Location: Food Animal Metabolism Research
2025 Annual Report
Objectives
Objective 1: Determine the absorption, distribution, metabolism, and excretion of emerging and legacy chemicals in food animals.
Sub-objective 1.A: Determine the metabolism and disposition of [14C]-nitrofurazone in broiler chickens.
Sub-objective 1.B: Determine the ADME of [14C]-PBDEs 47, 99, and 153 in laying turkeys.
Sub-objective 1.C: Determine the ADME of 1,3,7,8-tetrabromo [14C]-dibenzo-p-dioxin in laying hens.
Sub-objective 1.D: Determine the ADME of a defined mix of PFAS, including perfluorohexane sulfonic acid (PFHxS) in lactating cattle.
Sub-objective 1.E: Determine the fate of PFAS originating in a contaminated water source during the life cycle of laying hens.
Sub-objective 1.F: Determine the ADME of [14C]-(-)-trans-'9-tetrahydrocannabinol (THC) and/or [14C]-cannabidiol (CBD) in lactating dairy goats.
Sub-objective 1.G: Determine the accumulation and depuration kinetics of THC and CBD in feedlot cattle supplemented with dietary hemp.
Sub-objective 1.H: Evaluation of cellular uptake, translocation, and toxicity of microplastics using cell models.
Sub-objective 1.I: Determination of the fate of microplastics in laying hens.
Sub-objective 1.J: Determination of the uptake and depuration of microplastics in lactating dairy goats.
Objective 2: Develop and validate sensitive and accurate rapid analytical tools to detect emerging and legacy residues in food animals and food animal systems.
Sub-objective 2.A: Develop ambient ionization mass spectrometric detection and quantitation techniques of chemicals in matrices easily collected from live animals (blood, hair, urine, saliva).
Sub-objective 2.B: Develop ambient ionization mass spectrometric detection and quantitation techniques of chemicals in postmortem matrices (blood, tissues).
Objective 3: Determine levels and sources of emerging and legacy chemical or biological residues in the domestic food supply.
Sub-objective 3.A: In cooperation with regulatory agencies, determine the levels of dioxins, furans, and PBDEs in the U.S. meat supply.
Sub-objective 3.B: Determine the source(s) contributing to high background levels of PBDEs in commercial turkey.
Approach
Consumers loathe the idea of chemical residues in milk, meat, and eggs even though quantifiable risk of harm from chemicals in U.S. livestock products is exceedingly low. Regardless, consumers equate trace-levels of chemical residues in food with poor product quality and safety. Consequently, producers, regulatory officials, industry representatives, and consumers agree that chemical residues in food should be minimized to the greatest extent possible.
We propose to conduct absorption, distribution, metabolism, and excretion (ADME) studies on legacy and emerging chemicals for which significant data gaps exist. These chemicals include hemp-derived cannabinoids, a legacy antibiotic (nitrofurazone), halogenated persistent pollutants, and environmentally relevant microplastic contaminants (Objective 1). Basic ADME studies will allow the science-based selection of target matrices (saliva, urine, milk, liver, kidney, fat, etc.) and ‘marker compounds’ (parent compound or metabolites) of critical importance to the development of practical rapid screening technologies (Objective 2). ADME studies also provide data from which pre-harvest residue accumulation rates and post-exposure depuration rates can be calculated. Such data will facilitate the marketing of essentially residue-free animals in instances of known animal exposures. In some cases, especially for highly potent halogenated hydrocarbons and emerging contaminants, the U.S. government has a vested interest in ensuring that residues in remain well below regulatory thresholds. Under Objective 3, we propose a continuation of a 25-year cooperative effort with the USDA Food Safety and Inspection Service (FSIS) to survey the U.S. meat supply for dioxins and dioxin-like chemical residues. This survey has been critical to the discovery of environmental sources of dioxins and has been critical to reducing food animal exposures. We also propose to continue discovery efforts to elucidate contamination sources of livestock-based foods.
Collectively, the goal of this proposal is to develop science-based solutions that minimize consumer exposures to chemical residues in food animal products.
Progress Report
Research efforts relating to Objective 1, “Determine the absorption, distribution, metabolism, and excretion of emerging and legacy chemicals in food animals”. The analytical phase of a study to investigate the effects of a perfluoroalkyl substance (PFAS) -specific sorbent included in the diets of broiler chickens to help eliminate residues of PFAS was completed. Data has been analyzed and a draft of the manuscript prepared and awaiting cooperator approval.
The analytical phase (~1250 samples) of a two-part study to investigate the rates of life-time (6-weeks) PFAS accumulation (13 perfluorinated carboxylic acids and 12 perfluorinated sulfonates) in broiler chickens (n=70) and the lifetime (32-weeks) accumulation with subsequent depuration (9-weeks) of PFAS in laying hens (n=72) is ongoing. Analysis of PFAS in broiler chicken plasma, liver, and breast muscle is complete, with thigh muscle, gizzard, and skin samples processed but not analyzed. Analysis of laying hen plasma, liver, breast muscle, and thigh muscle are complete, with gizzard, skin, egg yolk, and egg white samples processed but not analyzed. Analyses are ongoing.
A live-phase study in laying hens (n=24) was conducted using the [14C]-polyethylene terphthalate (PET) microplastic synthesized in-house was completed in Q1 of FY2025. Laying hens were provided with a single bolus dose and samples (blood plasma, feces, eggs, and tissues) were collected at various withdrawal timepoints. Sample processing and analysis are ongoing.
A live-phase study in lactating sheep (n=7) was conducted using PET microplastic synthesized in-house was completed in Q1 of FY2025. Lactating sheep were provided with a single bolus dose and samples (blood plasma, urine, feces, and tissues) were collected at various withdrawal timepoints. Sample processing and analysis are ongoing.
A live-phase study in broiler chickens (n=68) was conducted in Q4 of FY24 to assess the usefulness of microalgal natural products already used as feed additives for broiler chickens to increase PFAS excretion from previously contaminated animals. Tissue samples were collected at various timepoints during feed additive treatment to assess the rate of PFAS elimination from edible tissues. All tissue processing is complete. The analytical phase is complete for blood plasma and liver samples with breast muscle, thigh muscle, and gizzard currently being analyzed.
Synthetic methods for the production of bead materials that can sorb PFAS compounds are being developed. The synthesis and characterization of the materials are ongoing. These beads initially will be used to study the remediation of PFAS contaminated water.
A synthetic method was developed for the production of different polymeric nanomaterials including polystyrene, co-polystyrene-polymethylmethacrylate for use in studying the transfer of micro- and nanoplastics into edible tissues of food animals. Characterizations of these nanomaterials are ongoing.
Liver samples collected during the metabolism and disposition study for [14C]-nitrofurazone were sent to collect proteomic data to help identify a marker compound for testing purposes. Data from the proteomic work has been received and analysis is in progress to evaluate potential marker peptides that are markedly different in abundance from control samples.
Objective 2 is focused on the development of analytical methods capable of rapidly and sensitively measuring chemical analytes in food-animal matrices. A cooperative study with the U.S. Food and Drug Administration and USDA’s Food Safety and Inspection Service is ongoing to validate a multi-residue method for measure 32 perfluoroalkyl substances including precursors, carboxylates, and sulfonates in agricultural matrices including blood plasma, muscle, egg, milk, liver, other edible products, and animal feed. The method is a one-step extraction utilizing internal standards to quantify compounds in = 50ng/kg (parts per trillion) in all matrices validated. Current progress includes single laboratory validation with eventual multi-lab validation. Additionally, current PFAS methods were transferred to a new liquid chromatograph – tandem mass spectrometer (LC-MS/MS) which has greater sensitivity for PFAS compounds compared to current laboratory instrumentation.
A quantitative carbohydrate method utilizing liquid chromatography-high resolution mass spectrometry was developed in FY24 to assess the concentrations of mono-, di-, and trisaccharides in sugarbeet root tissues. In FY25, the method’s usefulness was assessed in analyzing carbohydrate levels in harvested sugarbeet samples.
Objective 3’s goal is to determine the levels of dioxins, furans, and polybrominated diphenyl ethers (PBDEs) in beef, swine, and siluriformes of the U.S. meat supply. The collection of pork fat, beef (fat and liver) and siluriform samples (~400 total) was completed by USDA Food Safety and Inspection Service in early Q1 2025. The quantification of dioxins and furans in all sample extracts is completed. Validation of the PBDE analytical method on the gas chromatograph – tandem mass spectrometer (GC-MS/MS) was completed so sample extracts can be analyzed into Q1 of FY2026.
Accomplishments
1. Polybrominated diphenyl ether levels in U.S. meat, poultry, and siluriformes. Polybrominated diphenyl ethers (PBDEs) are man-made compounds that were used in industrial and consumer products. As a result of their high use and characteristics they are considered highly persistent environmental contaminants. Due to their structure, PBDEs are able to accumulate in the fat of food animals and animal products to a high degree. As a result of this accumulation, consumers can be exposed to the compounds through various meat products including beef, pork, chicken, turkey, and siluriformes. ARS scientists in Fargo, North Dakota, have regularly measured PBDE levels in fat samples from beef, pork, chicken, and turkey that come directly from animal processing facilities for the last two decades. The PBDE levels have been steadily declining and consumer exposure estimates calculated on consumption were found to be within or lower than values reported in other countries. The data provides reassuring information to U.S. producers that U.S. meat and poultry exports are relatively safe from PBDEs.
2. Tissue distribution of nitrofurazone in broiler chickens. Nitrofurazone is a synthetic antibiotic, but its use is banned in food animals due to its potential carcinogenicity and mutagenicity. Regulatory agencies monitor nitrofurazone’s illegal use in animal derived food products through a metabolite or marker compound, semicarbizide. But the current marker compound used to assess possible nitrofurazone use produces false positive detections which can result in rejection of exported products. Therefore, finding a nitrofurazone specific marker compound is necessary and of global interest. ARS scientists in Fargo, North Dakota, generated [14C]-nitrofurazone compounds by incorporating carbon-14 at the furaldehyde carbon or carbonyl carbon. These compounds were used to perform distribution, metabolism, and depletion studies in broiler chickens. The studies provided important information in relation to nitrofurazone metabolite stability, protein binding, and strides toward identification of a potential marker compound for nitrofurazone.
3. Synthesis of polyethylene terephthalate (PET) and generation of PET nanoparticles. Microplastics are environmental contaminants which are commonly measured in feed and water used for animal and human consumption. Due to the presence of microplastics in feed and water, there has been interest in measuring the transfer of these contaminants into animals and animal products used for human consumption. For these studies, often a label is incorporated into the compound or particle of interest. For this purpose, ARS scientists in Fargo, North Dakota, undertook the synthesis of polyethylene terephthalate (PET) to incorporate a carbon-14 radiolabel into the molecule for tracing capabilities. This radiolabeled microplastic was synthesized for use in absorption and elimination studies with laying hens and lactating sheep to determine the extent of distribution in animal products. These studies provide evidence on the absorption and elimination of PET microplastics in animal products.
Review Publications
Singh, A., Smith, D.J. 2025. Disposition of orally administered [14C]-nitrofurazone in broiler chickens. Journal of Agricultural and Food Chemistry. https://doi.org/10.1021/acs.jafc.4c07931.
Lupton, S.J., Smith, D.J., Howey, E.B., Predgen, A.S., Schmidt, C.E., Scholljegerdes, E., Ivey, S., Esteban, E., Johnston, J.J. 2025. Tissue histology and depuration of per- and polyfluoroalkyl substances (PFAS) from dairy cattle with lifetime exposures to PFAS contaminated drinking water and feed. Food Additives & Contaminants. Part A: Chemistry, Analysis, Control, Exposure & Risk Assessment. https://doi.org/10.1080/19440049.2024.2444560.
Singh, A., Shelver, W.L., Smith, D.J. 2025. Synthesis of 14C-labeled polyethylene terephthalate and generation of 14C-nanoparticles for fate and disposition studies. Journal of Labelled Compounds and Radiopharmaceuticals. https://doi.org/10.1002/jlcr.4137.
Lupton, S.J. 2025. Polybrominated diphenyl ethers (PBDEs) in US meat, poultry, and siluriformes: 2018 – 19 levels, trends, and estimated consumer exposures. Food Additives & Contaminants. Part A: Chemistry, Analysis, Control, Exposure & Risk Assessment. 42(4):452-464. https://doi.org/10.1080/19440049.2025.2457947.
Shelver, W.L., Mcgarvey, A.M., Billey, L.O. 2024. Disposition of [14C]-polystyrene microplastics after oral administration to lactating sheep. Food Additives & Contaminants. Part A: Chemistry, Analysis, Control, Exposure & Risk Assessment. 41:(9)1132-1143. https://doi.org/10.1080/19440049.2024.2379382.
Shelver, W.L., Billey, L.O., Mcgarvey, A.M., Hoselton, S.A., Banerjee, A. 2024. The effects of concentration, duration of exposure, size and surface function of polymethyl methacrylate micro/nanoplastics on human liver cells. Ecotoxicology and Environmental Safety. 287. Article 117240. https://doi.org/10.1016/j.ecoenv.2024.117240.