Biotransformation of estrogens in nitrifying activated sludge under aerobic and alternating anoxic/aerobic conditions.

Natural and synthetic estrogens present in municipal wastewater can be biodegraded during treatment, particularly in activated sludge. The objective was to assess the extent of transformation of 17-beta-estradiol (E2) and 17-alpha-ethinylestradiol (EE2) by nitrifying activated sludge and evaluate potential relationships between availability of oxygen, nitrification rate, and estrogen removal. For each batch experiment, two reactors were set up--aerobic and alternating anoxic/aerobic-which were then amended with E2 and EE2 from methanolic stock solutions. The EE2 was persistent under anoxic conditions; under aerobic conditions, the observed level of its removal was 22%. The E2 was readily converted to estrone (El)--faster under aerobic (nitrifying) than anoxic (denitrifying) conditions. During the initial anoxic conditions, a metabolite consistent with 17-alpha-estradiol transiently accumulated and was subsequently removed when the reactor was aerated. Higher removal rates of estrogens were associated with higher nitrification rates, which supports the contention that the nitrifying biomass was responsible for their removal.     

Abiotic transformation of estrogens in synthetic municipal wastewater: An alternative for treatment?

The abiotic transformation of estrogens, including estrone (E1), estradiol (E2), estriol (E3) and ethinylestradiol (EE2), in the presence of model vegetable matter was confirmed in this study. Batch experiments were performed to model the catalytic conversion of E1, E2, E3 and EE2 in synthetic wastewater. Greater than 80% reduction in the parent compounds was achieved for each target chemical after 72 h with the remaining concentration distributed between aqueous and solid phases as follows: 13% and 7% for E1, 10% and 2% for E2, 6% and 2% for E3, and 8% and 3% for EE2, respectively. Testosterone, androstenedione and progesterone were also monitored in this study, and their concentrations were found to be in agreement with initially spiked amount. Data collected under laboratory conditions provided the basis for implementing new abiotic wastewater treatment technologies that use inexpensive materials.     

Photodegradation of the steroid hormones 17beta-estradiol (E2) and 17alpha-ethinylestradiol (EE2) in dilute aqueous solution

The photochemical transformation of natural estrogenic steroid 17beta-estradiol (E2) and the synthetic oral contraceptive 17alpha-ethinylestradiol (EE2) has been studied in dilute non buffered aqueous solution (pH 5.5-6.0) upon monochromatic (254 nm) and polychromatic (lambda>290 nm) irradiation. Upon irradiation at 254 nm, the quantum yields of E2 and EE2 photolysis were similar and evaluated to be 0.067+/-0.007 and 0.062+/-0.007, respectively. Upon polychromatic excitation, and by using phenol as chemical actinometer, the photolysis efficiencies have been determined to be 0.07+/-0.01 and 0.08+/-0.01 for E2 and EE2, respectively. For both estrogens, photodegradation by-products were identified with GC/MS and LC/MS. In a first step, a model compound--5,6,7,8-tetrahydro-2-naphthol (THN)--, which represents the photoactive phenolic group, was used to obtain basic photoproduct structural informations. Numerous primary and secondary products were observed, corresponding to hydroxylated phenolic- or quinone-type compounds.     

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.     

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.     

Biotransformation of ethinylestradiol by microalgae

The capability of biotransformation of 11 microalgae strains was tested on ethinylestradiol (EE). Seven strains were ineffective whilst Selenastrum capricornutum, Scenedesmus quadricauda, Scenedesmus vacuolatus and Ankistrodesmus braunii biotransformed the substrate. EE was converted by S. capricornutum in three products (ethinylestradiol glucoside, 3-β-d-glucopyranosyl-2-hydroxyethinylestradiol, and 3-β-d-glucopyranosyl-6β-hydroxyethinyl estradiol) in 40%, 5%, and 5% yields, respectively. S. quadricauda transformed EE into 17-ethinyl-1,4-estradien-10,17β-diol-3-one (12%) and A. braunii transformed EE into 6--hydroxy-ethinylestradiol (25%).

It is noteworthy that EE is converted in 92% yield in ethinylestradiol glucoside by S. capricornutum when using optimal algal density conditions.     

Development and modeling of a flat plate serpentine reactor for photocatalytic degradation of 17-ethinylestradiol

A flat plate serpentine reactor modified from ultraviolet disinfection pool in municipal wastewater treatment plants was developed for the removal of 17-ethinylestradiol (EE2) for the first time. The photocatalytic degradation performance of EE2 was investigated in this serpentine reactor under different conditions such as inlet concentrations, loaded catalyst concentrations, incident radiations fluxes, and flow velocities. More than 98 % of EE2 was removed under certain conditions within 120 min. An integrated model including a six-flux adsorption–scattering model and a modified flow diffusion model was established to investigate the effect of radiation field and flow velocities, respectively. A satisfactory agreement was observed between the model simulation and experimental results, showing a potential for design and scale-up of photocatalytic reactor for wastewater treatment.     

Factors controlling the biodegradation of 17beta-estradiol, estrone and 17alpha-ethinylestradiol in different natural soils

We conducted a series of laboratory microcosm incubations with [(14)C]-labeled 17beta-estradiol (E2), estrone (E1) and 17alpha-ethinylestradiol (EE2) in 17 different natural soils to characterize hormone mineralization. A significantly higher mineralization was observed for E1 (2.0-37.6%) and E2 (4.2-50.2%) than for EE2 (0.5-2.6%) in all test soils after 21 days. Soil physical or chemical parameters were not related to estrogen mineralization. Although sorption parameters varied greatly for E2 (K(F)=21.9-317.5 mL g(-1)), for E1 (K(F)=46.0-517.5 mL g(-1)) and for EE2 (K(F)=29.9-326.1 mL g(-1)) this apparently did not control estrogen bioavailability since it showed no effects on hormone mineralization. In order to elucidate the controlling factors, experiments with combined additions of radiolabeled estrogens and different substrates were conducted. Additions of ammonium nitrate or alanine to soil samples generally increased EE2 mineralization, thus indicating N-limitation. Additions of glucose induced higher E2 and EE2 degradation in comparison to control samples which is attributed to co-metabolism. Additions of saw dust, catechol or streptomycin influenced the microbial population in the test soils and affected the mineralization of E2 and EE2. Thus, we clearly demonstrate that different microbial communities are responsible for E2 and EE2 degradation in soils. We suggest that EE2 is mineralized by white-rot fungi and E2 by bacteria.     

Identification and behavior of reaction products formed by chlorination of ethynylestradiol

Six products were formed by reaction of ethynylestradiol (EE2) with sodium hypochlorite in buffered solutions. 4-Chloroethynylestradiol (4-ClEE2) and 2,4-dichloroethynylestradiol (2,4-diClEE2) were identified as the two major reaction products, using preparative HPLC, MS, and NMR. When EE2 reacted with chlorine at different pHs (pH 5, 7, and 9) or chlorine concentrations (0.2, 1, 2, and 5 mmol/l, corresponding to molar ratios to EE2, 1, 5, 10, and 25, respectively), the formation of 4-ClEE2 and 2,4-diClEE2 was observed under the above conditions, and the highest yields were 20 and 52 mol%, respectively. EE2 was consumed almost completely within 5 min of chlorination by addition of chlorine of more than 1 mmol/l (molar ratio to EE2, 5). On the other hand, the two products existed in highly chlorinated solutions after 60 min (4ClEE2, 1-6 mol%; 2,4-diClEE2, 3-25 mol%). The estrogenic activities of 4-ClEE2 by estrogen receptor alpha or beta binding assay were similar to those of the parent EE2, and the activities of 2,4-diClEE2 were lower about 10 times.     

Alteration of steroid hormone profile in juvenile turbot (Psetta maxima) as a consequence of short-term exposure to 17alpha-ethynylestradiol

The synthetic estrogen 17alpha-ethynylestradiol (EE2) is among the most potent xenoestrogens found in treated sewage. Adverse effects have been reported in fish exposed to this chemical, such as vitellogenin induction or alteration of gonad structure. In this study, the effects of EE2 on sex steroid profile in juvenile turbot (Psetta maxima) were investigated in a 15-day in vivo experiment. An analytical method was developed for the simultaneous gas chromatography/mass spectrometry determination of 14 steroids in gonads, plasma and bile. The impact of EE2 on the endocrine status of juvenile turbot was sex-dependent, since males were much more sensitive than females. Even at very low concentration (3.5 ng l(-1)), EE2 depressed androgen production as well as plasma levels in males. Androstenedione and 11-ketotestosterone were highly affected by EE2-exposure, since their production was suppressed in EE2-treated males. Furthermore, testis estrogen production and conjugation increased as a consequence of EE2 exposure. Overall, the ratio of androgens to estrogens in EE2-exposed males dropped in testis and plasma and reached values observed in females. These results indicate that juvenile male turbots are susceptible to hormonal imbalance as a consequence of short-term exposure to environmentally relevant EE2 levels.     

Successive identification of biodegradation rates for multiple sequentially reactive contaminants in groundwater.

At the field scale, the biodegradation rate is usually estimated from analytical solutions to single species transport with first-order reactions, using measured data as input. Because many contaminants, e.g., chlorinated solvents, are degraded in a sequential pattern, with degradation products further reacting to produce new species, it is of great interest to quantify the transformation rate of every reaction. The conventional inverse solutions for identifying the transformation rates are limited to single species problems. In the present study, we propose a successive optimization approach to identify the biodegradation rate for each species by using a previously developed analytical solution to multi-species first-order reactive transport using data obtained at the field scale. By specifying a link between analytical solutions to sequentially reactive transport problems and optimization methods and assuming constant transport parameters (velocity, dispersivities, and retardation factors), the first-order transformation rates are optimized successively from parent species to its daughter species.     

Bottom-up approach for the reaction of xenobiotics and their metabolites with model substances for natural organic matter by electrochemistry-mass spectrometry (EC-MS)

Risk assessment of xenobiotics requires a comprehensive understanding of their transformation in the environment. As most of the transformation processes usually involve a redox reaction or a hydrolysis as the first steps of the transformation, we applied an approach that uses an electrochemical cell to investigate model “redox” reactions in aqueous solutions for environmental processes. We investigated the degradation of a variety of xenobiotics from polar to nonpolar and analyzed their degradation products by on-line coupling of electrochemistry with mass spectrometry (EC-MS). Furthermore, we evaluated possible binding reactions with regard to the generation of non-extractable residues with some model substances (catechol, phthalic acid, γ-l-Glutamyl-l-cysteinyl-glycine (GSH) and l-histidine) deduced from a natural organic matter (NOM) structure model and identified possible binding-sites.Whereas typically investigations in soil/water-systems have been applied, we used to our knowledge for the first time a bottom-up approach, starting from the chemicals of interest and different model substances for natural organic matter to evaluate chemical binding mechanisms (or processes) in the EC-MS under redox conditions. Under oxidative conditions, bindings of the xenobiotics with catechol, GSH and histidine were found, but no reactions with the model compound phthalic acid were observed. In general, no chemical binding has yet been found under reductive conditions. In some cases (i.e. benzo[a]anthracene) the oxidation product only underwent a binding reaction, whereas the xenobiotic itself did not undergo any reactions.EC-MS is a promising fast and simple screening method to investigate the environmental behavior of xenobiotics and to evaluate the potential risks of newly synthesized substances.     

Influence of redox potential (Eh) on the availability of arsenic species in soils and soils amended with biosolid

A study was done on the influence of redox potential on the mobility and availability of the various arsenic chemical forms in a Mollisol soil from central Chile amended with biosolid. Arsenic availability was strongly dependent on the applied redox potential. As expected, under reducing conditions (-200 mV vs Hg/Hg(2)Cl(2)) arsenic availability increased significantly, and arsenic was found mainly as arsenite. On the contrary under oxidizing conditions (200 mV vs Hg/Hg(2)Cl(2)) arsenic solubility decreased markedly and was governed by the presence of arsenate. The greatest concentration of organic arsenic species was found under reducing conditions, which would indicate that methylated species may participate in the transformation of arsenate to arsenite. In biosolid-amended soils the concentrations of methylated species increased as a function of time under reducing conditions, which can be attributed to the greater microbial activity resulting from the organic matter supply from the biosolid to soil. In all the systems, a high concentration of As(V) was found under reducing conditions, indicating that the chemical kinetics for the conversion of arsenate to arsenite is slow. Along time, the content of As(V) increased in the control soils, which may be attributed to the possible dissolution of iron oxides and hydroxides under reducing conditions.     

Bottom-up approach for the reaction of xenobiotics and their metabolites with model substances for natural organic matter by electrochemistry–mass spectrometry (EC–MS)

Risk assessment of xenobiotics requires a comprehensive understanding of their transformation in the environment. As most of the transformation processes usually involve a redox reaction or a hydrolysis as the first steps of the transformation, we applied an approach that uses an electrochemical cell to investigate model “redox” reactions in aqueous solutions for environmental processes. We investigated the degradation of a variety of xenobiotics from polar to nonpolar and analyzed their degradation products by on-line coupling of electrochemistry with mass spectrometry (EC–MS). Furthermore, we evaluated possible binding reactions with regard to the generation of non-extractable residues with some model substances (catechol, phthalic acid, γ-l-Glutamyl-l-cysteinyl-glycine (GSH) and l-histidine) deduced from a natural organic matter (NOM) structure model and identified possible binding-sites.Whereas typically investigations in soil/water-systems have been applied, we used to our knowledge for the first time a bottom-up approach, starting from the chemicals of interest and different model substances for natural organic matter to evaluate chemical binding mechanisms (or processes) in the EC–MS under redox conditions. Under oxidative conditions, bindings of the xenobiotics with catechol, GSH and histidine were found, but no reactions with the model compound phthalic acid were observed. In general, no chemical binding has yet been found under reductive conditions. In some cases (i.e. benzo[a]anthracene) the oxidation product only underwent a binding reaction, whereas the xenobiotic itself did not undergo any reactions.EC–MS is a promising fast and simple screening method to investigate the environmental behavior of xenobiotics and to evaluate the potential risks of newly synthesized substances.     

Investigation of the reaction of hexabromocyclododecane with polysulfide and bisulfide in methanol/water solutions

Reactions of hexabromocyclododecane (HBCD), a widely used brominated flame retardant, with reduced sulfur species (e.g., polysulfide and bisulfide) were investigated in well-defined solutions under anoxic conditions. It is likely that reactions of HBCD with reduced sulfur species such as polysulfides and bisulfide present in anoxic subregions of coastal water bodies and sediments could have a significant impact on the fate of HBCD. The second-order reaction rate constant of HBCD with polysulfides in 80% methanol/20% water at 40°C is 2.2 (±0.3)×10(-2) M(-1) s(-1). The second-order reaction rate constant of HBCD with bisulfide is 8.9 (±2.8)×10(-4) M(-1) s(-1) under the same conditions. The formation of two products was observed with either of the two reduced sulfur species. The experiments also indicate that the γ-isomer of HBCD is reacting significantly faster with reduced sulfur species than the α-isomer.     

Histological alternation and vitellogenin induction in adult rare minnow (Gobiocypris rarus) after exposure to ethynylestradiol and nonylphenol

Adult rare minnow (Gobiocypris rarus) were exposed to 0, 1, 5, and 25 ng/l (nominal concentrations) of 17alpha-ethynylestradiol (EE2) and 3, 10, and 30 microg/l (nominal concentrations) of 4-nonylphenol (NP) under flow-through conditions for a period of 28 d. Low mortality was observed at 5 and 25 ng/l EE2 and the growth of fish reduced significantly at 25 ng/l EE2 compared to controls. However, the gonadosomatic indices (GSI) of male fish were significantly higher in 1 ng/l EE(2) treatments and in 10 and 30 microg/l NP treatments (p<0.05). Renal somatic indices (RSI) of male fish in EE2 treatments were significantly higher than those in controls (p<0.05). In contrast, significantly decreased GSI and RSI of female fish could only be observed in 5 and 25 ng/l EE(2) treatments (p<0.05). Hepatosomatic indices (HSI) of male fish were significantly higher in 25 ng/l EE(2) treatments. However, significantly increased of HSI of female fish could only be observed in 1 ng/l EE(2) treatments. Plasma vitellogenin (VTG) induction could be observed in males after exposed to different concentrations of EE2 and NP, and plasma VTG concentrations in females exposed to 5 and 25 ng/l EE(2) were also significantly higher than in controls (p<0.05). At level higher than 5 ng/l EE2 or 30 microg/l NP, hepatic tissue and renal tissue impairment of males could be observed. The pathological male liver was associated with a hypertrophy of hepatocytes and damages to cellar structure and accumulated eosinophilic material. Renal tissue showed different pathological effects which was reflected by accumulated eosinophilic material, hemorrhages within the kidney tubules and hypertrophy of the tubular epithelia. Also at these levels of exposure, feminization of male fish could be noticed and parts of males manifested the testis-ova phenomenon. Ovaries of female rare minnow in 25 ng/l EE2 treatment group were degenerated. Therefore when exposed to EE2 and NP even at environmental observed concentrations, adverse effects could occur in the reproductive system of adult fishes. The observed hepatic tissue and renal tissue impairment should be due to the induction and accumulation of VTG in organs, especially in males.     

Simulation of aerosols and gas-phase species over Europe with the Polyphemus system. Part II: Model sensitivity analysis for 2001

This paper presents a multi-pollutant sensitivity study of an air quality model over Europe with a focus on aerosols. Following the evaluation presented in the companion paper, the aim here is to study the sensitivity of the model to input data, mathematical parameterizations and numerical approximations. To that end, 30 configurations are derived from a reference configuration of the model by changing one input data set, one parameterization or one numerical approximation at a time. Each of these configurations is compared to the same reference simulation over two time periods of the year 2001, one in summer and one in winter. The sensitivity of the model to the different configurations is evaluated through a statistical comparison between the simulation results and through comparisons to available measurements. The species studied are ozone (O₃), nitrogen dioxide (NO₂), sulfur dioxide (SO₂), ammonia (NH₃), coarse and fine aerosol particles (PM_c and PM_2.5), sulfate, nitrate, ammonium, chloride and sodium.For all species, the modeled concentrations are very sensitive to the parameterization used for vertical turbulent diffusion and to the number of vertical levels. For the other configurations considered in this work, the sensitivity of the modeled concentration to configuration choice varies with the species and the period of the year. O₃ is impacted by options related to boundary conditions. PM_c is sensitive to sea-salt related options, to options influencing deposition and to options related to mass transfer between gas and particulate phases. PM_2.5 is sensitive to a larger number of options than PM_c: sea-salt, boundary conditions, heterogeneous reactions, aqueous chemistry and gas/particle mass transfer. NO₂ is strongly influenced by heterogeneous reactions. Nitrate shows the highest variability of all species studied. As with NO₂, nitrate is strongly sensitive to heterogeneous reactions but also to mass transfer, thermodynamic related options, aqueous chemistry and computation of the wet particle diameter. While SO₂ is mostly sensitive to aqueous chemistry, sulfate is also sensitive to boundary conditions and, to a lesser extent, to heterogeneous reactions. As with nitrate, ammonium is largely impacted by the different configuration choices, although the sensitivity is slightly lower than for nitrate. NH₃ is sensitive to aqueous chemistry, mass transfer and heterogeneous reactions. Chloride and sodium are impacted by sea-salt related options, by options influencing deposition and by options concerning the aqueous-phase module.