Age effect on perfluorooctanoate (PFOA) plasma concentration in post-weaning rats following oral gavage with ammonium perfluorooctanoate (APFO).

The relationship between age and plasma concentration of perfluorooctanoate (PFOA) in young rats was investigated. The study was conducted in two phases in which male and female rats between 3 and 8 weeks of age were administered the ammonium salt of PFOA (APFO) by single oral gavage at either 10 or 30mg/kg. In Phase I, APFO was administered at a dose of 10mg/kg body weight to 27-, 34-, 38-, 48-, and 55-day-old male and female rats. Plasma was collected 24h after the dose. In Phase II, APFO doses of either 10 or 30mg/kg body weight were given to groups of 23-, 30-, and 32-day-old male and female rats, and plasma was collected at 2 and 24h after the dose (separate groups), and urine was collected for 24h. PFOA concentrations were measured by LC/MS/MS. In Phase I, plasma concentrations of PFOA were not dependent on age for rats 5 weeks of age and older; however, in 4-week-old rats, male plasma PFOA concentrations were 5-6 times lower than during weeks 5-8, and female plasma PFOA concentrations were 2.5-4 times higher than subsequent weeks. In Phase II, plasma samples collected 2h post-dosing indicated no significant difference in the PFOA uptake by age in females; although, in males, plasma PFOA concentrations were significantly less in 32-day-old rats, approximating one-half of the values observed at 23 and 30 days of age. Plasma samples collected 24h after dosing from 3- to 5-week-old rats indicated a slightly but significantly higher male plasma concentration at 30 and 32 days of age as compared to 23 days of age for the 30mg/kg dose group only. Significantly lower (approximately 10-fold) plasma PFOA concentrations occurred in 32-day-old females as compared with 23- and 30-day-old females at both 2 and 24h after the dose. Although statistically significant changes in urine PFOA concentrations did not occur between age and dose groups within sex, urine PFOA concentrations generally supported plasma elimination. At 23 days of age, the ratio of male to female plasma PFOA concentrations was approximately 2-3:1 compared to approximately 30:1 at 32 days of age. An unexplainable inconsistency in PFOA plasma concentrations for both sexes was noted when comparing Phase I values for 27-day-old rats to Phase II values for 23- and 30-day-old rats. The Phase I values for the 27-day-old rats of both sexes were five to six times lower than Phase II values for the 23- and 30-day-old rats. However, Phase I values for 34-day-old rats were comparable to Phase II values for 32-day-old rats. Despite this anomaly between the 23-, 27-, and 30-day-old rat values, there is strong evidence that age-dependent changes in the elimination of PFOA develop in female rats between 3 and 5 weeks of age, with a consistent marked difference occurring after 30 days of age.

Perfluorooctanoic acid: relationship between repeated inhalation exposures and plasma PFOA concentration in the rat.

A large database exists describing the pharmacokinetic behavior of perfluorooctanoic acid (PFOA) following oral exposure. The objective of this study was to examine the concentration- and time-dependence of the pharmacokinetics of inhaled PFOA in rat plasma to determine equivalent inhalation and oral (from literature values) exposure levels. The study was comprised of two separate experiments: a single 6-h inhalation exposure and repeated inhalation exposures for 3 weeks (6h per day, 5 days per week). In both experiments, male and female rats were exposed nose-only to aerosol atmospheres of either 0, 1, 10, or 25mg/m(3) PFOA. In the single exposure experiment, blood was drawn via the tail vein pre-exposure, four times concurrent to exposure, and six times post-exposure up to 24h. In the repeated exposure experiment, blood was collected immediately before and after exposure 3 days per week. Plasma PFOA concentrations were quantitated by liquid chromatography-mass spectrometry (LC-MS). Following the single exposures, plasma PFOA concentrations were directly proportional to airborne concentrations in both male and female rats. Elimination of PFOA from the plasma was sex-dependent, with female rats eliminating PFOA much more rapidly than male rats. Following repeated PFOA exposure, there was little daily PFOA carryover observed in plasma samples from female rats, while males demonstrated an accumulative pattern over the 3-week period. Peak post-exposure PFOA plasma concentrations in female rats averaged 1, 2, and 4 microg/mL when exposed to 1, 10, and 25mg/m(3) PFOA, respectively, and returned to baseline levels by the time of the next pre-exposure sample collection. Male rats reached steady state plasma concentrations of 8, 21, and 36 microg/mL (ppm) after 3 weeks of exposure to 1, 10, and 25mg/m(3) PFOA, respectively. These results demonstrate that the pharmacokinetic properties of inhaled PFOA in male and female rats are similar to those observed in male and female rats following oral dosing with PFOA. It is thus possible to use this internal dose metric (plasma PFOA) for route-to-route dose extrapolation, with inhalation exposures of 1, 10, and 25mg/m(3) PFOA corresponding to oral doses of approximately 0.3, 1.0, and 2.0mg/kg in rats.

Penetration of ammonium perfluorooctanoate through rat and human skin in vitro.

Rat and human epidermal membranes were mounted onto in vitro diffusion cells with an exposure area of 0.64 cm2, and skin integrity was confirmed using electrical impedance. Following membrane selection, Fluorad FC-118, a 20% aqueous solution of ammonium perfluorooctanoate (AFPO), was applied to the epidermal surface of each skin replicate at approximately 150 microL/cm2 and the donor chamber opening occluded with Parafilm. Serial receptor fluid samples were collected hourly from 1 to 6 h and at 12, 24, 30, and 48 h and analyzed by liquid chromatography-mass spectrometry (LC-MS) for APFO anion (PFO-). For rat skin, the time to steady-state penetration (6500+/-3000 ng APFO x cm(-2) x h(-1)) occurred in less than 12 h, which was sustained until termination (48 h). Based on the concentration of the applied test material, the permeability coefficient (Kp) for APFO in rat skin was calculated to be 3.25+/-1.51 x 10(-5) cm/h. By end of the 48-h exposure period, only a small portion of the total APFO applied (1.44+/-1.13%) had penetrated through rat skin. For human skin, steady-state penetration of APFO (190+/-57 ng APFO x cm(-2) x h(-1)) was reached by 12 h. Based on the concentration of the applied test material, the permeability coefficient for APFO in human skin was calculated to be 9.49+/-2.86 x 10(-7) cm/h. By the end of the 48-h exposure period, only a negligible amount of the total APFO applied (0.048+/-0.01%) had penetrated through human skin. Thus, under infinite dose and occlusive conditions, the steady-state penetration of APFO from a 20% solution was approximately 34-fold faster through rat skin than human skin.

Validation of human physiologically based pharmacokinetic model for vinyl acetate against human nasal dosimetry data.

Vinyl acetate has been shown to induce nasal lesions in rodents in inhalation bioassays. A physiologically based pharmacokinetic (PBPK) model for vinyl acetate has been used in human risk assessment, but previous in vivo validation was conducted only in rats. Controlled human exposures to vinyl acetate were conducted to provide validation data for the application of the model in humans. Five volunteers were exposed to 1, 5, and 10 ppm 13C1,13C2 vinyl acetate via inhalation. A probe inserted into the nasopharyngeal region sampled both 13C1,13C2 vinyl acetate and the major metabolite 13C1,13C2 acetaldehyde during rest and light exercise. Nasopharyngeal air concentrations were analyzed in real time by ion trap mass spectrometry (MS/MS). Experimental concentrations of both vinyl acetate and acetaldehyde were then compared to predicted concentrations calculated from the previously published human model. Model predictions of vinyl acetate nasal extraction compared favorably with measured values of vinyl acetate, as did predictions of nasopharyngeal acetaldehyde when compared to measured acetaldehyde. The results showed that the current PBPK model structure and parameterization are appropriate for vinyl acetate. These analyses were conducted from 1 to 10 ppm vinyl acetate, a range relevant to workplace exposure standards but which would not be expected to saturate vinyl acetate metabolism. Risk assessment based on this model further concluded that 24 h per day exposures up to 1 ppm do not present concern regarding cancer or non-cancer toxicity. Validation of the vinyl acetate human PBPK model provides support for these conclusions.

Pharmacokinetics of perfluorooctanoate in cynomolgus monkeys.

The pharmacokinetics of perfluorooctanoate (PFOA) in cynomolgus monkeys were studied in a six-month oral capsule dosing study of ammonium perfluorooctanoate (APFO) and in a single-dose iv study. In the oral study, samples of serum, urine, and feces were collected every two weeks from monkeys given daily doses of either 0, 3, 10, or 20 mg APFO/kg. Steady-state was reached within four weeks in serum, urine, and feces. Serum PFOA followed first-order elimination kinetics after the last dose, with a half-life of approximately 20 days. Urine was the primary elimination route. Mean serum PFOA concentrations at steady state in the 3, 10, and 20 mg/kg-day dose groups, respectively, were 81, 99, and 156 microg/ml in serum; 53, 166, and 181 microg/ml in urine; and, 7, 28, and 50 microg/g in feces. Mean liver concentrations reached 16, 14, and 50 microg/g in the 3, 10, and 20 mg/kg groups, respectively. In the iv study, three monkeys per sex were given a single dose of 10 mg/kg potassium PFOA. Samples were collected through 123 days. The terminal half-life of PFOA in serum was 13.6, 13.7, and 35.3 days in the three male monkeys and 26.8, 29.3, and 41.7 days in the three females. Volume of distribution at steady state was 181 +/- 12 and 198 +/- 69 ml/kg for males and females, respectively. Based on the result of both the oral and iv studies, the elimination half-life is approximately 14-42 days, and urine is the primary route of excretion.

The Tinsley LCR Databridge Model 6401 and electrical impedance measurements to evaluate skin integrity in vitro.

Electrical impedance is used to confirm skin integrity for in vitro dermal regulatory testing and as a tool to evaluate skin condition to determine the irritation and corrosion potential of various chemicals and personal care products. In this experiment, samples of dermatomed human skin were mounted onto static diffusion cells (0.64 cm2) maintained at 32 degrees C. Following equilibration with 0.9% saline in the donor and receptor chambers, an impedance measurement was taken with a Tinsley LCR Databridge Model 6401 set in the resistance mode (R) and in (a) the serial-equivalent mode (SER) with an alternating current (AC) frequency of 100 hertz (Hz), (b) SER and 1000 Hz, (c) parallel-equivalent mode (PAR) and 100 Hz, and (d) PAR and 1000 Hz. With the databridge set in the SER-equivalent mode and an AC frequency of 1000 Hz, the minimum (7.2 kOmega), maximum (10.0 kOmega), and median (8.6 kOmega) impedance values exhibited a limited response range (2.8 kOmega). However, when the Tinsley 6401 was set in the PAR-equivalent mode at the lower AC frequency of 100 Hz the minimum (16.7 kOmega), maximum (134.6 kOmega), median (83.2 kOmega), and range (117.9 kOmega) of values were the highest obtained. The results confirm that the operator-selected settings on the Tinsley LCR Databridge Model 6401 affect the impedance measurement and the dynamic range of values observed for dermatomed human skin in vitro.

The metabolism of beta-chloroprene: preliminary in-vitro studies using liver microsomes.

Based on analogy with butadiene and isoprene, the metabolism of beta-chloroprene (2-chloro-1,3-butadiene, CD) to reactive intermediates is likely to be a key determinant of tumor development in laboratory rodents exposed to CD by inhalation. The purpose of this study is to identify species differences in toxic metabolite (epoxide) formation and detoxification in rodents and humans. The in-vitro metabolism of CD was studied in liver microsomes of B6C3F1 mice, Fischer/344 and Wistar rats, Syrian hamsters, and humans. Microsomal oxidation of CD in the presence of NADP(+), extraction with diethyl ether, and analysis by GC-mass selective detection (MSD) indicated that (1-chloroethenyl)oxirane (CEO) was an important metabolite of CD in the liver microsomal suspensions of all species studied. Other potential water-soluble oxidative metabolites may have been present. The oxidation of CD was inhibited by 4-methyl pyrazole, an inhibitor of CYP 2E1. CEO was sufficiently volatile at 37 degrees C for vial headspace analysis using GC-MSD single ion monitoring (m/z=39). CEO was synthesized and used to conduct partition measurements along with CD and further explore CEO metabolism in liver microsomes and cytosol. The liquid-to-air partition coefficients for CD and CEO in the microsomal suspensions were 0.7 and 58, respectively. Apparent species differences in the uptake of CEO by microsomal hydrolysis were hamster approximately human>rats>mice. Hydrolysis was inhibited by 1,1,1-trichloropropene oxide, a competitive inhibitor of epoxide hydrolase. A preliminary experiment indicated that the uptake of CEO in liver cytosol by GSH conjugation was hamster>rats approximately mice (human cytosol not yet tested). In general, the results suggest that metabolism may help explain species differences showing a greater sensitivity for CD-induced tumorigenicity in mice, for example, compared with hamsters. Additional experiments are in progress to quantify the kinetic parameters of CD oxidation and CEO metabolism by enzymatic hydrolysis and conjugation by glutathione S-transferase for in cytosol. A future goal is to use the kinetic rates to parameterize a physiologically based toxicokinetic model and relate the burden of toxic metabolite to the cancer dose-response observed in experimental animals.