Perfluorooctanesulfonate (PFOS) Conversion from N-Ethyl-N-(2-hydroxyethyl)-perfluorooctanesulfonamide (EtFOSE) in male Sprague Dawley rats after inhalation exposure.

Ethyl-N-(2-hydroxyethyl)-perfluorooctanesulfonamide (EtFOSE) was one of the key building blocks for many of the perfluorooctanesulfonyl-based chemistry and laboratory studies have shown that EtFOSE can metabolically degrade to perfluorooctanesulfonate (PFOS). Non-occupational contribution sources to PFOS are thought to occur in general population via diets, drinking water, air and dust. For workers, however, the exposure route was mostly airborne and the exposure source was predominantly to precursor compounds such as EtFOSE. We undertook this study to investigate how much EtFOSE was converted to PFOS in the serum for male rats after 6h of exposure to EtFOSE vapor (whole body) at ambient temperature, which simulated a work place exposure scenario. There were no abnormal clinical observations and all rats gained weight during study. Interim tail-vein blood samples, collected up to 21 days after exposure, were analyzed for Et-FOSE and PFOS concentrations by LC-MS/MS. Upon inhalation exposure, the biotransformation of EtFOSE to PFOS in serum in the male rats was rapid and very little EtFOSE was detected in the serum within 24h after EtFOSE exposure. The highest conversion to PFOS in serum after exposure to EtFOSE vapor appeared to occur between Day 8-14 post exposure. Considering the potential surface and fur adsorption of test compound in the whole-body exposure system, our data would support that at least 10% of the inhaled EtFOSE was biotransformed to PFOS in the serum based on the range of lower 95% CI (confidence interval) values. This information is valuable because it quantitatively translates EtFOSE exposure into serum PFOS concentration, which serves as a matrix for internal dosimetry (of PFOS exposure) that can be used as an anchor across species as well as between different exposure routes.

Four-week dietary supplementation with 10- and/or 15-fold basal choline caused decreased body weight in Sprague Dawley rats.

Choline is an essential nutrient utilized for phosphatidylcholine biosynthesis and lipoprotein packaging and secretion. Recently, choline supplementation has been used by athletes and the public for weight loss. However, the potential toxicological impact of choline dietary supplementation requires further investigation. This study examined the effects of choline dietary supplementation in Sprague Dawley rats for 4 weeks. Rats were fed diets containing basal choline levels (control) or 5-, 10-, or 15-fold (5×, 10×, or 15×) basal diet concentration. In groups fed choline-supplemented diets, there were no toxicologically relevant findings in clinical observations, food intake, clinical chemistry, liver weights, or liver histopathology. However, decreased mean body weights (8.5-10.2%) and body weight gains (24-31%) were noted for the 10× choline-supplemented (females only) and 15× choline-supplemented (both sexes) groups relative to the control groups from day 3 onward. These body weight effects were not related to a persistent reduction in average food intake. Serum cholesterol was increased in the 15× choline-supplemented male rats relative to the controls, an expected effect of choline supplementation; however, there were no changes in the serum cholesterol of female rats. Serum choline concentrations were increased in female rats relative to the male rats across all treatment groups. The maximum tolerated dose for male and female rats were the 15× and 10× choline supplements, respectively, based on decreased mean body weight and body weight gains. This study supported the conclusions of a clinical trial that showed a high choline diet can decrease body weight in humans.

Hepatocellular hypertrophy and cell proliferation in Sprague-Dawley rats following dietary exposure to ammonium perfluorooctanoate occurs through increased activation of the xenosensor nuclear receptors PPARα and CAR/PXR.

Ammonium perfluorooctanoate (APFO), a processing aid used in the production of fluoropolymers, produces hepatomegaly and hepatocellular hypertrophy in rodents. In mice, APFO-induced hepatomegaly is associated with increased activation of the xenosensor nuclear receptors, PPARα and CAR/PXR. Although non-genotoxic, chronic dietary treatment of Sprague-Dawley (S-D) rats with APFO produced an increase in benign tumours of the liver, acinar pancreas, and testicular Leydig cells. Most of the criteria for establishing a PPARα-mediated mode of action for the observed hepatocellular tumours have been previously established with the exception of the demonstration of increased hepatocellular proliferation. The present study evaluates the potential roles for APFO-induced activation of PPARα and CAR/PXR with respect to liver tumour production in the S-D rat and when compared to the specific PPARα agonist, 4-chloro-6-(2,3-xylidino)-2-pyrimidinylthioacetic acid (Wy 14,643). Male S-D rats were fed APFO (300 ppm in diet) or Wy 14,643 (50 ppm in diet) for either 1, 7, or 28 days. Effects of treatment with APFO included: decreased body weight; hepatomegaly, hepatocellular hypertrophy, hepatocellular hyperplasia (microscopically and by BrdU labelling index), and hepatocellular glycogen loss; increased activation of PPARα (peroxisomal ß-oxidation and microsomal CYP4A1 protein); decreased plasma triglycerides, cholesterol, and glucose; increased activation of CAR (CYP2B1/2 protein) and CAR/PXR (CYP3A1 protein). Responses to treatment with Wy 14,643 were consistent with increased activation of PPARα, specifically: increased CYP4A1 and peroxisomal ß-oxidation; increased hepatocellular hypertrophy and cell proliferation; decreased apoptosis; and hypolipidaemia. With the exception of decreased apoptosis, the effects observed with Wy 14,643 were noted with APFO, and APFO was less potent. These data clearly demonstrate an early hepatocellular proliferative response to APFO treatment and suggest that the hepatomegaly and tumours observed after chronic dietary exposure of S-D rats to APFO likely are due to a proliferative response to combined activation of PPARα and CAR/PXR. This mode of action is unlikely to pose a human hepatocarcinogenic hazard.

Pathology Working Group review and evaluation of proliferative lesions of mammary gland tissues in female rats fed ammonium perfluorooctanoate (APFO) in the diet for 2 years.

Perfluorooctanoate (PFO) is a perfluorinated carboxylate that is widely distributed in the environment. A 2-year chronic study was conducted in rats fed either 30 or 300 ppm of ammonium perfluorooctanoate (APFO). To investigate the possible relationship of APFO exposure to proliferative mammary lesions, a Pathology Working Group (PWG) review of the original slides was performed. The consensus reached by the PWG was that the incidence of mammary-gland neoplasms was not affected by chronic dietary administration of APFO. Therefore, feeding female rats up to 300 ppm of APFO resulted in no increase in proliferative lesions of the mammary tissue.

Perfluorooctane sulfonate alters gut microbiota-host metabolic homeostasis in mice.

Perfluorooctane sulfonate (PFOS) is a persistent environmental chemical whose biological effects are mediated by multiple mechanisms. Recent evidence suggests that the gut microbiome may be directly impacted by and/or alter the fate and effects of environmental chemicals in the host. Thus, the aim of this study was to determine whether PFOS influences the gut microbiome and its metabolism, and the host metabolome. Four groups of male C57BL/6 J mice were fed a diet with or without 0.003 %, 0.006 %, or 0.012 % PFOS, respectively. 16S rRNA gene sequencing, metabolomic, and molecular analyses were used to examine the gut microbiota of mice after dietary PFOS exposure. Dietary PFOS exposure caused a marked change in the gut microbiome compared to controls. Dietary PFOS also caused dose-dependent changes in hepatic metabolic pathways including those involved in lipid metabolism, oxidative stress, inflammation, TCA cycle, glucose, and amino acid metabolism. Changes in the metabolome correlated with changes in genes that regulate these pathways. Integrative analyses also demonstrated a strong correlation between the alterations in microbiota composition and host metabolic profiles induced by PFOS. Further, using isolated mouse cecal contents, PFOS exposure directly affected the gut microbiota metabolism. Results from these studies demonstrate that the molecular and biochemical changes induced by PFOS are mediated in part by the gut microbiome, which alters gene expression and the host metabolome in mice.

Toxicological evaluation of potassium perfluorobutanesulfonate in a 90-day oral gavage study with Sprague-Dawley rats.

Perfluorobutanesulfonate (PFBS) is a surfactant and degradation product of substances synthesized using perfluorobutanesulfonyl fluoride. A 90-day rat oral gavage study has been conducted with potassium PFBS (K+PFBS). Rats were dosed with K+PFBS at doses of 60, 200, and 600mg/kg-day body weight. The following endpoints were evaluated: clinical observations, food consumption, body weight, gross and microscopic pathology, clinical chemistry, and hematology. In addition, functional observation battery and motor activity assessments were made. Histological examination included tissues in control and 600 mg/kg-day groups. Additional histological examinations were performed on nasal cavities and turbinates, stomachs, and kidneys in the 60 and 200 mg/kg-day groups. No treatment-related mortality, body weight, or neurological effects were noted. Chromorhinorrhea (perioral) and urine-stained abdominal fur were observed in males at 600mg/kg-day. Red blood cell counts, hemoglobin, and hematocrit values were reduced in males receiving 200 and 600mg/kg-day; however, there were no adverse histopathological findings in bone marrow. Total protein and albumin were lower in females at 600mg/kg-day. There were no significant changes in clinical chemistry in either sex. All rats appeared normal at sacrifice. Microscopic changes were observed only at the highest dose in the stomach. These changes consisted of hyperplasia with some necrosis of the mucosa with some squamous metaplasia. These effects likely were due to a cumulative direct irritation effect resulting from oral dosing with K+PFBS. Histopathological changes were also observed in the kidneys. The changes observed were minimal-to-mild hyperplasia of the epithelial cells of the medullary and papillary tubules and the ducts in the inner medullary region. There were no corresponding changes in kidney weights. Clinical chemistry parameters related to kidney function were unchanged. These kidney findings are likely due to a response to high concentration of K+PFBS in tubules and ducts and represent a minimal-to-mild effect. Microscopic changes of an equivocal and uncertain nature were observed in the nasal mucosa and were likely attributable to the route of dosing (oral gavage). The NOAEL for the female rat in this study was 600 mg/kg-day (highest dose of study). The NOAEL for the male rat was 60 mg/kg-day based on hematological effects.

A two-generation oral gavage reproduction study with potassium perfluorobutanesulfonate (K+PFBS) in Sprague Dawley rats.

Perfluorobutanesulfonate (PFBS) is a surfactant and degradation product of substances based on perfluorobutanesulfonyl fluoride. A two-generation reproductive rat study has been conducted with potassium PFBS (K(+)PFBS). Parental-generation (P) rats were dosed orally by gavage with 0, 30, 100, 300 and 1000mg K(+)PFBS/kg/day for 10 weeks prior to and through mating (males and females), as well as during gestation and lactation (females only). First generation (F1) pups were dosed similarly, beginning at weaning. Second generation (F2) pups were not directly dosed but potentially exposed to PFBS through placental transfer and nursing, and the study was terminated 3 weeks after their birth. Endpoints evaluated included body weight, food consumption, clinical signs, estrus cycling, sperm quality, pregnancy, natural delivery, litter outcomes, and developmental landmarks. The no-observable-adverse effect dose level (NOAEL) in the parental generations (P and F1) was 100mg/kg/day. In the 300 and 1000mg/kg/day dose group rats, there were (1) increased liver weight (absolute or relative) and corresponding increased incidence of adaptive hepatocellular hypertrophy (male only) and (2) increased incidence of minimal to mild microscopic findings in the medulla and papilla of the kidneys (male and female). There were no K(+)PFBS treatment-related effects on fertility or reproduction among the P or the F1 rats. There were no microscopic changes in male or female reproductive organs, and no biologically relevant effects on sperm parameters, mating, estrous cycles, pregnancy, and natural delivery in the P- or F1-generations. There were no K(+)PFBS treatment-related effects on survival of pups in the two-generation study. Litter size and average pup birth weight per litter were not statistically significantly different from controls in any dose group. In the F1-generation, terminal body weight was reduced in males at 1000mg/kg/day. Preputial separation was slightly delayed (approximately 2 days) at this dose, a finding consistent with the body weight reduction. Essentially no effects were observed in the F1 females. F2 pups had normal body weights. The reproductive NOAEL was >1000mg/kg/day in both generations.

A comparison of the pharmacokinetics of perfluorobutanesulfonate (PFBS) in rats, monkeys, and humans.

Materials derived from perfluorobutanesulfonyl fluoride (PBSF, C(4)F(9)SO(2)F) have been introduced as replacements for eight-carbon homolog products that were manufactured from perfluorooctanesulfonyl fluoride (POSF, C(8)F(17)SO(2)F). Perfluorobutanesulfonate (PFBS, C(4)F(9)SO(3)(-)) is a surfactant and potential degradation product of PBSF-derived materials. The purpose of this series of studies was to evaluate the pharmacokinetics of PFBS in rats, monkeys, and humans, thereby providing critical information for human health risk assessment. Studies included: (1) intravenous (i.v.) elimination studies in rats and monkeys; (2) oral uptake and elimination studies in rats; and (3) human serum PFBS elimination in a group of workers with occupational exposure to potassium PFBS (K(+)PFBS). PFBS concentrations were determined in serum (all species), liver (rats), urine (all species), and feces (rats). In rats, the mean terminal serum PFBS elimination half-lives, after i.v. administration of 30mg/kg PFBS, were: males 4.51+/-2.22h (standard error) and females 3.96+/-0.21h. In monkeys, the mean terminal serum PFBS elimination half-lives, after i.v. administration of 10mg/kg PFBS, were: males 95.2+/-27.1h and females 83.2+/-41.9h. Although terminal serum half-lives in male and female rats were similar, without statistical significance, clearance (CL) was significantly greater in female rats (469+/-40mL/h) than male rats (119+/-34mL/h) with the area under the curve (AUC) significantly larger in male rats (294+/-77microg.h/mL) than female rats (65+/-5microg.h/mL). These differences were not observed in male and female monkeys. Volume of distribution estimates suggested distribution was primarily extracellular in both rats and monkeys, regardless of sex, and urine appeared to be a major route of elimination. Among 6 human subjects (5 male, 1 female) followed up to 180 days, the geometric mean serum elimination half-life for PFBS was 25.8 days (95% confidence interval 16.6-40.2). Urine was observed to be a pathway of elimination in the human. Although species-specific differences exist, these findings demonstrate that PFBS is eliminated at a greater rate from human serum than the higher chain homologs of perfluorooctanesulfonate (PFOS) and perfluorohexanesulfonate (PFHxS). Thus, compared to PFOS and PFHxS, PFBS has a much lower potential for accumulation in human serum after repeated occupational, non-occupational (e.g., consumer), or environmental exposures.

Perfluoroalkyl acids and related chemistries--toxicokinetics and modes of action.

The perfluoroalkyl acid salts (both carboxylates and sulfonates, hereafter designated as PFAAs) and their derivatives are important chemicals that have numerous consumer and industrial applications. However, recent discoveries that some of these compounds have global distribution, environmental persistence, presence in humans and wildlife, as well as toxicity in laboratory animal models, have generated considerable scientific, regulatory, and public interest on an international scale. The Society of Toxicology Contemporary Concepts in Toxicology Symposium, entitled "Perfluoroalkyl Acids and Related Chemistries: Toxicokinetics and Modes-of-Action Workshop" was held February 14-16, 2007 at the Westin Arlington Gateway, Arlington, VA. In addition to the Society of Toxicology, this symposium was sponsored by 3M Company, DuPont, Plastics Europe, and the U.S. Environmental Protection Agency. The objectives of this 3-day meeting were to (1) provide an overview of PFAA toxicity and description of recent findings with the sulfonates, carboxylates, and telomer alcohols; (2) address the toxicokinetic profiles of various PFAAs among animal models and humans, and the biological processes that are responsible for these observations; (3) examine the possible modes of action that determine the PFAA toxicities observed in animal models, and their relevance to human health risks; and (4) identify the critical research needs and strategies to fill the existing informational gaps that hamper risk assessment of these chemicals. This report summarizes the discourse that occurred during the symposium.

Perfluorooctane sulfonate-induced changes in fetal rat liver gene expression.

In utero exposure of laboratory rats to perfluorooctane sulfonate (PFOS, C(8)F(17)SO(3)(-)), a chemically stable surfactant that is widely disseminated in the environment and present in serum samples from wildlife and humans, is associated with decreased neonatal survival, and growth deficits as well as hepatomegaly. This hepatomegaly in newborn rats exposed to PFOS in utero resembles that observed in adults and is characterized by peroxisome proliferation and decreased liver triglycerides, both of which are suspected to be manifested through PPARalpha-mediated transcriptional regulation. The purpose of the present investigation was to determine whether these changes in metabolic status are a reflection of transcriptional changes in fetal rat liver using global gene expression array analyses. Gravid Sprague-Dawley rats were administered 3mg/kg PFOS by gavage daily from gestational day 2-20 and terminated on day 21. Although there was no treatment-related frank terata, there was a substantial effect of PFOS on the perinatal hepatic transcriptome-225 unique transcripts were identified as statistically increased and 220 decreased by PFOS exposure; few transcripts were changed by more than two-fold. Although the PPARalpha transcript (Ppara) itself was not affected, there was a significant increase in expression of gene transcripts associated with hepatic peroxisomal proliferation as well as those responsible for fatty acid activation, transport and oxidation pathways (both mitochondrial and peroxisomal). Additional metabolic pathways altered by in utero PFOS exposure were a stimulation of fetal hepatic fatty acid biosynthesis and a net reduction of Cyp7a1 transcript, which is required for bile acid synthesis. There were minimal effects on the expression of thyroid-related gene transcripts. In conclusion, gene expression analysis provides strong evidence indicating transcriptional control of the altered metabolic status of neonates following PFOS exposure in utero, much of which appears to be under the influence of a functional perinatal PPARalpha regulatory pathway.

Thyroid hormone status and pituitary function in adult rats given oral doses of perfluorooctanesulfonate (PFOS).

INTRODUCTION: Perfluorooctanesulfonate (PFOS) is widely distributed and persistent in humans and wildlife. Prior toxicological studies have reported decreased total and free thyroid hormones in serum without a major compensatory rise in thyrotropin (TSH) or altered thyroid gland histology. Although these animals (rats, mice and monkeys) might have maintained an euthyroid state, the basis for hypothyroxinemia remained unclear. We undertook this study to investigate the causes for the PFOS-induced reduction of serum total thyroxine (TT4) in rats. HYPOTHESES: We hypothesized that exposure to PFOS may increase free thyroxine (FT4) in the rat serum due to the ability of PFOS to compete with thyroxine for binding proteins. The increase in FT4 would increase the availability of the thyroid hormone to peripheral tissues for utilization, metabolic conversation, and excretion. We also hypothesized that PFOS does not directly interfere with the regulatory functions of the hypothalamic-pituitary-thyroid (HPT) axis in rats. EXPERIMENTS: Three experimental designs were employed to test these hypotheses. (1) Female Sprague-Dawley (SD) rats were given a single oral dose of 15 mg potassium PFOS/kg body weight. At intervals of 2, 6, and 24h thereafter, measurements were made for serum FT4, TT4, triiodothyronine (TT3), reverse triiodothyronine (rT3), thryrotropin (TSH), and PFOS concentrations, as well as liver PFOS concentrations, UDP-glucuronosyltransferase 1A (UGT1A) family mRNA transcripts, and malic enzyme (ME) mRNA transcripts and activity. (2) To provide evidence for increased uptake and metabolism of thyroxine (T4), 125 I-T4 was given to male and female SD rats by intravenous injection, followed in 2h by a single oral dose of 15 mg potassium PFOS/kg body weight. 125 I radioactivity was determined in urine and feces collected over a 24-h period and in serum and liver collected at 24h. (3) To assess the potentials effect of PFOS on the hypothalamic-pituitary-thyroid axis, over an 8-day period, groups of male SD rats were given PFOS (3mg/kg-d), propyl thiouracil (PTU, 10 microg/mL in water), or PTU and PFOS in combination, with controls receiving 0.5% Tween 20 vehicle. On days 1, 3, 7, and 8, TT4, TT3, and TSH were monitored. On day 8, pituitaries were removed and placed in static culture for assessment of thyrotropin releasing hormone (TRH)-mediated release of TSH. RESULTS: (1) PFOS transiently increased FT4 and decreased TSH within 6h, with values returning to control levels by 24h. TT4 was decreased by 55% over a 24-h period. TT3 and rT3 were decreased at 24h to a lesser extent than TT4. ME mRNA transcripts were increased at 2h and activity was increased at 24h. UGT1A mRNA transcripts were increased at 2 and 6h. (2) 125 I decreased in serum and liver relative to controls and consistent with a reduction in serum TT4. Concomitantly, 125 I activity was increased in urine and feces collected from PFOS-treated rats. (3) During the 8 days of dosing with PFOS, TSH was not elevated in male rats, while TT4 and TT3 were decreased. Pituitary response to TRH-mediated TSH release was not diminished after 8-daily oral doses of PFOS. CONCLUSIONS: These findings suggest that oral dosing in rats with PFOS results in transiently increased tissue availability of the thyroid hormones and turnover of T4 with a resulting reduction in serum TT4. PFOS does not induce a classical hypothyroid state under dosing conditions employed nor does it alter HPT activities.

An interlaboratory study of perfluorinated alkyl compound levels in human plasma.

We conducted an interlaboratory study which differed from the typical study of this type because of its emphasis on comparing intralaboratory variability in results. We sent specimens to six laboratories experienced in the analysis of perfluorinated alkyl compounds in blood matrices and that use stringent procedures to control and assure accuracy and precision. Each received an identical set of 60 plasma specimens that were analyzed in six completely independent batches. Split specimens were included so that within- and between-batch coefficients of variation could be calculated. All laboratories used liquid chromatography-tandem mass spectrometry (LC-MS/MS). The concentrations of perfluorooctanesulfonate (PFOS), perfluorooctanoate (PFOA), and perfluorohexanesulfonate (PFHxS) measured in the specimens in general showed a high level of agreement, although in some cases the agreement was only moderate. The average within- and between-batch coefficient of variation for PFOS was 9.1% and 9.3%; for PFOA was 14.5% and 14.5%; and for PFHxS was 14.5% and 17.0%. The recent availability of labeled internal standards, among other advances, has facilitated improvement in the accuracy and precision of the assays. Considering the degree of between-subject variation in levels among people in background-exposed populations, the results indicate that biomarker-based epidemiologic studies of associations with health could have reasonable precision.

Reproductive and developmental toxicity of potassium perfluorohexanesulfonate in CD-1 mice.

Potassium perfluorohexanesulfonate (K+PFHxS) was evaluated for reproductive/developmental toxicity in CD-1 mice. Up to 3 mg/kg-d K+PFHxS was administered (n = 30/sex/group) before mating, for at least 42 days in F0 males, and for F0 females, through gestation and lactation. F1 pups were directly dosed with K+PFHxS for 14 days after weaning. There was an equivocal decrease in live litter size at 1 and 3 mg/kg-d, but the pup-born-to-implant ratio was unaffected. Adaptive hepatocellular hypertrophy was observed, and in 3 mg/kg-d F0 males, it was accompanied by concomitant decreased serum cholesterol and increased alkaline phosphatase. There were no other toxicologically significant findings on reproductive parameters, hematology/clinical pathology/TSH, neurobehavioral effects, or histopathology. There were no treatment-related effects on postnatal survival, development, or onset of preputial separation or vaginal opening in F1 mice. Consistent with previous studies, our data suggest that the potency of PFHxS is much lower than PFOS in rodents.

Half-life of serum elimination of perfluorooctanesulfonate,perfluorohexanesulfonate, and perfluorooctanoate in retired fluorochemical production workers.

BACKGROUND: The presence of perfluorooctanesulfonate (PFOS), perfluorohexanesulfonate (PFHS), and perfluorooctanoate (PFOA) has been reported in humans and wildlife. Pharmacokinetic differences have been observed in laboratory animals. OBJECTIVE: The purpose of this observational study was to estimate the elimination half-life of PFOS, PFHS, and PFOA from human serum. METHODS: Twenty-six (24 male, 2 female) retired fluorochemical production workers, with no additional occupational exposure, had periodic blood samples collected over 5 years, with serum stored in plastic vials at -80 degrees C. At the end of the study, we used HPLC-mass spectrometry to analyze the samples, with quantification based on the ion ratios for PFOS and PFHS and the internal standard (18)O(2)-PFOS. For PFOA, quantitation was based on the internal standard (13)C(2)-PFOA. RESULTS: THE ARITHMETIC MEAN INITIAL SERUM CONCENTRATIONS WERE AS FOLLOWS: PFOS, 799 ng/mL (range, 145-3,490); PFHS, 290 ng/mL (range, 16-1,295); and PFOA, 691 ng/mL (range, 72-5,100). For each of the 26 subjects, the elimination appeared linear on a semi-log plot of concentration versus time; therefore, we used a first-order model for estimation. The arithmetic and geometric mean half-lives of serum elimination, respectively, were 5.4 years [95% confidence interval (CI), 3.9-6.9] and 4.8 years (95% CI, 4.0-5.8) for PFOS; 8.5 years (95% CI, 6.4-10.6) and 7.3 years (95% CI, 5.8-9.2) for PFHS; and 3.8 years (95% CI, 3.1-4.4) and 3.5 years (95% CI, 3.0-4.1) for PFOA. CONCLUSIONS: Based on these data, humans appear to have a long half-life of serum elimination of PFOS, PFHS, and PFOA. Differences in species-specific pharmacokinetics may be due, in part, to a saturable renal resorption process.

Negative bias from analog methods used in the analysis of free thyroxine in rat serum containing perfluorooctanesulfonate (PFOS).

Decreases in serum total thyroxine (TT4) and free thyroxine (FT4) without a compensatory rise in thyroid stimulating hormone (thyrotropin or TSH) or histological changes of the thyroid have been observed in studies with perfluorooctanesulfonate (PFOS) treatments in rats. Prior observations do not fit the clinical profile of a hypothyroid state. PFOS is known to compete with fatty acids for albumin binding, and serum free fatty acids (FFA) are known to interfere with FT4 measurement using analog methods due to competition for protein binding. Therefore, we hypothesized that measured decreases in serum FT4 by analog methods in the presence of PFOS were due to carrier protein binding interference. We compared FT4 analog assay methods with a reference method using equilibrium dialysis (ED-RIA) for FT4 measurement in rat sera in vitro and in vivo. We also measured hepatic malic enzyme mRNA transcripts and activity as a marker for hepatic thyroid hormone response. PFOS did not reduce serum TT4 and FT4 in vitro at concentrations up to 200 microM. After three daily 5mg/kg oral doses of potassium PFOS to female rats, serum TSH and FT4 by ED-RIA were unchanged (although FT4 determined by two common analog methods was decreased), and malic enzyme was not suppressed. These data suggest that prior reports of reduced free thyroid hormone in the presence of PFOS were due to negative bias in analog methods and that short-term PFOS treatment does not suppress the physiological thyroid status in rats. A reference method such as ED-RIA should be used for determination of serum FT4 in the presence of PFOS.

Preliminary evidence of a decline in perfluorooctanesulfonate (PFOS) and perfluorooctanoate (PFOA) concentrations in American Red Cross blood donors.

The purpose of this pilot study was to determine whether perfluorooctanesulfonate (PFOS,C(8)F(17)SO(3)(-)) and perfluorooctanoate (PFOA,C(7)F(15)CO(2)(-)) concentrations in American Red Cross blood donors from Minneapolis-St. Paul, Minnesota have declined after the 2000-2002 phase-out of perfluorooctanesulfonyl-fluoride (POSF, C(8)F(17)SO(2)F)-based materials by the primary global manufacturer, 3M Company. Forty donor plasma samples, categorized by age and sex, were collected in 2005, and PFOS and PFOA concentrations were compared to 100 (non-paired) donor serum samples collected in 2000 from the same general population that were analyzed at the time using ion-pair extraction methods with tetrahydroperfluorooctanesulfonate as an internal standard. Eleven of the 100 samples originally collected were reanalyzed with present study methods that involved (13)C- labeled PFOA spiked into the donor samples, original samples, control human plasma, and the calibration curve prior to extraction, and was used as a surrogate to monitor extraction efficiency. Quantification was performed by high performance liquid chromatography tandem mass spectrometry methods. Among the 100 serum samples analyzed for PFOS, the geometric mean was 33.1 ng ml(-1) (95% CI 29.8-36.7) in 2000 compared to 15.1 ng ml(-1) (95% CI 13.3-17.1) in 2005 (p<0.0001) for the 40 donor plasma samples. The geometric mean concentration for PFOA was 4.5 ng ml(-1) (95% CI 4.1-5.0) in 2000 compared to 2.2 ng ml(-1) (95% CI 1.9-2.6) in 2005 (p<0.0001). The decrease was consistent across donors' age and sex. To confirm these preliminary findings, additional sub-sets of year 2000 samples will be analyzed, and a much larger biomonitoring study of other locations is planned.

Effects of perfluorooctane sulfonate on mallard and northern bobwhite quail exposed chronically via the diet.

Adult mallard ducks and northern bobwhite quail were exposed to 0, 10, 50, or 150mg perfluorooctane sulfonate (PFOS)/kg in the diet for up to 21 weeks. Adult health, body and liver weight, feed consumption, gross morphology and histology of body organs, and reproduction were examined. Due to mortality, birds exposed to 50 or 150mg PFOS/kg feed were terminated by Week 7. In quail, the lowest observable adverse effect level (LOAEL) was 10mg PFOS/kg feed based on decreased survivorship of 14-day-old quail offspring. For adult female quail fed 10mg/kg feed, there was a slight but statistically significantly PFOS-related increase in liver weight when compared to controls. When liver weight was normalized to body weight, the statistically significant differences were still observed indicating that PFOS affected liver size. However, no other pathological effects were observed livers of quail from this treatment group which suggests that this enlargement may have been an adaptive response. For adult mallards, no treatment-related effects on feed consumption, body or liver weight, growth, or reproductive performance were observed. There was a slightly greater incidence of small testes (length) in adult male mallards and quail exposed to 10mg PFOS/kg, feed when compared to controls. However, spermatogenesis was not affected and there was no effect on the rates of egg fertilization. Due to transfer to eggs, concentrations of PFOS measured in the liver and blood at study termination were greater in male birds than female birds.

Toxicological response of Sprague Dawley rats from inhalation exposure to perfluorooctane sulfonyl fluoride (POSF).

Perfluorooctane sulfonyl fluoride (POSF) was a volatile starting material in the production of perfluorooctane sulfonate (PFOS), a stable surfactant that has been extensively studied due to its ubiquitous environmental distribution and slow clearance in humans. Because the inhalation toxicity of POSF on repeated exposure has not been previously reported, the current study evaluated the inhalation toxicity of POSF at 30, 100, and 300ppm (v/v) in rats for up to 13 weeks with a four-week recovery period. The extent of PFOS formation was also measured because POSF hydrolyzed to form PFOS. In addition, detailed urinalysis and examination of the urinary bladder were included to determine if factors associated with the development of bladder cancer were present. Exposure to POSF at 300ppm was associated with reduction in body weight-gain, necrosis of laryngeal cartilage, increased lung and bronchi weight with septal thickening, and changes in alveolar macrophages. The microscopic observations in larynx and lung are consistent with likely hydrolysis of POSF to form hydrogen fluoride (HF). Exposure to POSF at 100 and 300ppm was associated with increased relative liver weight, hepatocellular hypertrophy (except for females exposed to 100ppm POSF), and lowering of serum cholesterol (male only). After 13 weeks of exposure to 30, 100, or 300ppm POSF, serum PFOS concentration approximated 7, 35, or 100µg/mL, respectively. Approximately 0.1% of inhaled POSF was converted to PFOS. No changes indicative of bladder effects were observed in these rats exposed to POSF at any dose.

Organic Anion Transporting Polypeptides Contribute to the Disposition of Perfluoroalkyl Acids in Humans and Rats.

Perfluoroalkyl sulfonates (PFSAs) such as perfluorohexane sulfonate (PFHxS) and perfluorooctane sulfonate (PFOS) have very long serum elimination half-lives in humans, and preferentially distribute to serum and liver. The enterohepatic circulation of PFHxS and PFOS likely contributes to their extended elimination half-lives. We previously demonstrated that perfluorobutane sulfonate (PFBS), PFHxS, and PFOS are transported into hepatocytes both in a sodium-dependent and a sodium-independent manner. We identified Na+/taurocholate cotransporting polypeptide (NTCP) as the responsible sodium-dependent transporter. Furthermore, we demonstrated that the human apical sodium-dependent bile salt transporter (ASBT) contributes to the intestinal reabsorption of PFOS. However, so far no sodium-independent uptake transporters for PFSAs have been identified in human hepatocytes or enterocytes. In addition, perfluoroalkyl carboxylates (PFCAs) with 8 and 9 carbons were shown to preferentially distribute to the liver of rodents; however, no rat or human liver uptake transporters are known to transport these PFCAs. Therefore, we tested whether PFBS, PFHxS, PFOS, and PFCAs with 7-10 carbons are substrates of organic anion transporting polypeptides (OATPs). We used CHO and HEK293 cells to demonstrate that human OATP1B1, OATP1B3, and OATP2B1 can transport PFBS, PFHxS, PFOS, and the 2 PFCAs (C8 and C9). In addition, we show that rat OATP1A1, OATP1A5, OATP1B2, and OATP2B1 transport all 3 PFSAs. In conclusion, our results suggest that besides NTCP and ASBT, OATPs also are capable of contributing to the enterohepatic circulation and extended human serum elimination half-lives of the tested perfluoroalkyl acids.