1a. Objectives (from AD-416)
All animals produce natural hormones in the healthy function and regulation of their endocrine system. When a hormone is eliminated from the body and gets into the environment, it can act upon other animals as an endocrine disrupting compound by binding to and activating receptors within hormone-responsive organs. In sensitive organisms this can occur at the parts per trillion level, and risks include sex reversals and cancer. The potential for livestock farms to serve as sources of hormones to the environment has been recognized for many years. To guide the objectives of this proposal, the following research questions (RQ) were formulated to evaluate the respective testable hypotheses (TH): RQ 1) Does dissolved organic carbon and particulates from lagoon waste or soil enhance the mobility of estrogens in mineral soil and thus increase the potential for off-site movement? TH 1) If the movement of estrogens is related to the presence of lagoon- or soil-derived dissolved organic carbon and particulates, then estrogen in the presence of high amounts of soil/lagoon dissolved organic carbon will increase their mobility compared to aqueous free estrogens. RQ 2) Does dissolved organic carbon and particulates of lagoon waste or soil offer estrogenic hormones protection from microbial degradation and increase their longevities in the soil or manure storage facilities? TH 2) If estrogenic hormones are bound to dissolved organic carbon or particulates and not easily accessible to microbial processes, then hormone in the presence of high amounts of dissolved organic carbon or particulates will increase persistence in the environment compared to free aqueous hormone. RQ 3) Do estrogen conjugates (i.e. polar metabolites) promote the mobility or persistence of estrogens in soil? TH 3) If conjugated hormones are more water soluble than their de-conjugated forms, then they are also more mobile in the soil and can potentially migrate to areas of reduced microbial degradation. RQ 4) Do estrogens and their conjugates from lagoon waste applied to fields move into groundwater and into surface waters? TH 4) If the mobility of estrogens in soil is dependant on hormone conjugation, the presence of dissolved organic carbon or particulates, and the low retention time in upper soil layers, then application of manures on tile drainage fields will increase the presence of hormone/hormone conjugates in ground and surface waters. To answer these research questions and test the hypotheses, the following experimental objectives were formed: Objective 1 – Determine the sorption characteristics of estradiol, estradiol glucuronide and sulfate conjugates with particulates originating from natural organic matter from soil and from animal manure lagoon liquors (laboratory); Objective 2 – Determine the degradation of the parent compounds estradiol, estradiol glucuronide and sulfate conjugates in the presence of these particulates and soil (laboratory); and Objective 3 – Determine how land application strategies (surface applied vs. injection) for lagoon liquor influence the fate and transport of estradiol in a field setting.
1b. Approach (from AD-416)
Objective 1: Degradation of hormones is mediated primarily by biological activity and can lead to misinterpretation of observed physical/chemical soil absorption processes. In order to identify these processes experimentally, the experiments for this objective will be conducted under sterile conditions. Four hormone concentrations and nine time points will be used and sampled. Qualititative and quantitative assessment of hormone degradation and soil absorption will be made by chromatographic and spectrometric methods, and absorption curves and models can then be determined. Objective 2: High and low organic soils will be spiked with radioactive hormone conjugates with and without lagoon waste under aerobic/anaerobic and sterile/non-sterile conditions to measure biological degradation in soil. Mass balances, qualitative assessment of hormone fate, and separation of hormone compartments into the various soil fractions under these conditions will be determined. Objective 3: Well-defined field-scale Discovery Farms will be used to measure the transport of hormones when injected into soil and followed by uncontrolled and controlled drainage (rainfall). Both well water and soil extracts will be analyzed for hormone content across two growing seasons.
3. Progress Report
Estrogens are potent agonists of the estrogen receptors, and when exposed exogenously can alter an organism’s normal hormone balance. This can lead to reproductive abnormalities, such as feminization of male organisms. A major source of estrogens into the environment is from animal waste, particularly in the microenvironment surrounding concentrated animal feeding operations. However, despite the consistent detection of hormones in the environment, our previous laboratory data suggested that estrogens should not be mobile in the environment, and that movement into groundwater must be governed by more than vertical transport in soil by rainfall. Therefore, mechanisms need to be discovered on how estrogens enter and persist in the environment. Last year we discovered two such mechanisms, and this year have continued investigations. 1. Flushed swine manure wastewaters are composed of two compartments, i.e. the dissolved aqueous compartment and the particulate compartment. The particulate compartment was discovered to be composed of a sedimentary (sludge) and suspended fraction (colloidal). This colloidal fraction has significant potential for long-range transport, but it is not known if the colloidal-bound estrogens are still biologically active. We developed a method to isolate the colloidal fraction by ultrafiltration, and to quantitate the bound estrogen by radiochemical means. An assay was then developed that could assess the biological activity. Only preliminary results on real-world samples have been generated, however, it is hoped that results of these assays will determine whether these mobile, estrogen-bound colloids are biologically active and/or environmentally stable. 2. Last year it was shown that water-soluble conjugates of estrogens have significant potential to be transported in runoff to contaminate surface and groundwater. This year our investigations demonstrated that the various water-soluble conjugates can be degraded in the environment, but at different rates and to different endpoints. The degradation of some estrogen conjugates yielded metabolites that were not biologically active compounds, while others were degraded back to biologically active parent estrogen. The observed results were fitted into a robust model so that predictions on estrogen fate and transport could be made based on soil type. These results are very significant in understanding the fate and transport behavior of manure-borne estrogens, and to devise manure management methods that reduce their environmental release.