastewater treatment plants and receiving waters and halogenated estrogens are likely produced during wastewater disinfection. However, to the best of our knowledge, no studies have concurrently characterized free, conjugated, and halogenated estrogens. We have developed a method that is capable of simultaneously quantifying free, conjugated, and halogenated estrogens in treated wastewater effluent, in which detection limits were 0.13-1.3 ng L(-1) (free), 0.11-1.0 ng L(-1) (conjugated), and 0.18-18 ng L(-1) (halogenated). An aqueous phase additive, ammonium fluoride, was used to increase the electrospray (negative mode) ionization efficiency of free and halogenated estrogens by factors of 20 and 2.6, respectively. The method was validated using treated effluent from the greater Boston metropolitan area, where conjugated and halogenated estrogens made up 60-70% of the steroidal estrogen load on a molar basis.

Carbon isotopic (13C and 14C) composition of synthetic estrogens and progestogens.

RATIONALE: Steroids are potent hormones that are found in many environments. Yet, contributions from synthetic and endogenous sources are largely uncharacterized. The goal of this study was to evaluate whether carbon isotopes could be used to distinguish between synthetic and endogenous steroids in wastewater and other environmental matrices. METHODS: Estrogens and progestogens were isolated from oral contraceptive pills using semi-preparative liquid chromatography/diode array detection (LC/DAD). Compound purity was confirmed by gas chromatography/flame ionization detection (GC/FID), gas chromatography/time-of-flight mass spectrometry (GC/TOF-MS) and liquid chromatography/mass spectrometry using negative electrospray ionization (LC/ESI-MS). The (13)C content was determined by gas chromatography/isotope ratio mass spectrometry (GC/IRMS) and (14)C was measured by accelerator mass spectrometry (AMS). RESULTS: Synthetic estrogens and progestogens are (13)C-depleted (δ(13)C(estrogen) = -30.0 ± 0.9 ‰; δ(13)C(progestogen) = -30.3 ± 2.6 ‰) compared with endogenous hormones (δ(13)C ~ -16 to -26 ‰). The (14)C content of the majority of synthetic hormones is consistent with synthesis from C(3) plant-based precursors, amended with 'fossil' carbon in the case of EE(2) and norethindrone acetate. Exceptions are progestogens that contain an ethyl group at carbon position 13 and have entirely 'fossil' (14)C signatures. CONCLUSIONS: Carbon isotope measurements have the potential to distinguish between synthetic and endogenous hormones in the environment. Our results suggest that (13)C could be used to discriminate endogenous from synthetic estrogens in animal waste, wastewater effluent, and natural waters. In contrast, (13)C and (14)C together may prove useful for tracking synthetic progestogens.

Inputs of fossil carbon from wastewater treatment plants to U.S. rivers and oceans.

Every day more than 500 million cubic meters of treated wastewater are discharged into rivers, estuaries, and oceans, an amount slightly less than the average flow of the Danube River. Typically, wastewaters have high organic carbon (OC) concentrations and represent a large fraction of total river flow and a higher fraction of river OC in densely populated watersheds. Here, we report the first direct measurements of radiocarbon (14C) in municipal wastewater treatment plant (WWTP) effluent. The radiocarbon ages of particulate and dissolved organic carbon (POC and DOC) in effluent are old and relatively uniform across a range of WWTPs in New York and Connecticut Wastewater DOC has a mean radiocarbon age of 1630 +/- 500 years B.P. and a mean delta13C of -26.0 +/- 1 per thousand. Mass balance calculations indicate that 25% of wastewater DOC is fossil carbon, which is likely derived from petroleum-based household products such as detergents and pharmaceuticals. These findings warrant reevaluation of the "apparent age" of riverine DOC, the total flux of petroleum carbon to U.S. oceans, and OC source assignments in waters impacted by sewage.