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Arsenic is a ubiquitous contaminant, with typical human dietary intake below 1 μg/kg bw/d and extreme drinking water exposures up to ∼50 μg/kg bw/d. The formation and binding of trivalent metabolites are central to arsenic toxicity and strong human evidence suggests special concern for early life exposures in the etiology of adult diseases, especially cancer. This study measured the metabolism and disposition of arsenite in neonatal mice to understand the role of maturation in metabolic activation and detoxification of arsenic. Many age-related differences were observed after gavage administration of arsenite, with consistent evidence in blood and tissues for higher exposures to trivalent arsenic species in neonatal mice related to the immaturity of metabolic and/or excretory functions. The evidence for greater tissue binding of arsenic species in young mice is consistent with enhanced susceptibility to toxicity based on metabolic and toxicokinetic differences alone. Lactational transfer from arsenite-dosed dams to suckling mice was minimal, based on no dosing-related changes in the levels of arsenic species in pup blood or milk collected from the dams. Animal models evaluating whole-life exposure to inorganic arsenic must use direct dosing in early neonatal life to predict accurately potential toxicity from early life exposures in children.

Chronic exposure to inorganic arsenic (iAs), a contaminant of water and food supplies, is associated with many adverse health effects. A notable feature of iAs metabolism is sequential methylation reactions which produce mono- and di-methylated arsenicals that can contain arsenic in either the trivalent (III) or pentavalent (V) valence states. Because methylated arsenicals containing trivalent arsenic are more potent toxicants than their pentavalent counterparts, the ability to distinguish between the +3 and +5 valence states is a crucial property for physiologically based pharmacokinetic (PBPK) models of arsenicals to possess if they are to be of use in risk assessment. Unfortunately, current analytic techniques for quantifying arsenicals in tissues disrupt the valence state; hence, pharmacokinetic studies in animals, used for model calibration, only reliably provide data on the sum of the +3 and +5 valence forms of a given metabolite. In this paper we show how mathematical modeling can be used to overcome this obstacle and present a PBPK model for the dimethylated metabolite of iAs, which exists as either dimethylarsinous acid, (CH(3))(2)As(III)OH (abbreviated DMA(III)) or dimethylarsinic acid, (CH(3))(2)As(V)(O)OH (abbreviated DMA(V)). The model distinguishes these two forms and sets a lower bound on how much of an organ’s DMA burden is present in the more reactive and toxic trivalent valence state. We conjoin the PBPK model to a simple model for DMA(III)-induced oxidative stress in liver and use this extended model to predict cytotoxicity in liver in response to the high oral dose of DMA(V). The model incorporates mechanistic details derived from in vitro studies and is iteratively calibrated with lumped-valence-state PK data for intravenous or oral dosing with DMA(V). Model formulation leads us to predict that orally administered DMA(V) undergoes extensive reduction in the gastrointestinal (GI) tract to the more toxic trivalent DMA(III).

Arsenic is a ubiquitous contaminant, with typical dietary intake below 1 μg/kg bw/d and drinking water exposures up to 50 μg/kg bw/d. Arsenic exposures are associated with human diseases and doses of toxicological concern are similar to typical dietary intake. Metabolism of arsenite to dimethylarsinate (DMAV) by arsenite-3-methyltransferase (As3MT) promotes clearance, but also generates reactive trivalent intermediates that bind extensively to cellular thiols. This study measured pentavalent and trivalent arsenic species in blood and tissues after oral and intravenous administration of arsenite (50 μg/kg bw). After oral administration, the intestine and liver contained elevated levels of AsIII and MMAIII, relative to erythrocytes, lung, and kidney, suggesting incomplete conversion to DMA during first-pass metabolism. However, blood concentrations of the predominant species, DMA, were similar for oral and intravenous dosing. While all tissues examined contained DMAIII, muscle, brain, and plasma had undetectable levels of MMAIII. Tissue levels of arsenic species were similar following intravenous vs. oral administration, except lower in the intestine. The results confirm the role of metabolism in producing fluxes of putatively toxic trivalent arsenic intermediates. Tissue dosimetry suggests that the intestine, liver, lung, and kidney could be more susceptible to effects of bound arsenic, relative to muscle and brain.

Polyfluorinated compounds (PFCs) are authorized for use as greaseproofing agents in food contact paper. As C8-PFCs (8-carbons) are known to accumulate in tissues, shorter-chain C6-PFCs (6-carbons) have replaced C8-PFCs in many food contact applications. However, the potential of C6-PFCs for human biopersistence has not been fully evaluated. For the first time, we provide internal exposure estimates to key metabolites of 6:2 fluorotelomer alcohol (6:2 FTOH), a monomeric component of C6-PFCs, to extend our understanding of exposure beyond estimates of external exposure. Pharmacokinetic data from published rat and human studies on 6:2 FTOH were used to estimate clearance and area under the curve (AUC) for its metabolites: 5:3 fluorotelomer carboxylic acid (5:3 A), perfluorohexanoic acid (PFHxA) and perfluoroheptanoic acid (PFHpA). Internal exposure to 5:3 A was the highest of evaluated metabolites across species and it had the slowest clearance. Additionally, 5:3 A clearance decreased with increasing 6:2 FTOH exposure. Our analysis provides insight into association of increased internal 5:3 A exposure with high biopersistence potential of 6:2 FTOH. Our results identify 5:3 A as an important biomarker of internal 6:2 FTOH exposure for use in biomonitoring studies, and are potentially useful for toxicological assessment of chronic dietary 6:2 FTOH exposure.

Oseltamivir is an antiviral drug approved to treat influenza in humans. Although the dosing regimen of this drug is well established for non-pregnant patients, it is not clear if the significant physiological alterations associated with pregnancy affect the pharmacokinetics of oseltamivir and, thus, warrant different dosing regimens to assure efficacy. In this study, we investigated the suitability of rhesus macaques as an animal model for studying oseltamivir pharmacokinetics during all trimesters of pregnancy in comparison to pre-pregnant conditions. Specifically, we compared the pharmacokinetics of oseltamivir and its pharmacologically active metabolite oseltamivir carboxylate in rhesus monkeys after intravenous and nasogastric administration of 2.5 mg oseltamivir phosphate/kg body weight given prior to and during the first, second, and third trimesters of pregnancy. Pregnancy had only a modest effect upon the pharmacokinetic parameters of oseltamivir and oseltamivir carboxylate. Monkeys treated intravenously in the third trimester had a reduction in Vd and CL, compared to non-pregnant monkeys. These changes did not occur in the other two trimesters. Pregnant monkeys treated intravenously had 20–25% decrease in AUC0-∞ of oseltamivir carboxylate and a corresponding increase in Vd and CL. Pregnant monkeys treated nasogastrically with oseltamivir phosphate demonstrated a pattern that recapitulated intravenous dosing. Taken together these data indicate that rhesus monkeys are an acceptable model for studying drug-pregnancy interactions.

Physiologically-based pharmacokinetic (PBPK) modeling analysis does not stand on its own for regulatory purposes but is a robust tool to support drug/chemical safety assessment. While the development of PBPK models have grown steadily since their emergence, only a handful of models have been accepted to support regulatory purposes due to obstacles such as the lack of a standardized template for reporting PBPK analysis. Here, we expand the existing guidances designed for pharmaceutical applications by recommending additional elements that are relevant to environmental chemicals. This harmonized reporting template can be adopted and customized by public health agencies receiving PBPK model submission, and it can also serve as general guidance for submitting PBPK-related studies for publication in journals or other modeling sharing purposes. The current effort represents one of several ongoing collaborations among the PBPK modeling and risk assessment communities to promote, when appropriate, incorporating PBPK modeling to characterize the influence of pharmacokinetics on safety decisions made by regulatory agencies.

Renal elimination and the resulting clearance of perfluorooctanoic acid (PFOA) from the serum exhibit pronounced sex differences in the adult rat. The literature suggests that this is largely due to hormonally regulated expression of organic anion transporters (OATs) on the apical and basolateral membranes of the proximal tubule cells that facilitate excretion and reabsorption of PFOA from the filtrate into the blood. Previously developed PBPK models of PFOA exposure in the rat have not been parameterized to specifically account for transporter-mediated renal elimination. We developed a PBPK model for PFOA in the male and female rat to explore the role of Oat1, Oat3, and Oatp1a1 in sex-specific renal reabsorption and excretion of PFOA. Descriptions of the kinetic behavior of these transporters were extrapolated from in vitro studies and the model was used to simulate time-course serum, liver, and urine data for intravenous (IV) and oral exposures in both sexes. Model predicted concentrations of PFOA in the liver, serum, and urine showed good agreement with experimental data for both the male and female rat indicating that in vitro derived physiological descriptions of transporter-mediated renal reabsorption can successfully predict sex-dependent excretion of PFOA in the rat. This study supports the hypothesis that sex-specific serum half-lives for PFOA are largely driven by expression of transporters in the kidney and contributes to the development of PBPK modeling as a tool for evaluating the role of transporters in renal clearance.

2-Phenoxyethanol (PhE) has been shown to induce hepatotoxicity, renal toxicity, and hemolysis at dosages ≥ 400 mg/kg/day in subchronic and chronic studies in multiple species. To reduce uncertainty associated with interspecies extrapolations and to evaluate the margin of exposure (MOE) for use of PhE in cosmetics and baby products, a physiologically-based pharmacokinetic (PBPK) model of PhE and its metabolite 2-phenoxyacetic acid (PhAA) was developed. The PBPK model incorporated key kinetic processes describing the absorption, distribution, metabolism and excretion of PhE and PhAA following oral and dermal exposures. Simulations of repeat dose rat studies facilitated the selection of systemic AUC as the appropriate dose metric for evaluating internal exposures to PhE and PhAA in rats and humans. Use of the PBPK model resulted in refinement of the total default UF for extrapolation of the animal data to humans from 100 to 25. Based on very conservative assumptions for product composition and aggregate product use, model-predicted exposures to PhE and PhAA resulting from adult and infant exposures to cosmetic products are significantly below the internal dose of PhE observed at the NOAEL dose in rats. Calculated MOEs for all exposure scenarios were above the PBPK-refined UF of 25.

Manufacturing of perfluorooctanoic acid (PFOA), a synthetic chemical with a long half-life in humans, peaked between 1970 and 2002, and has since diminished. In the United States, PFOA is detected in the blood of >99% of people tested, but serum concentrations have decreased since 1999. Much is known about exposure to PFOA in drinking water; however, the impact of non-drinking water PFOA exposure on serum PFOA concentrations is not well characterized. The objective of this research is to apply physiologically based pharmacokinetic (PBPK) modeling and Monte Carlo analysis to evaluate the impact of historic non-drinking water PFOA exposure on serum PFOA concentrations. In vitro to in vivo extrapolation was utilized to inform descriptions of PFOA transport in the kidney. Monte Carlo simulations were incorporated to evaluate factors that account for the large inter-individual variability of serum PFOA concentrations measured in individuals from North Alabama in 2010 and 2016, and the Mid-Ohio River Valley between 2005 and 2008. Predicted serum PFOA concentrations were within two-fold of experimental data. With incorporation of Monte Carlo simulations, the model successfully tracked the large variability of serum PFOA concentrations measured in populations from the Mid-Ohio River Valley. Simulation of exposure in a population of 45 adults from North Alabama successfully predicted 98% of individual serum PFOA concentrations measured in 2010 and 2016, respectively, when non-drinking water ingestion of PFOA exposure was included. Variation in serum PFOA concentrations may be due to inter-individual variability in the disposition of PFOA and potentially elevated historical non-drinking water exposures.

Many physiologically based pharmacokinetic (PBPK) models for environmental chemicals, drugs, and nanomaterials have been developed to aid risk and safety assessments using acslX. However, acslX has been rendered sunset since November 2015. Alternative modeling tools and tutorials are needed for future PBPK applications. This forum article aimed to: (1) demonstrate the performance of 4 PBPK modeling software packages (acslX, Berkeley Madonna, MATLAB, and R language) tested using 2 existing models (oxytetracycline and gold nanoparticles); (2) provide a tutorial of PBPK model code conversion from acslX to Berkeley Madonna, MATLAB, and R language; (3) discuss the advantages and disadvantages of each software package in the implementation of PBPK models in toxicology, and (4) share our perspective about future direction in this field. Simulation results of plasma/tissue concentrations/amounts of oxytetracycline and gold from different models were compared visually and statistically with linear regression analyses. Simulation results from the original models were correlated well with results from the recoded models, with time-concentration/amount curves nearly superimposable and determination coefficients of 0.86-1.00. Step-by-step explanations of the recoding of the models in different software programs are provided in the Supplementary Data. In summary, this article presents a tutorial of PBPK model code conversion for a small molecule and a nanoparticle among 4 software packages, and a performance comparison of these software packages in PBPK model implementation. This tutorial helps beginners learn PBPK modeling, provides suggestions for selecting a suitable tool for future projects, and may lead to the transition from acslX to alternative modeling tools.

Our previous report on pharmacokinetic (PK) evaluation of 6:2 fluorotelomer alcohol (6:2 FTOH) examined the biopersistence potential of its metabolites based on data published from single inhalation and occupational 6:2 FTOH exposure studies. We calculated internal exposure estimates of three key metabolites of 6:2 FTOH, of which 5:3 fluorotelomer carboxylic acid (5:3 acid) had the highest internal exposure and the slowest clearance. No oral repeated 6:2 FTOH exposure data were available at the time to fully characterize the biopersistence potential of the metabolite 5:3 acid. We recently received additional data on 6:2 FTOH and 5:3 acid, which included a 90-day toxicokinetic study report on repeated oral 6:2 FTOH exposure to rats. We reviewed the study and analyzed the reported 5:3 acid concentrations in plasma, liver, and fat using one-compartment PK modeling and calculated elimination rate constants (kel), elimination half-lives (t1/2) and times to steady state (tss) of 5:3 acid at three 6:2 FTOH doses. Our results showed that tss of 5:3 acid in plasma and evaluated tissues were approximately close to 1 year, such that the majority of highest values were observed at the lowest 6:2 FTOH dose, indicating its association with the biopersistence of 6:2 FTOH. The results of our PK analysis are the first to characterize biopersistence potential of the 5:3 acid after repeated oral exposure to the parent compound 6:2 FTOH based on steady state PK parameters, and therefore, may have an impact on future study designs when conducting toxicity assays for such compounds.

The widespread use and high abuse liability of tobacco products has received considerable public health attention, in particular for youth, who are vulnerable to nicotine addiction. In this study, adult and adolescent squirrel monkeys were used to evaluate age-related metabolism and pharmacokinetics of nicotine after intravenous administration. A physiologically-based pharmacokinetic (PBPK) model was created to characterize the pharmacokinetic behaviors of nicotine and its metabolites, cotinine, trans-3'-hydroxycotinine (3'-OH cotinine), and trans-3'-hydroxycotinine glucuronide (3'-OH cotinine glucuronide) for both adult and adolescent squirrel monkeys. The PBPK nicotine model was first calibrated for adult squirrel monkeys utilizing in vitro nicotine metabolic data, plasma concentration-time profiles and cumulative urinary excretion data for nicotine and metabolites. Further model refinement was conducted when the calibrated adult model was scaled to the adolescents, because adolescents appeared to clear nicotine and cotinine more rapidly relative to adults. More specifically, the resultant model parameters representing systemic clearance of nicotine and cotinine for adolescent monkeys were approximately two- to three-fold of the adult values on a per body weight basis. The nonhuman primate PBPK model in general captured experimental observations that were used for both model calibration and evaluation, with acceptable performance metrics for precision and bias. The model also identified differences in nicotine pharmacokinetics between adolescent and adult nonhuman primates which might also be present in humans.

The use of toxicokinetic (TK) data is becoming more prevalent in the evaluation of food ingredient safety as more TK information is being incorporated in safety data packages. Data demonstrating “1) the extent of absorption, 2) tissue distribution, 3) pathways and rates of metabolism, and 4) rate(s) of elimination” of food ingredients and their metabolites of intermediate and high toxicological potential may be useful for planning and designing toxicity studies, selecting doses for toxicity studies, addressing species differences, and understanding the potential modes of action to evaluate their safety. TK data reported in the literature or generated from mechanistic TK studies can be analyzed using mathematical methods, including compartment and noncompartment TK methods, whose predictions can enhance interpretation of observed effects. Because of recent advancements, several approaches have been developed to improve sensitivity of analyses of available TK data and reduce uncertainty for evaluating safety of food ingredients. An example of advanced TK methods is physiologically-based TK (PBTK) modeling that incorporates physiological/biochemical parameters into a TK framework to predict internal exposure. In this review, we discuss the utility of some TK methods and explore their relevance and potential value for food ingredient safety evaluation. We also describe the strengths and limitations of these TK methods and discuss current challenges and opportunities for expanding their application for evaluating safety of food ingredients. This review represents a state of science report, and not a guidance document, on the utility and relevance of TK methods for the safety evaluation of food ingredients.

Bisphenol A (BPA) has received considerable attention throughout the last decade due to its widespread use in consumer products. For the first time a physiologically based pharmacokinetic (PBPK) model was developed in neonatal and adult rats to quantitatively evaluate age-dependent pharmacokinetics of BPA and its phase II metabolites. The PBPK model was calibrated in adult rats using studies on BPA metabolism and excretion in the liver and gastrointestinal tract, and pharmacokinetic data with BPA in adult rats. For immature rats the hepatic and gastrointestinal metabolism of BPA was inferred from studies on the maturation of phase II enzymes coupled with serum time course data in pups. The calibrated model predicted the measured serum concentrations of BPA and BPA conjugates after administration of 100μg/kg of d6-BPA in adult rats (oral gavage and intravenous administration) and postnatal days 3, 10, and 21 pups (oral gavage). The observed age-dependent BPA serum concentrations were partially attributed to the immature metabolic capacity of pups. A comparison of the dosimetry of BPA across immature rats and monkeys suggests that dose adjustments would be necessary to extrapolate toxicity studies from neonatal rats to infant humans.

Bromate (BrO3 −), a by-product of ozonation of drinking water, induces nephrotoxicity in male rats at much lower doses than in female rats. This difference appears to be related to the development of α-2u-globulin nephropathy in males. To determine sex-dependent changes in mRNA and protein expression in the renal cortex attributable to α-2u-globulin nephropathy, we performed microarray and immunohistochemical analyses in proximal renal tubules of male and female F344 rats treated with KBrO3 for 28 days. Particular attention was paid to molecular biomarkers of renal tubular injury. Microarray analysis of male and female rats treated with BrO3 − at low doses (125 mg/L KBrO3) displayed marked sex-dependent changes in renal gene expression. The greatest differences were seen in genes encoding for cellular differentiation, apoptosis, ion transport, and cell proliferation. Differences by sex were especially prominent for the cell cycle checkpoint gene p21, the renal injury protein Kim-1, and the kidney injury and cancer biomarker protein osteopontin. Dose-related nephrotoxicity, assessed by hematoxylin and eosin staining, was greater in males compared to female rats, as was cellular proliferation, assessed by bromodeoxyuridine staining. The fraction of proximal renal cells with elevated 8-oxodeoxyguanosine (8-OH-dG) was only increased at the high dose and did not differ by sex. Dose-dependent increases in the expression of osteopontin were detected immunohistochemically only in male rats and were localized in proximal tubule cells. Similarly, BrO3 − treatment increased clusterin and Kim-1 staining in the proximal tubules; however, staining for these proteins did not differ appreciably between males and females. These data demonstrate both qualitative and quantitative differences in the response of male versus female kidneys to BrO3 −-treatment. These sex-dependent effects likely contribute to renal carcinogenesis of BrO3 − in the male rat.

Bromate (BrO(3)(-)) is a ubiquitous by-product of using ozone to disinfect water containing bromide (Br(-)). The reactivity of BrO(3)(-) with biological reductants suggests that its systemic absorption and distribution to target tissues may display non-linear behavior as doses increase. The intent of this study is to determine the extent to which BrO(3)(-) is systemically bioavailable via oral exposure and broadly identify its pathways of degradation. In vitro experiments of BrO(3)(-) degradation in rat blood indicate a rapid initial degradation immediately upon addition that is >98% complete at concentrations up to 66μM in blood. As initial concentrations are increased, progressively lower fractions are lost prior to the first measurement. Secondary to this initial loss, a slower and predictable first order degradation rate was observed (10%/min). Losses during both phases were accompanied by increases in Br(-) concentrations indicating that the loss of BrO(3)(-) was due to its reduction. In vivo experiments were conducted using doses of BrO(3)(-) ranging from 0.077 to 15.3mg/kg, administered intravenously (IV) or orally (gavage) to female F344 rats. The variable nature and uncertain source of background concentrations of BrO(3)(-) limited derivation of terminal half-lives, but the initial half-life was approximately 10min for all dose groups. The area under the curve (AUC) and peak concentrations (C(t=5')) were linearly related to IV dose up to 0.77mg/kg; however, disproportionate increases in the AUC and C(t=5') and a large decrease in the volume of distribution was observed when IV doses of 1.9 and 3.8mg/kg were administered. The average terminal half-life of BrO(3)(-) from oral administration was 37min, but this was influenced by background levels of BrO(3)(-) at lower doses. With oral doses, the AUC and C(max) increased linearly with dose up to 15.3mgBrO(3)(-)/kg. BrO(3)(-) appeared to be 19-25% bioavailable without an obvious dose-dependency between 0.077 and 1.9mg/kg. The urinary elimination of BrO(3)(-) and Br(-) was measured from female F344 rats for four days following administration of single doses of 8.1mgKBrO(3)/kg and for 15 days after a single dose of 5.0mgKBr/kg. BrO(3)(-) elimination was detected over the first 12h, but Br(-) elimination from BrO(3)(-) over the first 48h was 18% lower than expected based on that eliminated from an equimolar dose of Br(-) (15.5±1.6 vs. 18.8±1.2μmol/kg, respectively). The cumulative excretion of Br(-) from KBr vs. KBrO(3) was equivalent 72h after administration. The recovery of unchanged administered BrO(3)(-) in the urine ranged between 6.0 and 11.3% (creatinine corrected) on the 27th day of treatment with concentrations of KBrO(3) of 15, 60, and 400mg/L of drinking water. The recovery of total urinary bromine as Br(-)+BrO(3)(-) ranged between 61 and 88%. An increase in the fraction of the daily BrO(3)(-) dose recovered in the urine was observed at the high dose to both sexes. The deficit in total bromine recovery raises the possibility that some brominated biochemicals may be produced in vivo and more slowly metabolized and eliminated. This was supported by measurements of dose-dependent increases of total organic bromine (TOBr) that was eliminated in the urine. The role these organic by-products play in BrO(3)(-)-induced cancer remains to be established.

Partition coefficients (PCs) are used in physiologically based pharmacokinetic (PBPK) models to estimate the free concentration of a chemical in specific blood or organs. Biological PC(tissue:blood) (tissue to blood) values were determined for a series of nonvolatile herbicides, insecticides, and fungicides in liver, brain, skin, fat, kidneys, and muscle of male Sprague-Dawley rats using two different analytical methods. The free phase concentration (in phosphate-buffered saline) of a given chemical was measured in the presence and absence of tissue (including blood) and used to calculate the PC, defined as the ratio of the concentration of the chemical in saline to the concentration in the tissue. PCs were determined for 13 compounds with aqueous solubility ranging from 20 to 4100 mg/L, molecular weights from 187.3 to 342.2 g/mol, and log K (ow) values from -0.18 to 3.9. An ultrafiltration high-performance liquid chromatography (HPLC) method was implemented for compounds with log K (ow) near 0.1 or less and a negligible depletion solid-phase microextraction (nd-SPME) method for compounds with higher log K (ow). PC(tissue:saline) coefficients of variation were 0.13 (n = 3 compounds) on average for the HPLC method and 0.29 (n = 10 compounds) for the nd-SPME method. Presented here is one of the most comprehensive data sets of biological partition coefficients for herbicides, insecticides, and fungicides.

The pharmacokinetic behavior of the majority of jet fuel constituents has not been previously described in the framework of a physiologically based pharmacokinetic (PBPK) model for inhalation exposure. Toxic effects have been reported in multiple organ systems, though exposure methods varied across studies, utilizing either vaporized or aerosolized fuels. The purpose of this work was to assess the pharmacokinetics of aerosolized and vaporized fuels, and develop a PBPK model capable of describing both types of exposures. To support model development, n-tetradecane and n-octane exposures were conducted at 89 mg/m(3) aerosol+vapor and 1000-5000 ppm vapor, respectively. Exposures to JP-8 and S-8 were conducted at ~900-1000 mg/m(3), and ~200 mg/m(3) to a 50:50 blend of both fuels. Sub-models were developed to assess the behavior of representative constituents and grouped unquantified constituents, termed "lumps", accounting for the remaining fuel mass. The sub-models were combined into the first PBPK model for petroleum and synthetic jet fuels. Inhalation of hydrocarbon vapors was described with simple gas-exchange assumptions for uptake and exhalation. For aerosol droplets systemic uptake occurred in the thoracic region. Visceral tissues were described using perfusion and diffusion-limited equations. The model described kinetics at multiple fuel concentrations, utilizing a chemical "lumping" strategy to estimate parameters for fractions of speciated and unspeciated hydrocarbons and gauge metabolic interactions. The model more accurately simulated aromatic and lower molecular weight (MW) n-alkanes than some higher MW chemicals. Metabolic interactions were more pronounced at high (~2700-1000 mg/m(3)) concentrations. This research represents the most detailed assessment of fuel pharmacokinetics to date.

A physiologically based pharmacokinetic (PBPK) model was developed for bisphenol A (BPA) in adult rhesus monkeys using intravenous (iv) and oral bolus doses of 100 μg d6-BPA/kg (Doerge et al., 2010). This calibrated PBPK adult monkey model for BPA was then evaluated against published monkey kinetic studies with BPA. Using two versions of the adult monkey model based on monkey BPA kinetic data from Doerge et al. (2010) and Taylor et al. (2011), the aglycone BPA pharmacokinetics were simulated for human oral ingestion of 5 mg d16-BPA per person (Völkel et al., 2002). Völkel et al. were unable to detect the aglycone BPA in plasma, but were able to detect BPA metabolites. These human model predictions of the aglycone BPA in plasma were then compared to previously published PBPK model predictions obtained by simulating the Völkel et al. kinetic study. Our BPA human model, using two parameter sets reflecting two adult monkey studies, both predicted lower aglycone levels in human serum than the previous human BPA PBPK model predictions. BPA was metabolized at all ages of monkey (PND 5 to adult) by the gut wall and liver. However, the hepatic metabolism of BPA and systemic clearance of its phase II metabolites appear to be slower in younger monkeys than adults. The use of the current non-human primate BPA model parameters provides more confidence in predicting the aglycone BPA in serum levels in humans after oral ingestion of BPA.

A physiologically based pharmacokinetic (PBPK) model was developed for the conazole fungicide triadimefon and its primary metabolite, triadimenol. Rat tissue:blood partition coefficients and metabolic constants were measured in vitro for both compounds. Pharmacokinetic data for parent and metabolite were collected from several tissues after intravenous administration of triadimefon to male Sprague-Dawley rats. The model adequately simulated peak blood and tissue concentrations but predicted more rapid clearance of both triadimefon and triadimenol from blood and tissues. Reverse metabolism of triadimenol to triadimefon in the liver was explored as a possible explanation of this slow clearance, with significant improvement in model prediction. The amended model was extrapolated to humans using in vitro metabolic constants measured in human hepatic microsomes. Human equivalent doses (HEDs) were calculated for a rat no observable adverse effect level (NOAEL) dose of 3.4mg/kg/day using area under the concentration curve (AUC) in brain and blood for triadimefon and triadimenol as dosimetrics. All dosimetric-based HEDs were 25-30 fold above the human oral reference dose of 0.03mg triadimefon/kg/day, but did not account for intra-human variability or pharmacodynamic differences. Ultimately, derivations of this model will be able to better predict the exposure profile of these and other conazole fungicides in humans.

Perchlorate can perturb thyroid hormone (TH) homeostasis by competitive inhibition of iodide uptake by the thyroid gland. Until recently, the effects of perchlorate on TH homeostasis were examined by measuring serum concentrations of THs by immunoassay (IA) methods. IA methods are sensitive, but for TH analysis they are compromised by lack of adequate specificity. In this study, we determined the concentrations of six THs: L-thyroxine (T₄), 3,3',5-triiodo-L-thyronine (T₃), 3,3',5'-triiodo-L-thyronine (rT₃), 3,5-diiodo-L-thyronine, 3,3'-diiodo-L-thyronine, and 3-iodo-L-thyronine in the serum of rats administered perchlorate by isotope (¹³C₆-T₄)-dilution liquid chromatography-tandem mass spectrometry. The method recoveries for THs spiked into a serum matrix were between 97.0% and 115%, with a coefficient of variation of 2.1% to 9.4%. Rats were placed on an iodide-deficient or iodide-sufficient diet for 2.5 months, and for the last 2 weeks of that period they were provided drinking water either without or with perchlorate (10 mg/kg body weight/day). No significant differences in serum concentrations of T₃ and T₄ were observed between rats given iodide-deficient and iodide-sufficient diets for 2 or 2.5 months. After 24 h of perchlorate exposure, significantly lower concentrations of T₃ and T₄ were found in the serum of rats administered the iodide-deficient diet but not in rats administered the iodide-sufficient diet. However, after 2 weeks of perchlorate exposure, TH levels in rats fed the iodide-sufficient diet were also significantly lower than those in control rats. Our results suggest that perchlorate affects TH homeostasis and that such effects are more pronounced under iodide-deficient nutrition.

Jet Fuel 8 (JP-8) is a major fuel source used by US and NATO military. JP-8 is a complex mixture of aliphatic and aromatic isomers of hydrocarbons. Tissue/blood partition coefficient (PC) values are chemical-specific parameters used in modeling the kinetic behavior of chemicals. The partition coefficient values for n-alkanes tend to increase with the increasing carbon number, but less is known about the trend for isomers of n-alkanes. PC values were obtained for the n-alkane nonane (C9) and five of its isomers, namely 3-methyloctane, 4-ethylheptane, 2,3-dimethylheptane, 2,2,4-trimethylhexane, 2,2,4,4-tetramethylpentane. The blood:air and tissue:air PC values correlated with the published log octanol/water (O:W) PC values for n-nonane and its isomers. Experimentally determined blood:air and tissue:air PC values for n-nonane with the largest O:W value were greatest and smallest for the isomer 2,2,4,4-tetramethylpentane with the lowest O:W value. As expected the fat tissue had the highest PC values and muscle the lowest for n-nonane and its isomers. For each tissue, a linear relationship was observed between the tissue/blood PC values for the isomers of n-nonane and n-nonane. This suggests that tissue/blood PC values for all isomers of an alkane could be estimated using data collected from only a sub-set of alkanes of equal carbon number. These reported tissue/blood PC values will support the development of a jet fuel physiologically-based pharmacokinetic (PBPK) model.

Bisphenol A (BPA) is a high-production volume industrial chemical used in the manufacture of polycarbonate plastic products and epoxy resin-based food can liners. The presence of BPA in urine of >90% of Americans aged 6-60 is controversial because of the potential for endocrine disruption, particularly during perinatal development, as suggested by in vitro, experimental animal, and epidemiological studies. The current study used LC/MS/MS to measure serum pharmacokinetics of aglycone (active) and conjugated (inactive) BPA in adult and neonatal rhesus monkeys by oral (PND 5, 35, 70) and intravenous injection (PND 77) routes using d6-BPA to avoid sample contamination. The concentration-time profiles observed in adult monkeys following oral administration of 100 μg/kg bw were remarkably similar to those previously reported in human volunteers given a similar dose; moreover, minimal pharmacokinetic differences were observed between neonatal and adult monkeys for the receptor-active aglycone form of BPA. Circulating concentrations of BPA aglycone were quite low following oral administration (< 1% of total), which reflects the redundancy of active UDP-glucuronosyl transferase isoforms in both gut and liver. No age-related changes were seen in internal exposure metrics for aglycone BPA in monkeys, a result clearly different from developing rats where significant inverse age-related changes, based on immaturity of Phase II metabolism and renal excretion, were recently reported. These observations imply that any toxicological effect observed in rats from early postnatal exposures to BPA could over-predict those possible in primates of the same age, based on significantly higher internal exposures and overall immaturity at birth.

Bisphenol A (BPA) is an important industrial chemical used in the manufacture of polycarbonate plastic products and epoxy resin-based food can liners. The presence of BPA in urine of >90% of Americans aged 6-60 suggests ubiquitous and frequent exposure. The current study used LC/MS/MS to measure serum pharmacokinetics of aglycone (active) and conjugated (inactive) BPA in adult and neonatal Sprague-Dawley rats by oral and injection routes. Deuterated BPA was used to avoid issues of background contamination. Linear pharmacokinetics were observed in adult rats treated orally in the range of 0-200 microg/kg bw. Evidence for enterohepatic recirculation of conjugated, but not aglycone, BPA was observed in adult rats. Significant inverse relationships were observed between postnatal age and measures of internal exposures to aglycone BPA and its elimination. In neonatal rats treated orally, internal exposures to aglycone BPA were substantially lower than from subcutaneous injection. The results reinforce the critical role for first-pass Phase II metabolism of BPA in gut and liver after oral exposure that attenuates internal exposure to the aglycone form in rats of all ages. The internal exposures to aglycone BPA observed in adult and neonatal rats following a single oral dose of 100 microg/kg bw are inconsistent with effects mediated by classical estrogen receptors based on binding affinities. However, an impact on alternative estrogen signaling pathways that have higher receptor affinity cannot be excluded in neonatal rats. These findings emphasize the importance of matching aglycone BPA internal dosimetry with receptor affinities in experimental animal studies reporting toxicity.

Perchlorate can competitively inhibit iodide uptake by the thyroid gland (TG) via the sodium/iodide symporter, consequently reducing the production of thyroid hormones (THs). Until recently, the effects of perchlorate on TH homeostasis are being examined through measurement of serum levels of TH, by immunoassay (IA)-based methods. IA methods are fast, but for TH analysis, they are compromised by the lack of adequate specificity. Therefore, selective and sensitive methods for the analysis of THs in TG are needed, for assessment of the effects of perchlorate on TH homeostasis. In this study, we developed a method for the analysis of six THs: L-thyroxine (T(4)), 3,3',5-triiodo-L-thyronine (T(3)), 3,3',5'-triiodo-L-thyronine (rT(3)), 3,5-diiodo-L-thyronine (3,5-T(2)), 3,3'-diiodo-L-thyronine (3,3'-T(2)), and 3-iodo-L-thyronine (3-T(1)) in TG, using liquid chromatography (LC)-tandem mass spectrometry (MS/MS). TGs used in this study were from rats that had been placed on either iodide-deficient diet or iodide-sufficient diet, and that had either been provided with perchlorate in drinking water (10 mg/kg/day) or control water. TGs were extracted by pronase digestion and then analyzed by LC-MS/MS. The instrumental calibration range for each TH ranged from 1 to 200 ng/ml and showed a high linearity (r>0.99). The method quantification limits (LOQs) were determined to be 0.25 ng/mg TG for 3-T(1); 0.33 ng/mg TG for 3,3'- and 3,5-T(2); and 0.52 ng/mg TG for rT(3), T(3), and T(4). Rats were placed on an iodide-deficient or -sufficient diet for 2.5 months, and for the last 2 weeks of that period were provided either perchlorate (10 mg/kg/day) in drinking water or control water. Iodide deficiency and perchlorate administration both reduced TG stores of rT(3), T(3), and T(4). In iodide-deficient rats, perchlorate exacerbated the reduction in levels of THs in TG. With the advances in analytical methodology, the use of LC-MS/MS for measurement of hormone levels in TG will allow more comprehensive evaluations of the hypothalamic-pituitary-thyroid axis.

The major objective of this project was to characterize the systemic disposition of the pyrethroid, deltamethrin (DLT), in immature rats, with emphasis on the age dependence of target organ (brain) dosimetry. Postnatal day (PND) 10, 21, and 40 male Sprague-Dawley rats received 0.4, 2, or 10 mg DLT/kg by gavage in glycerol formal. Serial plasma, brain, fat, liver, and skeletal muscle samples were collected for up to 510 h and analyzed for DLT and/or 3-phenoxybenzoic acid (PBA) content by high-performance liquid chromatography. Toxicokinetic data from previous experiments of the same design with young adult (PND 90) rats (Kim, K.-B., Anand, S. S., Kim, H. J., White, C. A., and Bruckner, J. V. [2008]. Toxicokinetics and tissue distribution of deltamethrin in adult Sprague-Dawley rats. Toxicol. Sci. 101, 197-205) were used to compare to immature rat data. Plasma and tissue DLT levels were inversely related to age. Preweanlings and weanlings showed markedly elevated brain concentrations and pronounced salivation, tremors, choreoathetosis, and eventual fatalities. Plasma DLT levels did not reliably reflect brain levels over time. Plasma:brain ratios were time and dose dependent, but apparently not age dependent. Brain levels were better correlated with the magnitude of salivation and tremors than plasma levels. Hepatic intrinsic clearance of DLT progressively increased during maturation, as did the hepatic extraction ratio. Thus, limited capacity to metabolically inactivate DLT appeared primarily responsible for the inordinately high target organ doses and acute neurotoxicity in pups and weanling rats. Hepatic blood flow was not rate limiting in any age group. Limited DLT hydrolysis was manifest in vivo in the pups by relatively low plasma PBA levels. Elevated exposure of the immature brain to a pyrethroid may prove to be of consequence for long-term, as well as short-term neurotoxicity.

Understanding the potential differences in metabolic capacity and kinetics between various common laboratory species as well as between genders is an important facet of chemical risk assessment that is often overlooked, particularly for chemicals which undergo non-cytochrome P450 mediated metabolism. The use of physiologically based pharmacokinetic (PBPK) models to better describe chemical exposure is made more powerful by incorporation of high quality in vitro kinetic data. To this end, metabolism of the conazole fungicide triadimefon was studied in hepatic microsomes of both genders of SD rats and CD-1 mice. Triadimefon depletion and triadimenol formation were measured in each type of microsomes. Michaelis-Menten regressions were applied to metabolic data and V(MAX) and the Michaelis constant (K(M)) values calculated. Male SD rats metabolized triadimefon more rapidly than female SD rats or either gender of CD-1 mouse. K(M) values were in the micromolar range, indicating the possibility of competitive inhibition with endogenous substrates. Intrinsic clearances derived from kinetic parameters indicate that triadimefon metabolism is blood-flow limited in all organisms studied with the possible exception of female rat. The in vitro half-life method was investigated as a less resource intensive method for the derivation of intrinsic clearance, and was found to be useful as a complement to the traditional Michaelis-Menten approach.

Background Perchlorate (ClO4−) is an environmental contaminant known to disrupt the thyroid axis of many terrestrial and aquatic species. ClO4− competitively inhibits iodide uptake into the thyroid at the sodium/iodide symporter and disrupts hypothalamic–pituitary–thyroid (HPT) axis homeostasis in rodents. Objective We evaluated the proposed mode of action for ClO4−-induced rat HPT axis perturbations using a biologically based dose–response (BBDR) model of the HPT axis coupled with a physiologically based pharmacokinetic model of ClO4−. Methods We configured a BBDR-HPT/ClO4− model to describe competitive inhibition of thyroidal uptake of dietary iodide by ClO4− and used it to simulate published adult rat drinking water studies. We compared model-predicted serum thyroid-stimulating hormone (TSH) and total thyroxine (TT4) concentrations with experimental observations reported in these ClO4− drinking water studies. Results The BBDR-HPT/ClO4− model failed to predict the ClO4−-induced onset of disturbances in the HPT axis. Using ClO4− inhibition of dietary iodide uptake into the thyroid, the model underpredicted both the rapid decrease in serum TT4 concentrations and the rise in serum TSH concentrations. Conclusions Assuming only competitive inhibition of thyroidal uptake of dietary iodide, BBDR-HPT/ClO4− model calculations were inconsistent with the rapid decrease in serum TT4 and the corresponding increase in serum TSH. Availability of bound iodide in the thyroid gland governed the rate of hormone secretion from the thyroid. ClO4− is translocated into the thyroid gland, where it may act directly or indirectly on thyroid hormone synthesis/secretion in the rat. The rate of decline in serum TT4 in these studies after 1 day of treatment with ClO4− appeared consistent with a reduction in thyroid hormone production/secretion. This research demonstrates the utility of a biologically based model to evaluate a proposed mode of action for ClO4− in a complex biological process.

The developing central nervous system of the fetus and neonate is recognized as very sensitive to maternal or gestational hypothyroidism. Despite this recognition, there is still a lack of data concerning the relationship between thyroid-related biomarkers and neurological outcomes. We used propylthiouracil administered at 0, 3, or 10 ppm in drinking water from gestational d 2 until weaning to create hypothyroid conditions to study the relationship between hypothalamic-pituitary-thyroid axis compensation and impaired neurodevelopment. In addition to serum T(3), T(4), free T(4), and TSH concentrations, cerebrocortical T(3) concentration (cT(3)), hepatic type I and cerebrocortical type II (D2) 5'-deiodinase activity, and thyroidal mRNA for thyroglobulin and sodium iodide symporter were measured. Extracellular recordings from the CA1 region in hippocampal slices were obtained from both postnatal d 21-32 (pups) and postnatal d 90-110 (adults) rats to assess neurophysiological effects. Thyroidal mRNA for thyroglobulin and sodium iodide symporter were increased in pups but not in dams. Both propylthiouracil doses increased cerebrocortical D2 activity approximately 5-fold in pups but only 10 ppm increased D2 activity in dams. In dams, cT(3) concentrations were maintained at 3 ppm but fell 75% at 10 ppm. cT(3) concentration in pups fell 50% at 3 ppm and more than 90% at 10 ppm. In both 3 and 10 ppm pups, hippocampal baseline synaptic activity correlated negatively with cerebrocortical D2 activity. In 3 ppm adults, impaired long-term potentiation was evident. In summary, during depletion of serum T(4), D2 activity served as a sensitive marker of tissue thyroid status, an indicator of the brain's compensatory response to maintain cT(3), and correlated with a neurophysiological outcome.

We report on the transient and persistent effects of JP-8 jet fuel exposure on auditory function in rats. JP-8 has become the standard jet fuel utilized in the United States and North Atlantic Treaty Organization countries for military use and it is closely related to Jet A fuel, which is used in U.S. domestic aviation. Rats received JP-8 fuel (1000 mg/m(3)) by nose-only inhalation for 4 h and half of them were immediately subjected to an octave band of noise ranging between 97 and 105 dB in different experiments. The noise by itself produces a small, but permanent auditory impairment. The current permissible exposure level for JP-8 is 350 mg/m(3). Additionally, a positive control group received only noise exposure, and a fourth group consisted of untreated control subjects. Exposures occurred either on 1 day or repeatedly on 5 successive days. Impairments in auditory function were assessed using distortion product otoacoustic emissions and compound action potential testing. In other rats, tissues were harvested following JP-8 exposure for assessment of hydrocarbon levels or glutathione (GSH) levels. A single JP-8 exposure by itself at 1000 mg/m(3) did not disrupt auditory function. However, exposure to JP-8 and noise produced an additive disruption in outer hair cell function. Repeated 5-day JP-8 exposure at 1000 mg/m(3) for 4 h produced impairment of outer hair cell function that was most evident at the first postexposure assessment time. Partial though not complete recovery was observed over a 4-week postexposure period. The adverse effects of repeated JP-8 exposures on auditory function were inconsistent, but combined treatment with JP-8 + noise yielded greater impairment of auditory function, and hair cell loss than did noise by itself. Qualitative comparison of outer hair cell loss suggests an increase in outer hair cell death among rats treated with JP-8 + noise for 5 days as compared to noise alone. In most instances, hydrocarbon constituents of the fuel were largely eliminated in all tissues by 1-h postexposure with the exception of fat. Finally, JP-8 exposure did result in a significant depletion of total GSH that was observable in liver with a nonsignificant trend toward depletion in the brain and lung raising the possibility that the promotion of noise-induced hearing loss by JP-8 might have resulted from oxidative stress.

Jet Propellant 8 (JP-8) is a kerosene-based jet fuel used in the military and is composed of hundreds of hydrocarbons. A PBPK model was developed to assess the metabolic interactions of JP-8 vapor on two prominent constituents of JP-8 vapor, m-xylene (XYL) and ethylbenzene (EBZ). A limited number of rats were exposed to JP-8 vapor in a Leach chamber for 4 h to 380, 1100, or 2700 mg/m3 (total hydrocarbon). Several individual hydrocarbons were monitored in the chamber atmosphere, including XYL, EBZ, and the total hydrocarbon concentration. Blood and liver were harvested and analyzed by a novel headspace SPME/GC-MS method that allowed for identification of individual hydrocarbons and low limits of detection. The PBPK model was able to describe the metabolic interactions between XYL, EBZ, and a lumped aromatic fraction of JP-8 vapor estimated to be 18 to 25% of the fuel vapor. Competitive inhibition of XYL and EBZ metabolism was observed for JP-8 vapor inhalation exposures of 1100 and 2700 mg/m3. Future inhalation studies with jet fuel include aerosol exposures and expansion of the PBPK models to include other hydrocarbons such as n-alkanes and upper respiratory tract dosimetry of aerosol droplets.

The National Toxicology Program (NTP) and the National Institute of Environmental Health Sciences (NIEHS) established the NTP Center for the Evaluation of Risks to Human Reproduction (CERHR) in June 1998. The purpose of CERHR is to provide timely, unbiased, scientifically sound evaluations of human and experimental evidence for adverse effects on reproduction and development caused by agents to which humans may be exposed. Hydroxyurea was selected for evaluation by a CERHR expert panel because of (1) its increasing use in the treatment of sickle cell disease in children and adults, (2) knowledge that it inhibits DNA synthesis and is cytotoxic, and (3) published evidence of its reproductive and developmental toxicity in rodents. Hydroxyurea is FDA-approved for reducing the frequency of painful crises and the need for blood transfusions in adults with sickle cell anemia who experience recurrent moderate-to-severe crises. Hydroxyurea is used in the treatment of cancer, sickle cell disease, and thalassemia. It is the only treatment for sickle cell disease aside from blood transfusion used in children. Hydroxyurea may be used in the treatment of children and adults with sickle cell disease for an extended period of time or for repeated cycles of therapy. Treatment with hydroxyurea may bemore » associated with cytotoxic and myelosuppressive effects, and hydroxyurea is mutagenic.« less

Deltamethrin (DLT) is a type II pyrethroid insecticide widely used in agriculture and public health. DLT is a potent neurotoxin that is primarily cleared from the body by metabolism. To better understand the dosimetry of DLT in the central nervous system, a physiologically based pharmacokinetic (PBPK) model for DLT was constructed for the adult, male Sprague-Dawley rat that employed both flow-limited (brain, gastrointestinal [GI] tract, liver, and rapidly perfused tissues) and diffusion-limited (fat, blood/plasma, and slowly perfused tissues) rate equations. The blood was divided into plasma and erythrocytes. Cytochrome P450-mediated metabolism was accounted for in the liver and carboxylesterase (CaE)-mediated metabolism in plasma and liver. Serial blood, brain, and fat samples were taken for DLT analysis for up to 48 h after adult rats received 2 or 10 mg DLT/kg po. Hepatic biotransformation accounted for approximately 78% of these administered doses. Plasma CaEs accounted for biotransformation of approximately 8% of each dosage. Refined PBPK model forecasts compared favorably to the 2- and 10-mg/kg po blood, plasma, brain, and fat DLT profiles, as well as profiles subsequently obtained from adult rats given 1 mg/kg iv. DLT kinetic profiles extracted from published reports of oral and iv experiments were also used for verification of the model's simulations. There was generally good agreement in most instances between predicted and the limited amount of empirical data. It became clear from our modeling efforts that there is considerably more to be learned about processes that govern GI absorption and exsorption, transport, binding, brain uptake and egress, fat deposition, and systemic elimination of DLT and other pyrethroids. The current model can serve as a foundation for construction of models for other pyrethroids and can be improved as more definitive information on DLT kinetic processes becomes available.

Maternal exposure to high doses of trichloroethylene (TCE) and its oxidative metabolites, trichloroacetic acid (TCA) and dichloroacetic acid (DCA), has been implicated in eye malformations in fetal rats, primarily micro-/anophthalmia. Subsequent to a cardiac teratology study of these compounds ( Fisher et al. 2001 , Int. J. Toxicol. 20:257–267), their potential to induce ocular malformations was examined in a subset of the same experimental animals. Pregnant, Sprague-Dawley Crl:CDR BR rats were orally treated on gestation days (GDs) 6 to 15 with bolus doses of either TCE (500 mg/kg/day), TCA (300 mg/kg/day), DCA (300 mg/kg/day), or all- trans retinoic acid (RA; 15 mg/kg/day). The heads of GD 21 fetuses were not only examined grossly for external malformations, but were sectioned using a modified Wilson’s technique and subjected to computerized morphometry that allowed for the quantification of lens area, globe area, medial canthus distance, and interocular distance. Gross ocular malformations were essentially absent in all treatment groups except for the RA group in which 26% of fetuses exhibited micro-/anophthalmia. Using the litter as the experimental unit of analysis, lens area, globe area, and interocular distance were statistically significantly reduced in the DCA treatment group. Statistically significant reductions in lens and globe areas also occurred in the RA treatment group, all four ocular measures were reduced in the TCA treatment group but none significantly so, and TCE was without effect. Because DCA, TCA, and RA treatments were associated with significant reductions in fetal body weight (bw), data were also statistically analyzed after bw adjustment. Doing so dramatically altered the results of treatment group comparisons, but the severity of bw reduction and the degree of change in ocular measures did not always correlate. This suggests that bw reduction may not be an adequate explanation for all the changes observed in ocular measures. Thus, it is unclear whether DCA specifically disrupted ocular development even under these provocative exposure conditions. Clearly, however, if TCE is capable of disrupting ocular development in the Sprague-Dawley rat, a higher dose than that employed in the present study is required.

The primary aim of the Acute Exposure Guideline Level (AEGL) program is to develop scientifically credible limits for once-in-a-lifetime or rare acute inhalation exposures to high-priority, hazardous chemicals. The program was developed because of the need of communities for information on hazardous chemicals to assist in emergency planning, notification, and response, as well as the training of emergency response personnel. AEGLs are applicable to the general population, including children, the elderly, and other potentially susceptible subpopulations. AEGLs are the airborne concentrations of chemicals above which a person could experience notable discomfort or irritation (AEGL-1); serious, long-lasting health effects (AEGL-2); and life-threatening effects or death (AEGL-3). AEGLs are determined for five exposure periods (10 and 30 min and 1, 4, and 8 h). Physiologically based pharmacokinetic (PBPK) models can be very useful in the interspecies and time scaling often required here. PBPK models are used for the current article to predict AEGLs for trichlorethylene (TCE), based on the time course of TCE in the blood and/or brain of rats and humans. These AEGLs are compared to values obtained by standard time-scaling methods. Comprehensive toxicity assessment documents for each chemical under consideration are prepared by the National Advisory Committee for AEGLs, a panel comprised of representatives of federal, state, and local governmental agencies, as well as industry and private-sector organizations. The documents are developed according to National Research Council (NRC) guidelines and must be reviewed by the NRC Subcommittee on Acute Exposure Guideline Levels before becoming final. AEGLs for 18 chemicals have been published, and it is anticipated that 40 to 50 chemicals will be evaluated annually.

Decane, a 10-carbon n-alkane and one of the highest vapor phase constituents of jet propellent-8 (JP-8), was selected to represent the semivolatile fraction for the initial development of a physiologically based pharmacokinetic (PBPK) model for JP-8. Rats were exposed to decane vapors at time-weighted average concentrations of 1200, 781, or 273 ppm in a 32-L Leach chamber for 4 h. Time-course samples for 1200 ppm and end-of-exposure samples for 781 and 273 ppm decane exposures were collected from blood, brain, liver, fat, bone marrow, lung, skin, and spleen. The pharmacokinetics of decane could not be described by flow-limited assumptions and measured in vitro tissue/air partition coefficients. A refined PBPK model for decane was then developed using flow-limited (liver and lung) and diffusion-limited (brain, bone marrow, fat, skin, and spleen) equations to describe the uptake and clearance of decane in the blood and tissues. Partition coefficient values for blood/air and tissue/blood were estimated by fitting end-of-exposure pharmacokinetic data and assumed to reflect the available decane for rapid exchange with blood. A portion of decane is speculated to be sequestered in "deep" pools in the body, unavailable for rapid exchange with blood. PBPK model predictions were adequate in describing the tissues and blood kinetics. For model validation, the refined PBPK model for decane had mixed successes at predicting tissue and blood concentrations for lower concentrations of decane vapor, suggesting that further improvements in the model may be necessary to extrapolate to lower concentrations.

Perchlorate (ClO4-), a contaminant in drinking water, competitively inhibits active uptake of iodide (I-) into various tissues, including mammary tissue. During postnatal development, inhibition of I- uptake in the mammary gland and neonatal thyroid and the active concentration ClO4- in milk indicate a potentially increased susceptibility of neonates to endocrine disruption. A physiologically based pharmacokinetic (PBPK) model was developed to reproduce measured ClO4- distribution in the lactating and neonatal rat and predict resulting effects on I- kinetics from competitive inhibition at the sodium iodide symporter (NIS). Kinetic I- and ClO4- behavior in tissues with NIS (thyroid, stomach, mammary gland, and skin) was simulated with multiple subcompartments, Michaelis-Menten (M-M) kinetics and competitive inhibition. Physiological and kinetic parameters were obtained from literature and experiment. Systemic clearance and M-M parameters were estimated by fitting simulations to tissue and serum data. The model successfully describes maternal and neonatal thyroid, stomach, skin, and plasma, as well as maternal mammary gland and milk data after ClO4- exposure (from 0.01 to 10 mg/kg-day ClO4-) and acute radioiodide (2.1 to 33,000 ng/kg I-) dosing. The model also predicts I- uptake inhibition in the maternal thyroid, mammary gland, and milk. Model simulations predict a significant transfer of ClO4- through milk after maternal exposure; approximately 50% to 6% of the daily maternal dose at doses ranging from 0.01 to 10.0 mg ClO4-/kg-day, respectively. Comparison of predicted dosimetrics across life-stages in the rat indicates that neonatal thyroid I- uptake inhibition is similar to the adult and approximately tenfold less than the fetus.

Due to perchlorate's (ClO4-) ability to competitively inhibit thyroid iodide (I-) uptake through the sodium-iodide symporter (NIS), potential human health risks exist from chronic exposure via drinking water. Such risks may include hypothyroidism, goiter, and mental retardation (if exposure occurs during critical periods in neurodevelopment). To aid in predicting perchlorate's effect on normal I- kinetics, we developed a physiologically-based pharmacokinetic (PBPK) model for the adult male rat. The model structure describes simultaneous kinetics for both anions together with their interaction at the NIS, in particular, the inhibition of I- uptake by ClO4-. Subcompartments and Michaelis-Menten (M-M) kinetics were used to describe active uptake of both anions in the thyroid, stomach, and skin. Separate compartments for kidney, liver, plasma, and fat were described by passive diffusion. The model successfully predicts both 36ClO4- and 125I- kinetics after iv doses of 3.3 mg/kg and 33 mg/kg, respectively, as well as inhibition of thyroid 125I- uptake by ClO4- after iv doses of ClO4- (0.01 to 3.0 mg/kg). The model also predicts serum and thyroid ClO4- concentrations from 14-day drinking water exposures (0.01 to 30.0 mg ClO4-/kg/day) and compensation of perchlorate-induced inhibition of radioiodide uptake due to upregulation of the thyroid. The model can be used to extrapolate dose metrics and correlate observed effects in perchlorate toxicity studies to other species and life stages, such as rat gestation (Clewell et al., 2003). Because the model successfully predicts perchlorate's interaction with iodide, it provides a sound basis for future incorporation of the complex hypothalamic-pituitary-thyroid feedback system.

Ammonium perchlorate is used as an oxidizer in rocket fuel. It has become a groundwater contaminant, dissociating to ammonium cation and perchlorate anion. The perchlorate ion competes with iodide for uptake into the thyroid, reducing thyroid hormone production. Pregnant Sprague-Dawley rats were given either untreated or perchlorate (1 mg/kg-day) treated drinking water beginning on gestation day 2. One set of control and exposed dams was sacrificed on gestation day 20. The litters from the second set of control and exposed dams were crossed immediately after parturition and were sacrificed at postnatal day 10. Dam serum and thyroid, pooled fetal sera, and male and female pup sera were collected and analyzed for perchlorate, thyroid-stimulating hormone (TSH), triiodothyronine (T3), and thyroxine (T4). Control pups receiving perchlorate through lactation had serum levels at postnatal day 10 of 0.54 μg/ml and 0.56 μg/ml for male and female pups, respectively, whereas exposed fetuses had serum perchlorate levels of 0.38 ± 0.04 μg/ml. Female pups receiving perchlorate lactationally had significantly lower levels of serum T4 than control pups and prenatally exposed pups. Serum T4 levels in male pups were not affected by perchlorate. Serum thyroid hormone levels from gestational perchlorate exposure were restored to control values by postnatal day 10. In utero perchlorate-exposure decreased serum T4 levels in the fetus. Gestational studies in conjunction with a crossfostering study design helped discern thyroid hormonal changes caused by perchlorate exposure during the perinatal period.

Perchlorate, an environmental contaminant, is known to disturb the hypothalamus–pituitary–thyroid (HPT) axis by blocking iodide accumulation in the thyroid. Iodide deficiency can lead to hypothyroidism and goiter in rats. The objective of the study was to characterize the pharmacokinetics of perchlorate in male Sprague–Dawley rats relative to inhibition of thyroidal radiolabeled iodide uptake and onset of up-regulation of the HPT axis. Radiolabeled perchlorate (3.3 mg/kg 36 ClO − 4 ) was excreted in urine (99.5% over a 48-h period). 36 ClO − 4 is rapidly distributed into tissues with preferential sequestration into skin, gastrointestinal tract (GT), and thyroid. Calculated half-lives of 36 ClO − 4 from the skin, thyroid, plasma, GT, and GT contents were 32.0, 7.6, 7.3, 10.0, and 8.6 h, respectively. Perchlorate was very effective at inhibiting thyroidal uptake of radiolabeled iodide ( 125 I − ). In animals iv dosed with perchlorate followed by an iv challenge of 125 I − , thyroidal 125 I − uptake was diminished by 11, 29, 55, and 82% at 11 h postdosing in the 0.01, 0.1, 1.0, and 3.0 mg/kg perchlorate dose groups, respectively. In perchlorate drinking water studies, dose-dependent inhibition in thyroidal uptake of 125 I − initially occurred with corresponding increases in serum thyroid-stimulating hormone (TSH) levels and decreases in thyroid hormone levels. TSH stimulated recovery from the initial perchlorate blocking effects was evident during 14 days of treatment in the 1.0 and 3.0 mg/kg per day treatment groups. However, recovery of serum thyroid hormones at these doses was much slower despite evidence for iodide sufficiency in the thyroid. These results suggest that the typical homeostatic mechanisms of the thyroid may respond differently at high doses of perchlorate used in this rat study (above 1 mg/kg per day) or perchlorate may be acting on the HPT axis by mechanisms other than thyroidal 125 I − uptake inhibition.

Trichloroethylene (TCE), trichloroacetic acid (TCA), and dichloroacetic acid (DCA) are commonly found as groundwater contaminants in many regions of the United States. Cardiac birth defects in children have been associated with TCE, and laboratory studies with rodents report an increased incidence of fetal cardiac malformations resulting from maternal exposures to TCE, TCA, and DCA. The objective of this study was to orally treat pregnant CDR(CD) Sprague-Dawley rats with large bolus doses of either TCE (500 mg/kg), TCA (300 mg/kg), or DCA (300 mg/kg) once per day on days 6 through 15 of gestation to determine the effectiveness of these materials to induce cardiac defects in the fetus. All- trans retinoic acid (RA) dissolved in soybean oil was used as a positive control. Soybean oil is commonly used as a dosing vehicle for RA teratology studies and was also used in this study as a dosing vehicle for TCE. Water was used as the dosing vehicle for TCA and DCA. Fetal hearts were examined on gestation day (GD) 21 by an initial in situ, cardiovascular stereomicroscope examination, and then followed by a microscopic dissection and examination of the formalin-fixed heart. The doses selected for TCA and DCA resulted in a modest decrease in maternal weight gain during gestation (3% to 8%). The fetal weights on GD 21 in the TCA and DCA treatment groups were decreased 8% and 9%, respectively, compared to the water control group and 21% in the RA treatment group compared to soybean oil control group. The heart malformation incidence for fetuses from the TCE-, TCA-, and DCA-treated dams did not differ from control values on a per fetus or per litter basis. The rate of heart malformations, on a per fetus basis, ranged from 3% to 5% for TCE, TCA, and DCA treatment groups compared to 6.5% and 2.9% for soybean oil and water control groups. The RA treatment group was significantly higher with 33% of the fetuses displaying heart defects. For TCE, TCA, and DCA treatment groups 42% to 60% of the litters contained at least one fetus with a heart malformation, compared to 52% and 37% of the Utters in the soybean oil and water control groups. For the RA treatment group, 11 of 12 litters contained at least one fetus with a heart malformation. Further research is needed to quantify the spontaneous rates of heart defects for vehicle control rats and to explain the disparity between findings in the present study and other reported findings on the fetal cardiac teratogenicity of TCE, TCA, and DCA.

NASA and DoD use ammonium perchlorate (NH4ClO4) salt as an oxidizer in solid rocket propellants. Perchlorate salts are used in several applications such as flares, pyrotechnics, explosives, batteri...

Dichloroacetic acid (DCA) is a contaminant found in treated drinking water due to chlorination. DCA has been shown to be a complete hepatocarcinogen in both mice and rats. In this study we developed a rapid and sensitive high-performance liquid chromatography (HPLC) method to simultaneously detect DCA and its metabolites, oxalic acid, glyoxylic acid and glycolic acid in blood and urine samples of animals sub-chronically administered with DCA (2 g/l) in drinking water. Both urine and plasma samples were treated minimally before HPLC analysis. Separation and detection of DCA and its metabolites were achieved using an anion-exchange column and a conductivity detector. The mobile phase consisted of an initial concentration of 0.01 mM sodium hydroxide in 40% methanol followed by a linear gradient from 0.01 mM to 60 mM sodium hydroxide in 40% methanol for 30 min. The lower detection limit for DCA and each of its three major metabolites was 0.05 microg/ml. DCA and its metabolites gave a linear response range from 0.05 to 100 microg/ml. Plasma DCA was also analyzed by gas chromatography (GC), and the results obtained correlated with those from the HPLC method (correlation coefficient=0.999). While available HPLC techniques offer sensitive procedures to detect either glycolic acid or oxalic acid, the described HPLC method has the unique advantage of determining simultaneously the parent compound (DCA) and its three major metabolites (oxalic acid, glyoxylic acid and glycolic acid) in biological samples, without complex sample preparation.

Bisphenol A (BPA) was administered by gavage (2.5-300,000 g/kg body weight (bw)/day) to pregnant Sprague Dawley dams, newborn pups, and continuing into adulthood. Aglycone (i.e., un- conjugated and active) and conjugated (i.e., inactive) BPA were evaluated by liquid chromatography electrospray tandem mass spectrometry (LC-ES/MS/MS) in serum to better interpret tox- icological endpoints measured in the study. Ethinyl estradiol (EE2, 0.5 and 5 g/kg bw/day) and the endogenous hormones, 17- estradiol (E2) and testosterone, were similarly evaluated. Mean BPA aglycone levels in vehicle and nacontrol rat serum (0.02- 0.5 ng/ml) indicated sample processing artifact, consistent with literature reports of a propensity for postexposure blood contam- ination by BPA. Direct measurements of BPA-glucuronide in ve- hicle and nacontrol serum (2-10nM) indicated unintentional exposure and metabolism at levels similar to those produced by 2.5 g/kg bw/day BPA (7-10nM), despite careful attention to poten- tial BPA inputs (diet, drinking water, vehicle, cages, bedding, and dust) and rigorous dosing solution certification and delivery. The source of this exposure could not be identified, but interpretation of the toxicological effects, observed only at the highest BPA doses, was not compromised. Internal exposures to BPA and EE2 agly- cones were highest in young rats. When maximal serum concen- trations from the two highest BPA doses and both EE2 doses were compared with concurrent levels of endogenous E2, the ER bind- ing equivalents were similar to or above those of endogenous E2 in male and female rats of all ages tested. Such evaluations of estro- genic internal dosimetry and comprehensive evaluation of contam- ination impact should aid in extrapolating risks from human BPA exposures.

The widespread usage of methylphenidate (MPH) in the pediatric population has received considerable attention due to its potential effect on child development. For the first time a physiologically based pharmacokinetic (PBPK) model has been developed in juvenile and adult humans and nonhuman primates to quantitatively evaluate species- and age-dependent enantiomer specific pharmacokinetics of MPH and its primary metabolite ritalinic acid. The PBPK model was first calibrated in adult humans using in vitro enzyme kinetic data of MPH enantiomers, together with plasma and urine pharmacokinetic data with MPH in adult humans. Metabolism of MPH in the small intestine was assumed to account for the low oral bioavailability of MPH. Due to lack of information, model development for children and juvenile and adult nonhuman primates primarily relied on intra- and interspecies extrapolation using allometric scaling. The juvenile monkeys appear to metabolize MPH more rapidly than adult monkeys and humans, both adults and children. Model prediction performance is comparable between juvenile monkeys and children, with average root mean squared error values of 4.1 and 2.1, providing scientific basis for interspecies extrapolation of toxicity findings. Model estimated human equivalent doses in children that achieve similar internal dose metrics to those associated with pubertal delays in juvenile monkeys were found to be close to the therapeutic doses of MPH used in pediatric patients. This computational analysis suggests that continued pharmacovigilance assessment is prudent for the safe use of MPH.

A six-compartment physiologically based pharmacokinetic (PBPK) model for the B6C3F1 mouse was developed for trichloroethylene (TCE) and was linked with five metabolite submodels consisting of four compartments each. The PBPK model for TCE and the metabolite submodels described oral uptake and metabolism of TCE to chloral hydrate (CH). CH was further metabolized to trichloroethanol (TCOH) and trichloroacetic acid (TCA). TCA was excreted in urine and, to a lesser degree, metabolized to dichloroacetic acid (DCA). DCA was further metabolized. The majority of TCOH was glucuronidated (TCOG) and excreted in the urine and feces. TCOH was also excreted in urine or converted back to CH. Partition coefficient (PC) values for TCE were determined by vial equilibrium, and PC values for nonvolatile metabolites were determined by centrifugation. The largest PC values for TCE were the fat/blood (36.4) and the blood/air (15.9) values. Tissue/blood PC values for the water-soluble metabolites were low, with all PC values under 2.0. Mice were given bolus oral doses of 300, 600, 1200, and 2000 mg/kg TCE dissolved in corn oil. At various time points, mice were sacrificed, and blood, liver, lung, fat, and urine were collected and assayed for TCE and metabolites. The 1200 mg/kg dose group was used to calibrate the PBPK model for TCE and its metabolites. Urinary excretion rate constant values were 0. 06/hr/kg for CH, 1.14/hr/kg for TCOH, 32.8/hr/kg for TCOG, and 1. 55/hr/kg for TCA. A fecal excretion rate constant value for TCOG was 4.61/hr/kg. For oral bolus dosing of TCE with 300, 600, and 2000 mg/kg, model predictions of TCE and several metabolites were in general agreement with observations. This PBPK model for TCE and metabolites is the most comprehensive PBPK model constructed for P450-mediated metabolism of TCE in the B6C3F1 mouse.

Trichloroethylene (TCE), an environmental contaminant of National concern, is the focus of a new health risk assessment process incorporating the Proposed Cancer Risk Assessment Guidelines. This paper describes not only how TCE became an environmental problem for the Air Force, but also details the new Risk Assessment process envisioned by the Environmental Protection Agency's (EPA) National Center for Environmental Assessment (NCEA). Insights on epidemiological evaluations, both past and future, and their impact on the cancer classification of TCE are discussed. Examples of how physiologically based pharmacokinetics and dose-response characterization described in the new Cancer Guidelines are applied to TCE are provided. In addition, a variety of modeling techniques are discussed for the development of reference doses (oral exposure) and reference concentrations (inhalation exposures) for TCE. Finally, the role of risk communication is included. This new process provides an example of how interagency (EPA, Department of Defense. Department of Energy) and extramural (industry, academia) partnerships can provide greater gains to the nation, as a whole, than any of the parts on their own.

Atrazine (ATR) is a widely used chlorotriazine herbicide, a ubiquitous environmental contaminant, and a potential developmental toxicant. To quantitatively evaluate placental/lactational transfer and fetal/neonatal tissue dosimetry of ATR and its major metabolites, physiologically based pharmacokinetic models were developed for rat dams, fetuses and neonates. These models were calibrated using pharmacokinetic data from rat dams repeatedly exposed (oral gavage; 5mg/kg) to ATR followed by model evaluation against other available rat data. Model simulations corresponded well to the majority of available experimental data and suggest that: (1) the fetus is exposed to both ATR and its major metabolite didealkylatrazine (DACT) at levels similar to maternal plasma levels, (2) the neonate is exposed mostly to DACT at levels two-thirds lower than maternal plasma or fetal levels, while lactational exposure to ATR is minimal, and (3) gestational carryover of DACT greatly affects its neonatal dosimetry up until mid-lactation. To test the model's cross-species extrapolation capability, a pharmacokinetic study was conducted with pregnant C57BL/6 mice exposed (oral gavage; 5mg/kg) to ATR from gestational day 12 to 18. By using mouse-specific parameters, the model predictions fitted well with the measured data, including placental ATR/DACT levels. However, fetal concentrations of DACT were overestimated by the model (10-fold). This overestimation suggests that only around 10% of the DACT that reaches the fetus is tissue-bound. These rodent models could be used in fetal/neonatal tissue dosimetry predictions to help design/interpret early life toxicity/pharmacokinetic studies with ATR and as a foundation for scaling to humans.

The water disinfection byproduct bromate (BrO{sub 3}{sup −}) produces cytotoxic and carcinogenic effects in rat kidneys. Our previous studies demonstrated that BrO{sub 3}{sup −} caused sex-dependent differences in renal gene and protein expression in rats and the elimination of brominated organic carbon in their urine. The present study examined changes in renal cell apoptosis and protein expression in male and female F344 rats treated with BrO{sub 3}{sup −} and associated these changes with accumulation of 3-bromotyrosine (3-BT)-modified proteins. Rats were treated with 0, 11.5, 46 and 308 mg/L BrO{sub 3}{sup −} in drinking water for 28 days and renal sections were prepared and examined for apoptosis (TUNEL-staining), 8-oxo-deoxyguanosine (8-oxoG), 3-BT, osteopontin, Kim-1, clusterin, and p-21 expression. TUNEL-staining in renal proximal tubules increased in a dose-related manner beginning at 11.5 mg BrO{sub 3}{sup −}/L in female rats and 46 mg/L in males. Increased 8-oxoG staining was observed at doses as low as 46 mg/L. Osteopontin expression also increased in a dose-related manner after treatment with 46 mg/L, in males only. In contrast, Kim-1 expression increased in a dose-related manner in both sexes, although to a greater extent in females at the highest dose. Clusterin and p21 expression also increased in amore » dose-related manner in both sexes. The expression of 3-BT-modified proteins only increased in male rats, following a pattern previously reported for accumulation of α-2{sub u}-globulin. Increases in apoptosis in renal proximal tubules of male and female rats at the lowest doses suggest a common mode of action for renal carcinogenesis for the two sexes that is independent of α-2{sub u}-globulin nephropathy. - Highlights: • Bromate induced nephrotoxicity in both male and female rats by similar mechanisms. • Apoptosis was seen in both male and female rats at the lowest doses tested. • Bromate-induced apoptosis correlated to 8-oxo-deoxyguanosine formation. • Bromate increased the level of 3-bromotyrosine-modified proteins in male rats only. • These data identify possible novel mechanisms for bromate-induced nephrotoxicity.« less

The rat hepatoma cell line, H4IIE, serves as a useful tool to assess potential biological effects such as induction of cytochrome P4501A1 expression. The objectives of this study were twofold: to investigate the kinetic time course and dosimetry of PCB77 in rat hepatoma cells dosed with PCB77 and in liver of rats given ip doses of PCB77, and to compare in vitro and in vivo P4501A1 enzyme induction responses. For the 4-day time–course study, H4IIE cells were exposed with two doses of [ 14 C]PCB77 (0.9 and 3 μg/plate) and harvested at 15 and 30 min, 1, 2, 4, 8, and 12 hr, and 1, 2, 3, and 4 days. PCB77-derived radioactivity was detected in the cells as early as 15 min postdosing. For the dose–response study, the cells were dosed with various concentrations of PCB77 (0.00316–5.37 μg/plate) and harvested on Day 3 since ethoxyresorufin O -deethylase (EROD) activity in vitro reached its maximum on the third day postdosing. Time–course and dose–response studies revealed that only 1–3% of the total delivered dose was found in the cells, with the remainder in the media and adhering to the culture plates. For the dose–response study in vivo, male Fischer rats were dosed with a single ip injection of various concentrations of PCB77 (0.1–50 mg/kg body wt) and euthanized on Day 3. PCB77-derived radioactivity and EROD induction in vivo were measured. When EROD activity and PCB77-derived radioactivity in the rat hepatoma cells and in the rat liver were compared on an equivalent weight basis, there was a significant correlation ( r 2 = 0.985) between them. Prior to this study, no information on quantitative dosimetry and EROD activities of PCB77 has been reported to validate the in vitro assay with in vivo data.