Source: NORTH DAKOTA STATE UNIV submitted to
PERSISTENCE AND MOBILITY OF ESTROGENS IN THE ENVIRONMENT
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
0219284
Grant No.
2010-65102-20400
Project No.
ND05903
Proposal No.
2009-02432
Multistate No.
(N/A)
Program Code
94540
Project Start Date
Dec 1, 2009
Project End Date
Nov 30, 2013
Grant Year
2010
Project Director
Casey, F. X.
Recipient Organization
NORTH DAKOTA STATE UNIV
(N/A)
FARGO,ND 58105
Performing Department
School of Natural Resource Sciences
Non Technical Summary
Hormone detections have been associated with manure management practices; whish is of particular concern to the animal agriculture industry. The primary estrogen, E2, is potent to sensitive organisms at part per trillion concentrations. Laboratory studies have determined E2 to be short-lived in soils; nonetheless, it is frequently detected in the environment. This apparent disparity between laboratory and field studies could be the result of unknown conjugate and colloidal transport processes in complex field systems that are not usually observed in the laboratory. Small organic matter particles, or colloids, can potentially attach to hormones protecting them from microbial degradation or helping them move freely in the soil. Also, as the animal eliminates hormones from their body, they are predominantly in the conjugated form. These conjugated forms of the hormone are potentially much more mobile then the deconjugatged forms, because they can readily dissolve in water compared to the deconjugated forms. This project will evaluate how hormone conjugates and colloids may contribute to the greater than expected mobility and persistence of hormones in the environment. Laboratory and field experiments will be used to achieve the overall goal of this project. The results of this project can provide improved manure management strategies that reduce releases of reproductive hormones from manure. Results will be presented at national scientific meetings and submitted for publication in scientific journals. Also, an aim for this project is to develop practical solutions/applications evolving from our basic research. North Dakota State University is the state's Land Grant institution that has an Extension mission to provide outreach education to make basic research applicable. In the past, the PDs have participated in several Extension/Outreach talks to farmers, consultants, etc. to educate them about hormones and animal manures. The PDs feel that it is important to actively participate in this Land Grant mission through extension/outreach activities and demonstrations, such as on the Discovery Farm.
Animal Health Component
(N/A)
Research Effort Categories
Basic
80%
Applied
20%
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
1120210205050%
1330210205050%
Knowledge Area
133 - Pollution Prevention and Mitigation; 112 - Watershed Protection and Management;

Subject Of Investigation
0210 - Water resources;

Field Of Science
2050 - Hydrology;
Goals / Objectives
The overall goal of this project is to identify likely mechanisms that cause the greater than expected mobility and persistence of reproductive hormones in the environment. This information is necessary to develop manure handling best management practices, and developing predictive models. Three experimental objectives were formed to achieve the project goal: Objective 1 determine the sorption characteristics of 17beta-estradiol (E2) and its glucuronide (E2G) and sulfate (E2S) conjugates to soil in the presence and absence of dissolved organic carbon (DOC) and colloidal organic carbon (COC) originating from animal manure. This first objective will test the hypothesis that if the sorption of estrogens are related to the presence of liquid manure (slurry)-derived DOC and COC, then estrogen in the presence of high amounts of manure DOC or COC will increase their mobility. Laboratory experiments that test this first hypothesis will be an output, and will answer the question, "Does DOC and COC from lagoon slurry enhance the mobility of estrogens in mineral soil and thus increase the potential for off-site movement" An additional hypothesis that this first objective will test is, "if conjugated hormones are more soluble than their deconjugated forms, then they will not bind strongly to soil be more mobile. Outputs testing this second hypothesis will be laboratory experiments that answer the question, "Do estrogen conjugates promote the apparent mobility or persistence of deconjugated estrogens in soil" Objective 2 will be to measure the degradation of E2, E2G, and E2S in soil in the presence of manure slurry-derived DOC and COC. The hypothesis tested of this second objective is, "if estrogenic hormones are bound to DOC or COC and not easily accessible to microbial processes, then estrogens in the presence of high amounts of DOC or COC will increase their persistence in the environment compared to free aqueous estrogens." Outcomes from this second objective will be laboratory experiments that answer the question, "Does DOC and COC fractions of manure slurry offer estrogenic hormones protection from microbial degradation and increase their longevities in the soil or manure storage facilities" Objective 3 will determine how land application strategies for manure slurry influence the fate and transport of estrogens through soil to tile drainage and how this relates to manure slurry contents of DOC, COC, and estrogen conjugates. The hypothesis that this objective will test is, if the mobility of estrogens in soil is related to hormone conjugates and the presence of DOC or COC and the retention time in the near-surface horizons, then the detections of estrogens will be related to manure application on tile drainage fields, manure-borne DOC and COC, and increased drainage rates. The outcome of this objective will be a field study, where manure is applied to the soil and E2, E2S, and E2G are measured in runoff and tile drainage and correlated to DOC and COC contents. The research question that will be answered is, "Do estrogens and their conjugates from manure slurries applied to fields move with gravity water into tile drains and into surface waters"
Project Methods
The fist and second objective will be achieved using laboratory experiments in which the sorption and persistence, respectively, of E2, E2S, and E2G to soil is observed in the presence of animal manure slurry. Radioisotope methods will be used with batch sorption experiments in soils that have been sterilized. Various concentrations of aqueous dissolved estrogens and manure slurry will be combined with soil, and the sorption kinetics and sorption equilibrium processes will be identified. Also, incubation experiments will be done under sterile/natural and aerobic/anaerobic conditions to identify the degradation pathways. Radiolabel forms of E2G and E2S will first need to be synthesized for the batch and incubation experiments. The third objective will be achieved with a field experiment in which existing subsurface tile-drainage lines are accessed and sampled. Also, surface runoff from the field will be sampled. Swine (Sus scrofa domesticus) manure slurry will be injected or surface applied to the field and water samples from the subsurface tile drain, and run off will be taken, preserved with formaldehyde, and analyzed with liquid chromatography equipped with tandem mass spectrometers. Results from the field study will be correlated with the manure DOC and COC, and compared to laboratory experiments. Results will be presented at national scientific meetings to further the scientific research in this area. Also, an aim for this project is to develop practical solutions/applications evolving from our basic research. North Dakota State University is the state's Land Grant institution that has an Extension mission to provide outreach education to make basic research applicable. In the past, the PDs have participated in several Extension/Outreach talks to farmers, consultants, etc. to educate them about hormones and animal manures. The PDs feel that it is important to actively participate in this Land Grant mission through extension/outreach activities and demonstrations, such as on the Discovery Farm.

Progress 12/01/09 to 11/30/13

Outputs
Target Audience: Target audiences are soil scientist, environmental scientist, water quality specialist, environmental engineers, remediation specialist, waste water managers, and potential policy makers. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? Presented at conferences and publication in peer-reviewed journals. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? 1) The fate of estradiol (E2) in anaerobic lagoons was investigated use benchtop scaled-down anaerobic digestors. It was found that estradiol converted to the less biologically potent estrone (E1) in less than 4 days and reduced the total potent estrogenicity by 50 fold. 2) The fate of the estradiol conjugate 17sulfate-17beta-estradiol (E2-17S) was investigated as a natural process that might explain the greater than expected detections of E2 in the environment. Soil batch studies indicated that E2-17S persisted much greater than E2, where E2-17S levels remained in the aqueous phase for the duration of the 14 day experiment while E2 only persists for a few hours. Additionally, the E2-17S did not bind as strongly to soil as the parent E2. These results indicate that E2-17S may contribute to the parent E2 detected in the environment and facilitate its apparent mobility. 3) Analytical proceedures were developed and reported that greatly help to close the experimental mass balance of highly labile estrogenic compounds. These analytical proceedures greatly increase the ability to understand the fate and transport processes of these compounds. 4) Analysis of samples from large undisturbed soil lysimeters where cow urine was applied provided insight into the fate and transport of natural estrogens, E2, E1, and their sulfate and glucuronide conjugates. Frequently detected E2 conjugates indicated that they facilitated the transport of E2 and E1.

Publications

  • Type: Journal Articles Status: Accepted Year Published: 2014 Citation: Heldur Hakk, Lawrence Sikora, Francis X. M. Casey, Gerald L. Larsen. Fate of 17b-Estradiol in Anaerobic Lagoon Digesters. J. Environmental Quality.
  • Type: Journal Articles Status: Published Year Published: 2013 Citation: X Bai, FXM Casey, H Hakk, TM DeSutter, PG Oduor, E Khan. 2013. Dissipation and transformation of 17b-estradiol-17-sulfate in soilwater systems. Journal of hazardous materials 260, 733-739
  • Type: Journal Articles Status: Accepted Year Published: 2013 Citation: SL Shrestha, FXM Casey, H Hakk, G Padmanabhan. 2013. Radioassay-Based Approach to Investigate Fate and Transformation of Conjugated and Free Estrogens in an Agricultural Soil. Environmental engineering science 30 (2), 89-96
  • Type: Journal Articles Status: Accepted Year Published: 2014 Citation: X Bai, FXM Casey, H. Hakk, T DeSutter, P Oduor, and E. Khan 2014. Sorption and Metabolism of 17beta-Estradiol-17-Sulfate in Sterilized Soil-Water Systems. Chemosphere
  • Type: Theses/Dissertations Status: Accepted Year Published: 2014 Citation: Xuelian Bai. 2014. Fate and transport of estrogen sulfate conjugate 17beta-estradiol-17-sulfate in soil water systems. Ph.D. Dissertation. North Dakota Sate University
  • Type: Theses/Dissertations Status: Accepted Year Published: 2014 Citation: Katrin Chambers. 2014. Bioavailability of 17beta-estradiol associated with soil and manure colloidal and dissolved fractions. M.S. Thesis. North Dakota State University
  • Type: Conference Papers and Presentations Status: Published Year Published: 2013 Citation: Chambers, K., F.X.M. Casey, H. Hakk, T.M. DeSutter, N. Shappell. 2013. Association of estradiol with manure and soil colloidal and dissolved fractions and potential bioactivity. Presented at the International Meetings of the ASA-CSSA-SSSA. Tampa, FL. November 3-6.


Progress 12/01/11 to 11/30/12

Outputs
OUTPUTS: In 2012, experiments were conducted to determine how dissolve organic carbon (DOC) and colloidal organic carbon (COC) derived from soil and manure affect the fate and transport of the estrogen, 17beta-estradiol (E2). This compound, E2, can disrupt the endocrine systems of aquatic organisms at very low concentrations (ppt levels). Radiolabelled (14C)-E2 was filtered with pure water and different combinations of soil and manure derived COC/DOC solutions. These experiments were designed to see whether the 14C-E2 preferentially associated with the different organic carbon fractions (i.e. COC or DOC), and whether these organic carbon fractions affected E2 persistence and binding to soil. Additionally, models were developed to help identify and potentially predict the fate and transport of the sulfate and glucuronide conjugates of E2. Estradiol is very insoluble and in order for an animal to excrete it from their body, it must first be made soluble by attaching a sulfate or glucuronide to the E2 molecule. The attachment of the sulfate (E2-S) or glucuronide (E2-G) to the E2 molecule is called conjugation. Between 60% and 90% of all estrogens excreted from farm animals are in the forms of conjugates. Experimental observations from soil batch experiments were used to develop fate and transport models of E2-S and E2-G and their metabolites. The model was solved numerically and applied inversely to the experimental observations using a global optimization method to quantify the sorption and transformation parameters of E2-S and E2-G and their metabolites. Also, a new laboratory analytical procedure was developed that only requires liquid chromatography and liquid scintillation to accurately quantify and qualify 14C-E2 its metabolites in laboratory experiments. Finally, field observations have continued on a field where subsurface agriculture drainage is installed and where manure has been applied. Other field observations of the E2 in the floodwaters were made. A total of four graduate students (2 MS and 2 PhDs) were involved in conducting the field and laboratory experiments, and developing the models. Experimental observation and model results were presented at two national scientific conferences. Also, the field and laboratory experiments, and the laboratory methods were published in three peer-review journals. PARTICIPANTS: Nothing significant to report during this reporting period. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
The laboratory experiments using the different DOC and COC fractions help to understand how these organic fractions facilitate E2 mobility and persistence in the environment. It was found that the DOC affected the solubility of E2, and that E2 can associate with the COC fractions of manure and soil, which can increase the mobility of E2 in water moving in the environment (e.g. field runoff, subsurface transport). The models that were developed helped identify the fate and transport parameters of the laboratory experiments. Ultimately, the desire is to have the laboratory experiments help to explain the field observations and potentially be used to develop better management practices. The new laboratory analytical procedure that was developed does not require the use of advanced analytical equipment, such as mass-spectrometry, to accurately quantify and qualify E2, its conjugates, and metabolites. This method is accessible to laboratories that do not have access to expensive analytical equipment. Furthermore, it allows a complete mass-balance determination and by quantifying the fate of estrogens and their metabolites in the aqueous, gas, and bound phases. Finally, E2 was observed in floodwaters in the Fargo-Moorhead community along the Red River of the North. The observations from the floodwaters indicated that E2 come from animal agriculture and/or wildlife habitat that had been flooded. No field measures of E2 have been analyzed from the subsurface drained field.

Publications

  • Casey, F.X.M. 2012. Sorption and degradation of estrogen conjugates in agricultural soils. 2012 Land Grant & Sea Grant National Water Conference, Portland, OR. USA.
  • Guy, A.C., T.M. DeSutter, F.X.M. Casey, R. Kolka and H. Hakk. 2012. Water quality, sediment, and soil characteristics near Fargo-Moorhead urban areas as affected by major flooding of the Red River of the North. J. Env. Qual. 41:554-563
  • Shrestha, S.L., F.X.M. Casey, H. Hakk, D.J. Smith and G. Padmanabhan. 2012. Fate and transformation of an estrogen conjugate and its metabolites in agricultural soils. Environ. Sci. Technol. 46:11047-11053


Progress 12/01/10 to 11/30/11

Outputs
OUTPUTS: In 2010/11, laboratory experiments were conducted to identify the potential for enhance estradiol transport and persistence by the mechanisms of i) associations with manure and soil colloidal and dissolve organic carbon (DOC), and ii) by estradiol derived from conjugates. Colloids and DOC were derived from from liquid swine manure and from soils where field experiments were conducted, and were characterized by size fractions. Radiolabelled (14C) estradiol was place in solution with the soil and manure colloids and DOC and measurements of estradiol partitioning or binding to the colloidal fractions were made. To identify the potential of estradiol conjugates of sulfate, a 14C-labelled sulfate conjugate of estradiol was synthesized. The 14C synthesized sulfate conjugate was then used in laboratory experiments to identify the persistence and binding of the conjugate and its metabolites in the soil. The soil experiments using the sulfate conjugate were then modeled. Field experiments were also commenced to identify whether swine manure derived estradiol can be transported through the soil depth to tile drainage installed at a depth of about 1 meter. Furthermore, the contribution of colloids/DOC and/or conjugates to the presence in the tile drainage is to be evaluated. Tile drainage accesses were installed and instruments to measure water flow and to sample water in the tile lines were installed. Additionally, wells were installed to monitor water table levels and to take groundwater samples. Our field research was highlighted in a tile-drainage tour of the farm where the research takes place. Additionally, research was presented to a multi-state tile-drainage workshop held in Fargo, ND and included researchers and practitioners from about 10 states. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Our laboratory experiments demonstrate that estradiol will associate with colloids derived from manures and can act to facilitate estradiol mobility in the environment. We found that the estradiol bound to colloids can also be dislodged. The implications of these results are that the colloids can facilitate estradiol movement above and through soil, perhaps to tile drainage water or groundwater, and then dissociate from the colloid to be free to negatively impact organisms. The laboratory experiments with the conjugate derived estradiol indicated that sulfate conjugates can transform into the potent estradiol molecule in soil solutions. The implication of these results from the conjugate studies is that conjugates, which are prevalent in animal manures, can contribute potent estradiol into the environment. A glucuronide conjugate of estradiol was detected in a field well water sample several meters below the land surface, which supports our conclusions that conjugates can enhance the mobility of estradiol through the soil. To date, manure was applied to our tile-drained field research site, but no results have been made.

Publications

  • Bai, X., F.X.M. Casey, H. Hakk, S.L. Shrestha, T.M. DeSutter, E. Khan and P.G. Oduor. 2011. Modeling Sorption and Degradation of 17beta-Estradiol-17-Sulfate in Agricultural Soils. In Annual Meetings Abstracts [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI.
  • Casey, F.X.M., and S. Shrestha. Persistence and Mobility of Estrogens in the Environment. NCERA 217. Fargo, ND. June 8, 2011. (Invited)
  • Chambers, K.B., F.X.M. Casey, H. Hakk, N.W. Shappell, T.M. DeSutter and E. Khan. 2011. Bioavailability of Colloidal Organic Carbon Bound Estrogen. In Annual Meetings Abstracts [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI.
  • Derby, N.E., H.H. Hakk, F.X.M. Casey and T.M. DeSutter. 2011. Effects of Composting Swine Manure on Nutrients and Estrogens. Soil Sci. 176:91-98.
  • Fan, Z., F.X.M. Casey, H. Hakk, G.L. Larsen, and E. Khan. 2011. Sorption, Fate, and Mobility of Sulfonamides in Soils. Water, Air, & Soil Pollution. 218:49-61.
  • Schuh, M., F.X.M. Casey, H. Hakk, T. DeSutter, K. Richards, E. Khan, and P. Oduor. 2011. An On-Farm Survey of Spatial and Temporal Stratifications of 17beta-Estradiol Concentrations. Chemosphere 82:1683-1689.
  • Schuh, M.C., F.X.M. Casey, H. Hakk, T.M. DeSutter, K.G. Richards, E. Khan, and P.G. Oduor. 2011. Effects of Field-Manure Applications on Stratified 17beta-Estradiol Concentrations. J. Hazard. Mater. 192:748-752.
  • Shrestha, S.L. 2011. Fate and Transformation of a Conjugated Natural Hormone 17beta-Estrradiol-3-Glucuronide in Soil-Water Systems. Ph.D. dissertation. North Dakota State Univ., Fargo.
  • Shrestha, S.L., X. Bai, D.J. Smith, H. Hakk, F.X.M. Casey, G.L. Larsen, and G. Padmanabhan. 2011. Synthesis and characterization of radiolabeled 17beta-estradiol conjugates. J. Labeled Compounds Radiopharmaceuticals. 54:267-271.
  • Zitnick, K.K., N.W. Shappell, H. Hakk, T.M. DeSutter, E. Khan, F.X.M. Casey. 2011. Effects of Liquid Swine Manure on Dissipation of 17beta-Estradiol in Soil. J. Hazard. Mater.


Progress 12/01/09 to 11/30/10

Outputs
OUTPUTS: Two methods to synthesis radiolabeled (14C) sulfate and glucuronide 17beta-estradiols were developed. One peer-reviewed publication was accepted regarding these synthesis methods. Soil batch sorption experiments were conducted using the 14C synthesized glucuronide and sulfate conjugates of estradiol to identify the potential of these conjugates in contributing estrogens to the environment. For the estradiol conjugate research, one presentation at an international scientific meeting was made, and one invited present at an international scientific meeting was given. Additionally, laboratory soil batch experiments were used to identify the effect of manure-derived colloidal (COC) and dissolved organic carbon (DOC) on the binding of estrogen to soil. A MS thesis was completed and a peer-review publication was accepted based on this laboratory study on the effect of COC/DOC on estradiol sorption and persistence in soil. For the field study portion of this project, a subsurface tiled-drained field was instrumented with wells and accesses to the existing tile drainage lines this summer. In the fall, two plots on this field received manure (one with controlled tile drainage and one with no controlled drainage). A third plot was considered a control and no manure was applied. Initial samples for water quality and quantity analysis have been taken from the three field plots. PARTICIPANTS: Not relevant to this project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Radiolabeled (14C) glucuronide (17beta-E2-3-G) and sulfate (17beta-E2-17-S) conjugates of estradiol and were successfully synthesized using enzymatic and chemical approaches, respectively, which permitted their use for laboratory scale fate and transport experiments in soil-water systems. Though our objective was measuring and modeling the movement of endocrine disrupting compounds in the environment, these studies are but a small portion of the potential studies in which radiolabeled conjugates could be used. Since glucuronidation and sulfation are the major conjugation pathways in vertebrates for not only steroid hormones, but other xenobiotics, then radiolabeled glucuronides and sulfates of other emerging contaminants may also be synthesized following the approaches provided in this paper, or with appropriate modifications of them. The facilitated transport and increased persistence of E2 may be caused by its association with manure-borne COC and/or DOC. Laboratory results showed that DOC/COC can lead to a greater persistence of E2 and reduced binding potential to the soil. Also, conjugates of E2 were shown to be more mobile and persistent in soil. The conjugates could result in the greater than expected mobility and persistence of estrogens in the environment. These laboratory results improve our understanding of the fate and transport of E2 in the environment, and help to explain field data. Also, the field and laboratory results together will improve management and treatment methods useful in improving water quality.

Publications

  • Bai, X., F.X.M. Casey, T. DeSutter, H. Hakk, P. Oduor and E. Khan. 2010. Sorption and Degradation of 17beta-Estradiol-17Sulfate in Agricultural Soils. In Annual Meetings Abstracts [CD-ROM]. ASA, CSSA, and SSSA, Madison, WI.
  • Casey, F.X.M., S. Shrestha, H. Hakk, D. Smith, G.L. Larsen, and G. Padmanabhan. 2009. The Fate and Transport of Reproductive Hormones in the Environment. In Fate and Transport of Microconstituents within the Subsurface Environment During Water Reuse Applications. American Geophysical Union, San Francisco, CA, December 14-18, 2009.
  • Shrestha, S.L., X. Bai, D.J. Smith, H. Hakk, F.X.M. Casey, G.L. Larsen, and G. Padmanabhan. 2010. Synthesis and characterization of radiolabeled 17beta-estradiol conjugates. J. Labeled Compounds Radiopharmaceuticals. (Accepted)
  • Zitnick, K.K. 2010. Effects of Liquid Swine Manure on Sorption of 17beta-estradiol to Soil. M.S. thesis North Dakota State Univ., Fargo.