Quantification of steroid sex hormones using solid-phase extraction followed by liquid chromatography-mass spectrometry.

In this study, the occurrence of trace amounts of natural and synthetic steroid estrogens in the aquatic environment was studied using liquid chromatography coupled with electrospray mass spectrometry, following solid-phase extraction (SPE). The SPE was performed with C18 and NH2 cartridges. The first objective was to develop a reliable method for analyzing steroid estrogens (resulting from human and animal excretions) in different matrices. The method developed was then applied to quantify the occurrence of natural and synthetic hormones (estrone [E1], 17beta-estradiol [betaE2], 17alpha-estradiol [alphaE2], estriol [E3], and 17alpha-ethinylestradiol [EE2]) in environmental samples in surface water and wastewater treatment plant (WWTP) influent and effluent. In the WWTP influents, betaE2, alphaE2, and E3 were identified as ranging up to 72.6 ng/L in WWTP influent and to 16 ng/L in WWTP effluent. Analysis o f surface wa ter sampled upstream from the WWTP revealed the presence of all five estrogens, at levels up to 19.8 ng/L. These concentrations of estrogens pose an issue for large and small communities, because they are higher than the recommended guidelines for estrogen-active compounds and because a lot of communities use surface water as drinking-water sources.     

Steroid estrogens, conjugated estrogens and estrogenic activity in farm dairy shed effluents

Agricultural wastes are a source of steroid estrogens and, if present, conjugated estrogens may add to the estrogen load released to soil and aquatic environments. Dairy shed effluent samples were collected from 18 farms for analysis of steroid estrogens by GC-MS, conjugated estrogens by LC-MS-MS, and estrogenic activity by E-screen in vitro bioassay. 17α-estradiol was found at highest concentrations (median 730 ng l⁻¹), followed by estrone (100 ng l⁻¹) and 17β-estradiol (24 ng l⁻¹). Conjugated estrogens (estrone-3-sulfate, 17α-estradiol-3-sulfate and 17β-estradiol-3,17-disulfate) were measured in most samples (12-320 ng l⁻¹). Median estrogenic activity was 46 ng l⁻¹ 17β-estradiol equivalents. Conjugated estrogens contributed up to 22% of the total estrogen load from dairy farming, demonstrating their significance. Steroid estrogens dominated overall estrogenic activity measured in the samples. Significantly, 17α-estradiol contributed 25% of overall activity, despite potency 2% that of 17β-estradiol, highlighting the importance in environmental risk assessments of this previously neglected compound.     

Oxidation of natural and synthetic hormones by the horseradish peroxidase enzyme in wastewater

Steroid estrogens, including both natural estrogens (e.g., estrone - E1; 17beta-estradiol - E2; and estriol - E3) and synthetic estrogens (e.g., 17alpha-ethinylestradiol - EE2), are known as endocrine-disrupting compounds. The objective of this research was to evaluate the feasibility of the enzymatic oxidation of estrogens and to optimize this process in municipal wastewater contaminated with steroid estrogens using horseradish peroxidase (HRP) and hydrogen peroxide. An initial HRP activity of 0.02 U ml(-1) was sufficient to completely remove EE2 from the synthetic solution, although greater HRP doses (up to 0.06 U ml(-1)) were required to remove E1, E2 and E3. The optimal molar peroxide-to-substrate ratio was determined to be approximately 0.45. Based on the Michaelis-Menten kinetics, the HRP had an increasing reactivity with E1, E3, E2, and EE2, in increasing order. In real activated sludge process effluent, an HRP dose of 8-10 U ml(-1) was required to completely remove all of the studied estrogens, while only 0.032 U ml(-1) of HRP was necessary to treat synthetic water containing the same estrogen concentrations.     

How seasonality affects the flow of estrogens and their conjugates in one of Japan's most populous catchments

A detailed study of the free and conjugated estrogen load discharged by the eight major sewage treatment plants into the Yodo River basin, Japan was carried out. Sampling campaigns were focused on the winter and autumn seasons from 2005 to 2008 and the free estrogens estrone(E1), 17β-estradiol(E2), estriol(E3), 17α-ethynylestradiol(EE2) as well as their conjugated (sulfate and glucuronide) forms. For both sewage effluent and river water E2 and E1 concentrations were greatest during the winter period (December-March). This coincides with the period of lowest rainfall and lowest temperatures in Japan. E1 was the dominant estrogenic component in effluent (means of 10-50 ng/L) followed by E2 (means of 0.5-3 ng/L). The estrogen sulfate conjugates were found intermittently in the 0.5-1.7 ng/L concentration range in the sewage effluents. The greatest estrogen exposure was found to be in the Katsura River tributary which exceeded 1 ng/L E2-equivalents during the winter period.     

Estrogens and synthetic androgens in manure slurry from trenbolone acetate/estradiol implanted cattle and in waste-receiving lagoons used for irrigation

The increasing size of concentrated animal feeding operations has led to a concomitant increase in the land-application of manure, which has spawned research on the concentrations and environmental risk assessment of natural and synthetic hormones in animal manures. 17β-Trenbolone acetate (TBA) is widely used in the United States for improving daily gains in beef cattle and is often administered in combination with 17β-estradiol (17β-E2). Trenbolone (TB) and E2 isomers and their metabolites were quantified in manure collection pits and lagoon effluent from beef cattle implanted with the commercial anabolic preparation Ravoler-S (containing 140 mg 17β-trenbolone acetate and 28 mg 17β-E2). Manure pit and lagoon effluent samples were collected weekly for 9 weeks post implanting and analyzed using reverse-phase liquid chromatography tandem mass spectrometry. 17α-TB was the most abundant androgen with the highest concentration observed 2 weeks post implant. 17β-TB and trendione peaked at the end of week 2 and 4, respectively. For the estrogens, the highest concentrations for estrone (E1), estriol (E3), and 17α-E2 were observed after week 4, 6, and 8, respectively. 17β-E2 concentrations were the lowest of the estrogens and erratic over time. In lagoon water, which is used for irrigation, 17α-TB and E1 had the highest detected hormone concentrations (1.53 and 1.72 μg L⁻¹, respectively). Assuming a 1-2 order dilution during transport to surface water, these hormone levels could lead to concentrations in receiving waters that exceed some of the lowest observable effect levels (LOELs) reported for hormones (e.g., 0.01-0.03 μg L⁻¹).     

Occurrence of free estrogens,conjugated estrogens,and bisphenol A in fresh livestock excreta and their removal by composting in North China

An efficient pretreatment and analytical method was developed to investigate the occurrence and fate of four free estrogens (estrone (E1), 17β-estradiol (17β-E2), estriol (E3), and 17α-ethinylestradiol (EE2)), four conjugated estrogens (estrone-3-sulfate sodium salt (E1-3S), 17β-estradiol-3-sulfate sodium salt (E2-3S), estrone-3-glucuronide sodium salt (E1-3G), and 17β-estradiol-3-glucuronide sodium salt (E2-3G)), and bisphenol A (BPA) in three livestock farms raising beef cattle, cows, sheep, swine, and chickens in Qi County, which is located in North China. The results demonstrated that one cow and one beef cattle excreted 956.25–1,270.41 and 244.38–319.99 μg/day of total (free and conjugated) estrogen, respectively, primarily through feces (greater than 91 %), while swine excreted 260.09–289.99 μg/day of estrogens, primarily through urine (98–99 %). The total estrogen excreted in sheep and broiler chicken feces was calculated to be 21.64–28.67 and 4.62–5.40 μg/day, respectively. It was determined that conjugated estrogens contributed to 21.1–21.9 % of the total estrogen excreted in cow feces and more than 98 % of the total estrogen excreted in swine urine. After composting, the concentration of total estrogen decreased by 18.7–59.6 %; however, increased levels of BPA were measured. In treated compost samples, estrogens were detected at concentrations up to 74.0 ng/g, which indicates a potential risk of estrogens entering the surrounding environment.     

Estrogens and synthetic androgens in manure slurry from trenbolone acetate/estradiol implanted cattle and in waste-receiving lagoons used for irrigation

The increasing size of concentrated animal feeding operations has led to a concomitant increase in the land-application of manure, which has spawned research on the concentrations and environmental risk assessment of natural and synthetic hormones in animal manures. 17β-Trenbolone acetate (TBA) is widely used in the United States for improving daily gains in beef cattle and is often administered in combination with 17β-estradiol (17β-E2). Trenbolone (TB) and E2 isomers and their metabolites were quantified in manure collection pits and lagoon effluent from beef cattle implanted with the commercial anabolic preparation Ravoler-S (containing 140 mg 17β-trenbolone acetate and 28 mg 17β-E2). Manure pit and lagoon effluent samples were collected weekly for 9 weeks post implanting and analyzed using reverse-phase liquid chromatography tandem mass spectrometry. 17α-TB was the most abundant androgen with the highest concentration observed 2 weeks post implant. 17β-TB and trendione peaked at the end of week 2 and 4, respectively. For the estrogens, the highest concentrations for estrone (E1), estriol (E3), and 17α-E2 were observed after week 4, 6, and 8, respectively. 17β-E2 concentrations were the lowest of the estrogens and erratic over time. In lagoon water, which is used for irrigation, 17α-TB and E1 had the highest detected hormone concentrations (1.53 and 1.72 μg L⁻¹, respectively). Assuming a 1–2 order dilution during transport to surface water, these hormone levels could lead to concentrations in receiving waters that exceed some of the lowest observable effect levels (LOELs) reported for hormones (e.g., 0.01–0.03 μg L⁻¹).     

Steroid estrogens in ocean sediments

This paper gives results from a study measuring the abundance of steroid hormones in ocean sediments in the proximity of a deep ocean sewage outfall. The outfall is discharge point for an enhanced primary sewage treatment plant and sediment samples were taken adjacent and 7 km from the outfall. All samples contained steroid estrogens at nanogram per gram levels with higher concentrations at the 7 km sampling site. The concentration of estrone ranged from (0.16-1.17 ng/g), 17beta-estradiol (0.22-2.48 ng/g) and the synthetic 17alpha-ethinylestradiol (<0.05-0.5 ng/g). The values detected correspond with estimates based on the proportion of estrogens sorbed to particles in the effluent and the expected proportion of particles originating from sewage in the ocean sediments. The results suggest that estrogens associated with the particulate fraction aggregate on contact with high ionic strength seawater and settle to the seafloor after discharge through deep ocean outfalls.     

Sorption and desorption of three endocrine disrupters in soils

Sorption of the estrogens estrone (E1), 17beta-estradiol (E2) and 17alpha-ethynylestradiol (EE2) on four soils was examined using batch equilibrium experiments with initial estrogen concentrations ranging from 10 to 1000 ng mL-1. At all concentrations, >85% of the three estrogens sorbed rapidly to a sandy soil. E1 sorbed more strongly to soil than E2 or EE2. Partial oxidation of E2 to E1 was observed in the presence of soils. Autoclaving was more effective at reducing this conversion than inhibition with sodium azide or mercuric chloride, and had little effect on sorption, relative to the chemical microbial inhibitors. Sorption of EE2 was greater for fine-textured than coarse-textured soils, but greater than 90% of EE2 sorbed onto all four soils. The greatest degree of desorption of estrogens from the sandy soil occurred with the lowest initial concentration of 10 ng mL-1 and reached levels >or=80% for E1 and E2. Desorption of EE2 was greater in coarser textured soils than finer-textured soils. Again, relative desorption from all soils was greatest with low initial concentrations. Therefore, at environmentally relevant concentrations, estrogens quickly sorb to soils, and soils have a large capacity to bind estrogens, but these endocrine-disrupting compounds can become easily desorbed and released into the aqueous phase.     

Removal of estrogenic activity of natural and synthetic hormones from a municipal wastewater: efficiency of horseradish peroxidase and laccase from Trametes versicolor

Some researches studied the removal of steroid estrogens by enzymatic treatment, however none verified the residual estrogenicity after the enzymatic treatment at environmental conditions. In this study, the residual estrogenic activities of the key natural and synthetic steroid estrogens were investigated following enzymatic treatment with horseradish peroxidase (HRP) and laccase from Trametes versicolor. Synthetic water and municipal wastewater containing environmental concentrations of estrone, 17beta-estradiol, estriol, and 17alpha-ethinylestradiol were treated. Liquid chromatography-mass spectrometry analysis demonstrated that the studied steroid estrogens were completely oxidized in the wastewater reaction mixture after a 1-h treatment with either HRP (8-10 U ml(-1)) or laccase (20 U ml(-1)). Using the recombinant yeast assay, it was also confirmed that both enzymatic treatments were very efficient in removing the estrogenic activity of the studied steroid estrogens. The laccase-catalyzed process seemed to present great advantages over the HRP-catalyzed system for up-scale applications for the treatment of municipal wastewater.     

Sorption and Desorption of Three Endocrine Disrupters in Soils

Sorption of the estrogens estrone (E1), 17β-estradiol (E2) and 17α-ethynylestradiol (EE2) on four soils was examined using batch equilibrium experiments with initial estrogen concentrations ranging from 10 to 1000 ng mL^?1. At all concentrations, >85% of the three estrogens sorbed rapidly to a sandy soil. E1 sorbed more strongly to soil than E2 or EE2. Partial oxidation of E2 to E1 was observed in the presence of soils. Autoclaving was more effective at reducing this conversion than inhibition with sodium azide or mercuric chloride, and had little effect on sorption, relative to the chemical microbial inhibitors. Sorption of EE2 was greater for fine-textured than coarse-textured soils, but greater than 90% of EE2 sorbed onto all four soils. The greatest degree of desorption of estrogens from the sandy soil occurred with the lowest initial concentration of 10 ng mL^?1 and reached levels ≥80% for E1 and E2. Desorption of EE2 was greater in coarser textured soils than finer-textured soils. Again, relative desorption from all soils was greatest with low initial concentrations. Therefore, at environmentally relevant concentrations, estrogens quickly sorb to soils, and soils have a large capacity to bind estrogens, but these endocrine-disrupting compounds can become easily desorbed and released into the aqueous phase.     

Identification of new ozonation disinfection byproducts of 17β-estradiol and estrone in water

Estrogens are a class of micro-pollutants found in water at low concentrations (in the ng L⁻¹ range), but often sufficient to exert estrogenic effects due to their high estrogenic potency. Disinfection of waters containing estrogens through oxidative processes has been shown to lead to the formation of disinfection byproducts, which may also be estrogenic. The present work investigates the formation of disinfection byproducts of 17β-estradiol (E2) and estrone (E1) in the treatment of water with ozone. Experiments have been carried out at two different concentrations of the estrogens in ground water (100 ng L⁻¹ and 100 μg L⁻¹) and at varying ozone dosages (0-30 mg L⁻¹). Detection of the estrogens and their disinfection byproducts in the water samples has been performed by means of ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) with a triple quadrupole (QqQ) and a quadrupole-time of flight (QqTOF) instrument. Both E2 and E1 have been found to form two main byproducts, with molecular mass (MM) 288 and 278 in the case of E2, and 286 and 276 in the case of E1, following presumably the same reaction pathways. The E2 byproduct with MM 288 has been identified as 10epsilon-17beta-dihydroxy-1,4-estradieno-3-one (DEO), in agreement with previously published results. The molecular structures and the formation pathways of the other three newly identified byproducts have been suggested. These byproducts have been found to be formed at both high and low concentrations of the estrogens and to be persistent even after application of high ozone dosages.     

Environmental transport of endogenous dairy manure estrogens

Although estrogens originating from dairy manure applied to agricultural soils as a fertilizer can potentially contaminate surface water and groundwater, the variables that control transport are poorly understood. Our objective was to assess the potential for off-site movement of endogenous dairy cattle estrogens when manure is applied on fields at agronomically relevant fertilization rates. Estrone (E1), 17α-estradiol (α-E2), and 17β-estradiol (β-E2) were used in laboratory sorption, desorption, and transformation incubations with both manure and an agriculturally relevant soil. Sorption on manure containing 44% organic carbon exceeded sorption on soil containing 0.8% organic carbon by 20 to 150 times, following the pattern of β-E2 > α-E2 > E1. Approximately 20% of E1 and 17% of α-E2 were desorbed from manure, whereas only about 4% of β-E2 was desorbed. Thirty to seventy percent of α-E2 and β-E2 were converted to E1 in soil and manure, making it imperative that transformation reactions be considered when predicting transport and potential biological effects in the environment. Overall results indicate that high organic carbon concentrations and relatively low amounts of desorption inhibit the potential for off-site transport of endogenous dairy manure estrogens.     

Assessment of the importance of sorption for steroid estrogens removal during activated sludge treatment

Distribution coefficients (K(d)) between water and activated sludge particles (f(oc)=27.7+/-0.1%) were measured for the steroid estrogens (SE), estrone (E1), 17beta-estradiol (E2) and 17alpha-ethinylestradiol (EE2) in batch experiments. Experimental concentration levels ranged from environmentally realistic low ng/l to the high microg/l. In this range K(d)s were independent of their water concentration. The experimentally obtained K(d)s (with 95% confidence intervals) were 402+/-126 l/kg, 476+/-192 l/kg and 584+/-136 l/kg for E1, E2 and EE2, respectively. K(d)s were used to estimate the fraction of the total SE concentration that is expected to be sorbed in the activated sludge treatment tanks of a typical STP assuming equilibrium conditions. Assuming a suspended solids concentration of 4 g/l dissolved solids (ds), it was estimated that 61+/-9%, 66+/-13% and 70+/-6% of the total concentration of E1, E2 and EE2, respectively, would be sorbed during activated sludge treatment. The fraction of the SEs that was expected to be sorbed to suspended sludge particles in the effluents from a typical Danish STP was estimated to be only 0.20+/-0.06%, 0.24+/-0.10% and 0.29+/-0.07% of the total concentration of E1, E2 and EE2, respectively, at a suspended solids concentration of 5 mg/lds. For a typical STP the removal of steroid estrogens with excess sludge was estimated to be only 1.5-1.8% of the total loading if equilibrium conditions exists. Sorption is therefore not important for the fate of SEs in STPs compared to biodegradation.     

Europe-wide survey of estrogenicity in wastewater treatment plant effluents: the need for the effect-based monitoring

A pan-European monitoring campaign of the wastewater treatment plant (WWTP) effluents was conducted to obtain a concise picture on a broad range of pollutants including estrogenic compounds. Snapshot samples from 75 WWTP effluents were collected and analysed for concentrations of 150 polar organic and 20 inorganic compounds as well as estrogenicity using the MVLN reporter gene assay. The effect-based assessment determined estrogenicity in 27 of 75 samples tested with the concentrations ranging from 0.53 to 17.9 ng/L of 17-beta-estradiol equivalents (EEQ). Approximately one third of municipal WWTP effluents contained EEQ greater than 0.5 ng/L EEQ, which confirmed the importance of cities as the major contamination source. Beside municipal WWTPs, some treated industrial wastewaters also exhibited detectable EEQ, indicating the importance to investigate phytoestrogens released from plant processing factories. No steroid estrogens were detected in any of the samples by instrumental methods above their limits of quantification of 10 ng/L, and none of the other analysed classes of chemicals showed correlation with detected EEQs. The study demonstrates the need of effect-based monitoring to assess certain classes of contaminants such as estrogens, which are known to occur at low concentrations being of serious toxicological concern for aquatic biota.     

Effects of waterborne exposure to 4-nonylphenol on plasma sex steroid and vitellogenin concentrations in sexually mature male carp (Cyprinus carpio)

4-Nonylphenol (NP) has been shown to elicit estrogenic responses both in vivo and in vitro. The mechanism by which NP exerts estrogenic and other endocrine-modulating effects in vivo remains unclear, however. The goal of this study was to evaluate the ability of NP to elicit estrogenic responses through indirect mechanisms of action involving the modulation of endogenous steroid hormone concentrations. Sexually mature male common carp (Cyprinus carpio) were exposed to aqueous NP concentrations ranging from <0.05 to 5.4 microg NP/l for 28-31 d. Approximately 0.5-3.5 ppm of NP was detected in pooled plasma samples or tissue samples from the carp studied. NP exposure did not significantly increase plasma concentrations of 17beta-estradiol (E2), testosterone (T) or vitellogenin (VTG). Excluding outliers, plasma E2 concentrations ranged from <175 to 700 pg E2/ml. T concentrations ranged from 940 to 24,700 pg T/ml plasma. The greatest VTG concentration detected was 52 microg/ml. One-third of the plasma samples tested contained <1 microg VTG/ml. Overall, the results of this study did not support the hypothesis that exposure to waterborne NP can modulate concentrations of steroid hormones in the plasma of sexually mature male carp. The results did, however, raise a number of questions regarding the utility of estradiol equivalent (EEQ) estimates as a means of predicting in vivo effects of estrogenic substances. Furthermore, they provide information regarding the concentrations and variability of E2, T, and VTG in the plasma of sexually mature male carp, which may aid in design and interpretation of future studies.     

Spatial distribution and vertical profile of polybrominated diphenyl ethers and hexabromocyclododecanes in sediment core from Tokyo Bay, Japan

Brominated flame retardants (BFRs), hexabromocyclododecanes (HBCDs) and polybrominated diethyl ethers (PBDEs) were detected in three sediment cores and six surface sediments of Tokyo Bay, Japan. HBCDs were detected for the first time in this region with concentrations ranging from 0.056 to 2.3 ng/g dry wt, implying their widespread contamination, even though their concentrations were lower than summation operatorPBDEs (di- to nona-BDEs) and BDE-209. Levels of these compounds were higher near to the highly populated industrial area of the bay implicating industrial and human activities as sources of these compounds. Summation operatorPBDEs increased in the sediment layer up to the mid-1990s and decreased afterwards, whereas BDE-209 showed an increasing trend until now, following the usage of different commercial BDE mixtures. HBCDs first appeared in the mid-1970s and increased until today. The annual surficial flux of HBCDs (0.62-2.4 ng/cm2/yr) is equal to summation operatorPBDEs (0.95-2.6 ng/cm2/yr) but lower than that of BDE-209 (17-58 ng/cm2/yr).     

Spatial distribution and loading amounts of particle sorbed and dissolved perfluorinated compounds in the basin of Tokyo Bay

In this study, we analyzed over 30 types of PFCs, including precursors in both the dissolved phase and particle solid phase, in 50 samples of river water collected from throughout the Tokyo Bay basin. PFCs were detected in suspended solids (SSs) at levels ranging from <0.003-4.4 ng L(-1) (0.11-2470 ng g(-1) dry weight). The concentrations of PFCs in the SS were one to two order(s) of magnitude lower than those of PFCs in the dissolved phase. Relatively high levels of PFCs (total of 35 PFCs) in SS were observed in urbanized areas. The concentration of PFCAs, including PFOA and PFNA, were significantly correlated with the geographic index as artificial area (R(2) of the linear regression curve in a double logarithmic plot: 0.09-0.55). Conversely, PFOS and FOSA were significantly correlated with the arterial traffic area (R(2) in a double logarithmic plot: 0.29-0.55). Those spatial trends were similar to the trends in dissolved PFCs. We estimated the loading amount of PFCs into Tokyo Bay from six main rivers and found that more than 90% of the total PFCs reached Tokyo Bay in the dissolved phase. However, 40.0-83.5% of the long chain PFCAs (C12-C15), were transported as particle sorbed PFCs. Rain runoff events might increase the loading amount of PFCs in SS. Overall, the results presented herein indicate that greater attention should be given to PFCs, especially for longer chain PFCs in SS in addition to dissolved PFCs.     

17 β-estradiol and 17 α-ethinylestradiol mineralization in sewage sludge and biosolids

Natural steroid estrogens (e.g., 17 β-estradiol, E2), synthetic steroid estrogens (e.g., 17 α-ethinylestradiol, EE2) and pharmaceutical antibiotics (e.g., ciprofloxacin) are chemicals detected in biosolids and sewage sludges because they partition into the solids fraction during the wastewater treatment process. This research utilized a three-way factorial design (six media × two estrogens × three antibiotic treatments) to quantify cumulative E2 and EE2 mineralization over 133 d (MAX) in a range of sewage sludge and biosolid samples in the presence (4 and 40 mg kg^?1) and absence of ciprofloxacin. The same three-way factorial design was utilized to quantify the impact of the six media, E2 or EE2, and ciprofloxacin on cumulative soil respiration over 133 d (RESP). Minimal ciprofloxacin mineralization was observed (<0.05% over 133 d), but despite its persistence, ciprofloxacin had no significant effect on MAX of E2 or EE2, and, in general, no significant effect on RESP. MAX ranged from 38.38% to 48.44% for E2 but from only 0.72% to 24.27% for EE2 although RESP was relatively similar, ranging from 101.00 to 866.54 mg CO₂ in the presence of E2 and from 69.55 to 893.95 mg CO₂ in the presence of EE2. The sorption-limited bioavailability of EE2, which is inherently resistant to biodegradation due to chemical structure, as MAX and Freundlich sorption coefficients (K_f) were negatively correlated. As such, the K_f values of EE2 were largest in composted biosolids in which EE2 was particularly resistant to microbial degradation as the MAX of EE2 was <3%. In contrast, the MAX of E2 showed a positive association with the K_f values of E2 because some steps in the E2 transformation process have been found to occur in the sorbed phase. The MAX of E2 was significantly greater in the biosolid and composted biosolid media than in any other media, whereas the MAX of E2 decreased in the following order: secondary sewage sludge > primary sewage sludge > biosolid = composted biosolid. This suggests that sewage sludges in municipal lagoons and pre-treatment holding lagoons are a more favorable media for mineralization of EE2, whereas biosolids in post-treatment storage lagoons are a more favorable media for the mineralization of E2. The presence of ciprofloxacin will have no impact on the potential E2 or EE2 mineralization rates in these cases.     

Biochemical and biological assays of endocrine disrupting compounds in various manure matrices

Wastes generated by animal agriculture have complex matrices and present a significant challenge for achieving accurate measurements of estrogens. The objective of this study was to compare two different extraction methods and two different biochemical and biological estrogen assays on two dairy manure matrices. The two extraction methods studied in this research were a two-step base-toluene extraction and a one-step cyclohexane extraction. The two assays assessed were an enzyme-linked immunosorbent assay (ELISA) and a yeast estrogen screen (YES) bioassay. Estrogenic activity was measured directly as 17beta-estradiol (E2) through the ELISA method or as E2 equivalents (E2-eq) through the YES bioassay. Both extraction methods yielded reasonable recoveries from distilled water in the absence of matrix interferences. In manure samples, ELISA resulted in recoveries slightly higher than 100% in two types of dairy manures, but YES recoveries varied. The YES bioassay detected much higher estrogenic activities in dairy manure compared to the E2 concentrations measured by ELISA. The base-solvent extraction yielded higher E2 concentrations in dairy manure compared to the one-step cyclohexane extraction. These results suggest that manure matrices vary sufficiently that extraction methods must be optimized for specific assays utilized to quantify estrogens in manures.