Heldur Hakk

Perfluorooctanoic acid (PFOA) is a perfluorinated alkyl substance (PFAS) used as surfactant in a wide variety of industrial and consumer products. Over the past decade, concern has increased over the presence of PFOA in biosolids from wastewater treatment plants used as fertilizer on agricultural lands having the potential to enter the food chain. In this study, the uptake of 14C-PFOA from soil by alfalfa was determined, as was the bioavailability of 14C-PFOA-incurred into alfalfa in Sprague-Dawley rats. Alfalfa leaves accumulated PFOA to as high as 4-5 µg/g of dry leaf, approximately 10 times higher than accumulation in the stem. Alfalfa was ground for feeding to 15 female Sprague-Dawley rats (175-200 g). Animals within metabolism cages were fed 10 g of feed (6 g alfalfa + 4 g ground rat chow) twice a day for 14 days (equivalent to 50 ug-PFOA/kg/day). At the end of the feeding period, rats (n=3) were sacrificed at withdrawal days of 0, 3, 7, 11, and 14 days. During the feeding and withdrawal phases, urine and feces were collected daily. At sacrifice, blood, liver, kidney, adipose, muscle, skin, brain, heart, adrenals, spleen, lungs, and thymus were removed and assayed for 14C-PFOA by combustion and LSC analysis. Rats eliminated 72.8 ± 3.4% of the total dose via urine at 14-days, but urinary radioactivity fell below the LOD by day 3 of the withdrawal period. Fecal elimination was 6.5 ± 1.2 % of the dose and fell below the LOD by 2 days of withdrawal. The rapid and high elimination via urine indicates that a majority of the dose was absorbed. The uptake of PFOA into alfalfa was low from a high organic content soil, however, PFOA was highly bioavailable from the alfalfa when used as a feed component for rats. This study provides data for regulators investigating PFOA bioavailability and disposition in animals or animal products exposed to contaminated feed.

Polybrominated diphenyl ethers (PBDEs) levels in environmental matrices have generally declined following their phaseout as flame retardants. The objective of this study was to determine the absorp...

Metabolism, environmental fate, and low concentration food residue studies would be facilitated by the use of radiolabeled test articles that can be readily quantified within complex matrices. However, radiochemical approaches for such studies require high specific activities to allow analytical detection of correspondingly low masses of test article. The synthesis of high specific activity (>50 μCi/μmol) [¹⁴C]‐radiolabeled phenylbutazone presents a challenge using existing methodology, mainly due to the low solvent volumes required and vigorous refluxing needed to close the pyrazolidinedione ring. Herein, we report on the significant modification of an existing method that allows the synthesis of low masses of high specific activity (>50 μCi/μmol) [¹⁴C]‐phenylbutazone under mild conditions with simple purification and high yield. The closure of the pyrazolidinedione ring of 1,2‐diphenyl‐3,5‐pyrazolidinedione was accomplished as a first step with unlabeled 1,2‐diphenylhydrazine and diethyl malonate in 32% yield under gram‐scale conditions, which avoided the challenges of low solvent use and vigorous refluxing. Low mass of high specific activity n‐[1‐¹⁴C]‐butyl bromide was then added via a nucleophilic substitution reaction as a final step. Yields ranged from 65% to 92% during multiple synthetic attempts with unlabeled butyl bromide and were greatly influenced by reaction stoichiometry and the selection of base.

Seven animal drugs [penicillin G (PENG), sulfadimethoxine (SDMX), oxytetracycline (OTET), erythromycin (ERY), ketoprofen (KETO), thiabendazole (THIA), and ivermectin (IVR)] were used to evaluate the drug distribution between milk fat and skim milk fractions of cow milk. More than 90% of the radioactivity was distributed into the skim milk fraction for ERY, KETO, OTET, PENG, and SDMX, approximately 80% for THIA, and 13% for IVR. The distribution of drug between milk fat and skim milk fractions was significantly correlated to the drug’s lipophilicity (partition coefficient, log P, or distribution coefficient, log D, which includes ionization). Data were fit with linear mixed effects models; the best fit was obtained within this data set with log D versus observed drug distribution ratios. These candidate empirical models serve for assisting to predict the distribution and concentration of these drugs in a variety of milk and milk products.

Male Sprague–Dawley rats were dosed orally with 3 mg/kg of one of three hexabromocyclododecane (HBCD) diastereomers. Each diastereomer was well absorbed (73–83%), and distributed preferentially to lipophilic tissues. Feces were the major route of excretion; cumulatively accounting for 42% of dose for α-HBCD, 59% for ß-HBCD, and 53% for γ-HBCD. Urine was also an important route of HBCD excretion, accounting for 13% of dose for α-HBCD, 30% for ß-HBCD, and 21% for γ-HBCD. Total metabolism of HBCD diastereomers followed the rank order ß > γ > α, and was >65% of that administered. The metabolites formed were distinct in male rats: α-HBCD did not debrominate or stereoisomerize, but formed two hydroxylated metabolites; ß- and γ-HBCD were both extensively metabolized via pathways of stereoisomerization, oxidation, dehydrogenation, reductive debromination, and ring opening. ß-HBCD was biotransformed to two mercapturic acid pathway metabolites. The metabolites of ß- and γ-HBCD were largely distinct, and could possi...

The fate of [C]17β-estradiol ([C]E2) was monitored for 42 d in triplicate 10-L anaerobic digesters. Total radioactive residues decreased rapidly in the liquid layer of the digesters and reached a steady-state value of 22 to 26% of the initial dose after 5 d. High-performance liquid chromatography and liquid chromatography-tandem mass spectrometry analyses of the liquid layer of the anaerobic digesters indicated a rapid degradation of E2 to estrone (E1), which readily adsorbed to the sludge layer subsequent to its formation. Estrone was the predominant steroid identified under anaerobic digestion in the liquid layer or sorbed to sludge at 42 d. Methane formation represented 11.1 ± 5.7% of the initial E2 fortification with 0.3 to 0.5% of the starting E2 mineralized to carbon dioxide. Maximum [C]methane production appeared between Days 4 and 7. An estimate of estrogenicity of the final product based on reported estrogen equivalents for E1 and E2 was 2% of the original in active digesters. Anaerobic digestion of swine waste has several management benefits; moreover, this study demonstrated that it reduces the potential of environmental release of estrogens, which are known endocrine disruptors.

The metabolism of α- and γ-hexabromocyclododecane (HBCD) was investigated in adult C57BL/6 female mice. α- or γ-[¹⁴C]HBCD (3 mg/kg bw) was orally administered with subsequent urine and feces collection for 4 consecutive days; a separate group of mice was dosed and sacrificed 3 h postexposure to investigate tissue metabolite levels. Extractable and nonextractable HBCD metabolites were quantitated in liver, blood, fat, brain, bile, urine, and feces and characterized by LC/MS (ESI−). Metabolites identified were distinct between the two stereoisomers. In mice exposed to α-HBCD, four hydroxylated metabolites were detected in fecal extracts, and one of these metabolite isomers was consistently characterized in liver, brain, and adipose tissue extracts. In contrast, fecal extracts from mice exposed to γ-HBCD contained multiple isomers of monohydroxy-pentabromocyclododecene, dihydroxy-pentabromocyclododecene, and dihydroxy-pentabromocyclododecadiene, while in liver and adipose tissues extracts only a single monohydroxy-pentabromocyclododecane metabolite was observed. Both stereoisomers were transformed to metabolites which formed covalent bonds to proteins and/or lipids in the gut as suggested by high fecal nonextractables. The presence of tissue- and excreta-specific metabolic products after in vivo exposure to the two main HBCD stereoisomers supports previous toxicokinetic studies indicating that these two stereoisomers are biologically distinct. The distinct metabolic products identified in this study have the potential to aid in the identification of stereoisomer-specific HBCD exposures in future biomonitoring studies.

The excreted estrogen rate of all livestock in the United States is estimated at 134 kg d⁻¹. The influence of manure treatment on the fate of estrogens is critical in deciding the recycling of over 300 million dry tons of livestock produced annually. The effects of two common manure management practices, heated composting and ambient temperature decomposition, on the fate of 17β‐estradiol in poultry litter were determined. A mixture of poultry litter, wood chips, and straw was amended with [¹⁴C]17β‐estradiol and allowed to undergo decomposition with a laboratory‐scale heated composter (HC) or room temperature incubation (RTI) for 24 d. Radiolabel in the finished products was fractionated into water‐extractable, acetone‐extractable, nonextractable, and mineralized fractions. Total 17β‐estradiol radioactive residues in the HC and RTI (n = 2) treatments were not different (P > 0.05), except that statistically less 17β‐estradiol was mineralized to ¹⁴CO₂ during HC than RTI (1.1 vs. 10.0% for HC and RTI, respectively). Estrone was the major degradation product in extracts of HC and RTI treatments as determined by liquid chromatography/mass spectrometry analyses. The nonextractable residues indicated no quantitative differences among the humins between the treatments. An estimated 3% of the fortified estrogenicity remained after HC treatment, and 15% of the fortified estrogenicity remained after RTI treatment. If reduction of water‐removable, biologically active 17β‐estradiol is the treatment goal, then HC treatment would be slightly preferred over ambient temperature degradation. However, unmanaged, ambient temperature litter piles are less costly and time consuming for food animal producers and result in greater mineralization and similar immobilization of estradiol.