A comprehensive analysis of the bioaccumulation of three cyclic (namely, D4, D5, and D6) and five linear (namely, L2, L3, L4, L5, and L6) volatile methylsiloxanes (VMS) in biota was conducted by reviewing 50 published articles and laboratory reports on aquatic (82%) and terrestrial (18%) species. A total of 178 bioaccumulation values were evaluated employing a quantitative weight-of-evidence (qWoE) approach provided by the Bioaccumulation Assessment Tool. Considerable variability of bioconcentration factor (BCF) and bioaccumulation factor (BAF) values was observed for VMS in aquatic organisms, which ranged from below to above their respective regulatory threshold values. The frequency of evidence for nonbioaccumulation (i.e., BCF or BAF <2,000) was ≥50% for D6, L2, L5, and L6. Conversely, laboratory biomagnification factor (BMF) values were generally ≥71% less than 1, indicating low bioaccumulation potential through dietary uptake. Field BMFs displayed more variability but still suggested a low bioaccumulation potential. Despite the majority (≥65%) of trophic magnification factors (TMFs) in aquatic food webs being below 1, the broad range of TMF values for all VMS examined indicates potential uncertainties introduced by environmental factors in the target environments (e.g., chemical exposure gradient) and sampling bias. Overall, lines of evidence (≥71%) suggest that cyclic and linear VMS generally do not bioaccumulate in aquatic species, with dietary uptake being the primary pathway. For terrestrial species, primarily rats, predicted and field BMFs were less than 1 at 37 °C, primarily via respiratory elimination. Field TMFs for terrestrial food webs, including invertebrates and various bird species with body temperatures of 18 and 40 °C, respectively, were sourced from a single study, where D4 demonstrated trophic dilution, while D5 and D6 generally did not biomagnify in this terrestrial food web.

Octamethylcyclotetrasiloxane (D4) is a high production volume chemical that has been subject to thorough toxicological investigations. Animal studies with the substance were conducted with either Fischer 344 or Sprague Dawley CD rats. While the pharmacokinetic fate of D4 in Fischer rats is well understood, little information exists on Sprague Dawley CD rats, where reproductive effects have been demonstrated. The objective of this study was to explore the pharmacokinetic behavior in both rats, and to identify potential strain-specific differences. Fischer and Sprague Dawley CD rats were exposed for six hours to 700 ppm of 14C-D4 vapor either with or without preceding 14-day exposure to non-radiolabeled D4. Time-course data in blood, tissues and excreta were obtained through 168 h post-exposure and analyzed for both total radioactivity and parent D4. The data confirm that repeated exposure results in increased metabolism in both rat strains, confirming the findings of earlier studies of auto-induction of CYP2B1/2 by D4. The results also indicate that D4 is subject to strain-specific pharmacokinetic behavior, and that Fischer rats appear to metabolize D4 to a greater extent than Sprague Dawley CD rats.

To investigate the potential toxicity of Octamethylcyclotetrasiloxane (D4), studies in laboratory rats have used primarily one of two strains, Sprague-Dawley (SD) and Fischer-344 (F-344). Reproductive studies used SD rats whereas F-344 rats were used in D4 pharmacokinetics, metabolism, acute/subacute/chronic toxicity and oncogenicity studies. Here, we assessed specific endpoints related to D4 pharmacokinetics and biochemistry in SD and F-344 rats within a single study, which allows for direct comparisons between strain and sex. This assessment included determination of microsomal total P450, NADPH-cytochrome c reductase, epoxide hydrolase, CYP2B1/2, CYP1A1/2, CYP3A1/2, CYP2C11, and CYP2A1. Aside from slight brown pigment in the liver, the treated animals experienced no toxicologically significant weight loss, decrease in food consumption, or clinical signs. Concentrations of D4 in plasma and fat were generally greater in females relative to males in both strains. SD females appeared to have statistically significantly greater plasma and fat concentrations following 28 days of repeated exposure to D4 relative to F-344 rats, suggesting the existence of potential sex and strain differences in D4 pharmacokinetics. The effect of D4 exposure on liver enzyme expression was similar among and between sexes and strain and was consistent with that for phenobarbital-like inducers. Notable differences included a finding of elevated CYP2B1/2 protein levels without a similar magnitude of increase in CYP2B/1 activity and a greater degree of CYP3A1/2 induction (protein and activity) for female SD rats. The importance of these findings is unclear, however reduced CYP2B1/2 activity may give rise to lower rates of D4 metabolism and clearance, consistent with the higher tissue levels of D4 in SD relative to F-344 female rats.

Octamethylcyclotetrasiloxane (D4) and decamethylcyclopentasiloxane (D5) are low molecular weight cyclic volatile methyl siloxanes (cVMSs) primarily used as intermediates or monomers in the production of high molecular weight silicone polymers. The use of D4 as a direct ingredient in personal care products has declined significantly over the past 20 years, although it may be present as a residual impurity in a variety of consumer products. D5 is still used as an intentional ingredient in cosmetics, consumer products and in dry cleaning. Persons who may be exposed include occupational exposure for workers, and potential inhalation or dermal exposure for consumers and the general public. Because of the diverse use, especially of D5, and the potential for human exposure, a comprehensive program was undertaken to understand the kinetics, metabolism, enzyme induction and toxicity of D4 and D5 in rats following relevant routes of exposure. Physiologically based pharmacokinetic (PBPK) models utilizing these studies have been reported for D4 and D5 in the rat and human following dermal and inhalation exposures, with the oral uptake component of the model being limited in its description. Data from high dose oral studies in corn oil and simethicone vehicles and neat were used in the D4/D5 harmonized PBPK model development. It was uncertain if the inability to adequately describe the oral uptake was due to unrealistic high doses or unique aspects of the chemistry of D4/D5. Low dose studies were used to provide data to refine the description of oral uptake in the model by exploring the dose dependency and the impact of a more realistic food-like vehicle. Absorption, distribution, metabolism and elimination (ADME) of D4 and D5 was determined following a single low oral gavage dose of 14C-D4 and 14C-D5 at 30 and 100mg/kg body weight (bw), respectively, in a rodent liquid diet. Comparison of the low vs. high dose oral gavage administration of D4 and D5 demonstrated dose-dependent kinetic behavior. Data and modeling results suggest differences in metabolism between low and high dose administration indicating high dose administration results in or approaches non-linear saturated metabolism. These low dose data sets were used to refine the D4/D5 multi-route harmonized PBPK model to allow for a better description of the disposition and toxicokinetics of D4/D5 following oral exposure. With a refined oral uptake description, the model could be used in risk assessment to better define the internal dose of D4 and D5 following exposure to D4 and D5 via multiple routes.

Octamethylcyclotetrasiloxane (D4) and decamethylcyclopentasiloxane (D5) are used as intermediates or monomers in the synthesis of silicon-based polymers for industrial or consumer applications. D4 and D5 may remain as residual monomer in these polymers at less than 1000 ppm and may therefore be present as a minor impurity in consumer products. For D5, in addition to the manufacture of polymers, its uses include intentional addition to consumer products, personal care products and some dry-cleaning solvents. Two-year rodent chronic bioassays were conducted with both substances and borderline increases in the incidence of uterine tumors were observed, specifically, benign uterine adenoma with D4 and adenocarcinoma with D5. The effects profile and induction of uterine tumors share some similarity with that seen with chronic exposure to dopamine agonists. The current study investigated the potential for D4 and D5 to elicit dopamine agonist-like effects on estrous cyclicity. Separate groups of reproductively senescent female Fischer 344 rats (F344) were exposed via vapor inhalation to D4 (700 ppm, 9.3 mg/L) or D5 (160 ppm, 2.1 mg/L) or to a diet containing 0.0045, 0.045, or 4.5 ppm pergolide mesylate (PM), a potent dopamine agonist used here as a reference substance, from 11 through 24 months of age. The primary focus was to characterize the effects of D4 and D5 exposure on estrous cyclicity relative to that observed with PM. As a monitoring effort, circulating endogenous estradiol, progesterone, prolactin and corticosterone levels were evaluated monthly. A blood sample from each rat was obtained via tail vein in the afternoon after the daily inhalation exposure period once every 4 weeks. Histomorphologic examination of the major organs including the reproductive tract was conducted on all animals at study termination. This study has shown that chronic exposure to D4 and D5 can affect cyclicity in the reproductively senescent F344 rat. For each substance the effect on cyclicity involved reduction in the incidence of pseudopregnancy with a shift toward cycles more typical of younger animals. D4 and D5 induced an increase in estrous cycle repetition whereas D4 also increased the incidence of extended estrus. These shifts resulted in animals entering proestrus/estrus significantly more times over the duration of the study than seen in the control group. Similar effects were observed with the reference substance, PM. However, distinct differences in the timing and magnitude of the effects on the estrous cycle and impact on prolactin, progesterone, estradiol, and corticosterone suggest that D4 and D5 are not classical dopamine agonists even though a similar increased incidence of proestrus/estrus was also observed with PM. These results may prove important with respect to understanding D4- and D5-induced uterine tumor response in the F344 rat, given the relationship between increased incidence of uterine endometrium stimulation by endogenous estrogen as a consequence of extended or more frequent proestrus/estrus, uterine tumor risk, and questions of relevance to humans. Recent publications have summarized the existing data on D4 and D5, with emphasis on exploring the biological relevance of the uterine tumors (Klaunig et al., 2016a,b; Franzen et al., 2017; Dekant and Klaunig, 2016; Dekant et al., 2017). The authors concluded that although the mode of action has not yet been fully established, the data, including the findings from this study, indicate that the D4- and D5-induced uterine tumors observed in the rodent chronic bioassays have no relevance for human risk characterization based not only on the distinct species differences in regulation of the reproductive systems, but also the high exposure levels and duration required for expression in rats.

Octamethylcyclotetrasiloxane (D4) is a cyclic volatile methylsiloxane primarily used in the synthesis of silicon-based materials used in a variety of consumer products. This paper details the chronic toxicity and oncogenicity evaluation of D4 in the Fischer 344 rat. Animals were exposed to 0, 10, 30, 150, or 700 ppm D4 vapor for 6 h/day, 5 days/week for up to 104 weeks in whole-body inhalation chambers. Effects of two year chronic exposure included increased liver, kidney, testes, and uterine weight with correlating microscopic findings of hepatocellular hypertrophy (males only), chronic nephropathy (both sexes), interstitial cell hyperplasia, and cystic endometrial hyperplasia and endometrial adenoma, respectively. Upper respiratory tract irritation and lymphocytic leukocytosis were evident in both sexes. Increased neoplasia was demonstrated only in the uterus. Uterine endometrial adenomas were present in four of sixty animals exposed to 700 ppm D4 for 24 months. None were present in the other treatment groups. In contrast, in 700 ppm D4 group males the incidence of pituitary and pancreatic neoplasia was reduced as was thyroid c-cell adenoma/carcinoma in 700 ppm females. This study has identified that D4 is a mild respiratory irritant and increases liver and kidney weight without inducing neoplasia in these tissues. The increased incidence of uterine adenoma was the only treatment-related neoplastic finding associated with chronic exposure to D4.

Decamethylcyclopentasiloxane (D5) is a cyclic polydimethylsiloxane used in the synthesis of silicon-based materials and as a component in consumer products. Male and female Fischer 344 rats were exposed to D5 vapor (0, 10, 40, 160 ppm; whole-body inhalation) for 6 h/d, 5 d/wk, for up to 104 weeks. Microscopic examination of tissues revealed test article effects at 160 ppm in the upper respiratory tract (hyaline inclusions in males and females at 6, 12, and 24 months) and an increased incidence of uterine endometrial adenocarcinoma at 24-months. The hyaline inclusions were considered a non-adverse tissue response for lack of any other respiratory tract non-neoplastic or neoplastic changes. Uterine endometrial adenocarcinoma was not anticipated. Toxicity testing (mutagenicity/genotoxicity, acute, sub-acute and sub-chronic descriptive toxicity) performed prior to the conduct of the chronic bioassay provided no indication that the uterus was a potential target organ. The target organ and tumor type specificity (adenocarcinoma is a common spontaneous tumor in the aged Fischer 344 rat) suggests the effect is associated with estrous cycle alteration. A robust assessment of potential mode(s) of action responsible for the uterine tumors and relevance to humans is addressed in a companion manuscript (Klaunig et al., 2015).

Scientists from academia, industry, and government reviewed current international regulations for the screening of commercial chemicals for bioaccumulation in the context of the current state of bioaccumulation science. On the basis of this review, several recommendations were proposed, including a scientific definition for “bioaccumulative substances,” improved criteria for the characterization of bioaccumulative substances (including the trophic magnification factor and the biomagnification factor), novel methods for measuring and calculating bioaccumulation properties, and a framework for screening commercial chemicals for bioaccumulative substances. The proposed framework for bioaccumulation screening improves current practices by reducing miscategorization, making more effective use of available bioaccumulation data that currently cannot be considered, reducing the need for animal testing, providing simpler and cheaper test protocols for animal studies in case animal studies are necessary, making use of alternative testing strategies, including in vitro and in silico metabolic transformation assays, and providing a scientific foundation for bioaccumulation screening that can act to harmonize bioaccumulation screening among various jurisdictions.

With non-volatile compounds, high lipophilicity (i.e., fat:blood partition coefficients, Pf, in the range of several hundred to a thousand or higher) typically leads to concerns for bioaccumulation. To evaluate the extent to which highly cleared, lipophilic vapors are expected to accumulate in blood and tissues, we conducted pharmacokinetic (PK) analysis, using both a generic physiologically based (PBPK) model for inhalation of volatile compounds (VCs) and a more detailed PBPK model specifically developed for a highly lipophilic volatile (decamethylcyclopentasiloxane, D5). The generic PBPK model for inhalation of VCs in humans showed that highly metabolized, lipophilic compounds, with a low blood:air partition coefficient (Pb), do not accumulate in blood or systemic tissues with repeat exposures although a period of days to weeks may be required for fat to reach periodic steady state. VCs with higher Pb (in the hundreds) and lower hepatic extraction accumulate in blood on repeat exposures. The more detailed PBPK model for D5 also showed that this lipophilc VC does not accumulate in blood and predictions of the increases in D5 in fat with repeat exposures in rats agreed with experiments. In general, the major characteristic favoring accumulation of VCs in blood and systemic tissues is poor whole-body clearance, not lipophilicty. The term bioaccumulation should be used to refer to cases where repeat exposures lead to increases in VC blood (or central compartment) concentration. Based on this definition, highly cleared VCs, such as D5, would not be considered to bioaccumulate on repeat exposures.

Decamethylcyclopentasiloxane (D(5)), a volatile cyclic methyl siloxane (VCMS), is used in industrial and consumer products. Inhalation pharmacokinetics of another VCMS, octamethylcyclotetrasiloxane (D(4)), have been extensively investigated and successfully modeled with a multispecies physiologically based pharmacokinetic (PBPK) model. Here, we develop an inhalation PBPK description for D(5), using the D(4) model structure as a starting point, with the objective of understanding factors that regulate free blood and tissue concentrations of this highly lipophilic vapor after inhalation in rats and humans. Compared with D(4), the more lipophilic D(5) required deep compartments in lung, liver, and plasma to account for slow release from tissues after cessation of exposures. Simulations of the kinetics of a stable D(5) metabolite, HO-D(5), required diffusion-limited uptake in fat, a deep tissue store in lung, and its elimination by fecal excretion and metabolism to linear silanols. The combined D(5)/HO-D(5) model described blood and tissue concentrations of parent D(5) and elimination of total radioactivity in single and repeat exposures in male and female rats at 7 and 160 ppm. In humans, D(5) kinetic data are more sparse and the model structure though much simplified, still required free and bound blood D(5) to simulate exhaled air and blood time courses from 1 h inhalation exposures at 10 ppm in five human volunteers. This multispecies PBPK model for D(5) highlights complications in interpreting kinetic studies where chemical in blood and tissues represents various pools with only a portion free. The ability to simulate free concentrations is essential for dosimetry based risk assessments for these VCMS.

The disposition of decamethylcyclopentasiloxane (D5) in male and female Fischer 344 rats following single or repeated inhalation exposures was evaluated. Animals were administered a single 6-h nose-only exposure to 7 or 160 ppm 14C-D5 or fourteen 6-h nose-only exposures to unlabeled D5 followed on day 15 by a 6-h exposure to 14C-D5. Subgroups of exposed animals were used to evaluate body burden, distribution, elimination, and deposition on the fur. Retention of radioactivity following single and repeated exposures was relatively low (approximately 1-2% of inhaled D5). Radioactivity and parent D5 were widely distributed to tissues of both male and female rats, with the maximum concentration of radioactivity observed in most tissues by 3 h postexposure. Fat was a depot for D5, with elimination occurring much slower than observed for plasma and other tissues. In all groups, the primary route for elimination of radioactivity was through expired air. Analyses for parent D5 indicated that essentially all the radioactivity in the expired volatiles was unchanged D5. Repeated exposure gave rise to higher levels of parent D5 in the lung and fat of both sexes and in female liver relative to the single exposure. In fat, immediately after sacrifice approximately 50% of the radioactivity was attributed to parent. Five polar metabolites of D5 were identified in urine, with no parent D5 detected. Radiochromatograms demonstrated two peaks in feces. One corresponded to the retention time for D5. The second has been putatively identified as hydroxylated D5.

The pharmacokinetics of octamethylcyclotetrasiloxane (D4), a highly lipophilic and well-metabolized volatile cyclic siloxane, are more complex than those of other volatile hydrocarbons. The purpose of the present study was to evaluate rate constants for saturable metabolism in the body, to estimate possible presystemic D4 clearance by respiratory-tract tissues, and to assess rate constants for uptake of D4 after oral dosing. These experiments provided the opportunity to refine current physiologically based pharmacokinetic (PBPK) models for D4 and to independently estimate key model parameters by sensitive inhalation methods. The PBPK model could only be fitted to gas uptake results when metabolic capacity was included in the respiratory-tract epithelium. The model simulations were highly sensitive to the parameter for total percent of whole-body metabolism allocated to the respiratory tract, with optimal fits observed with this value equal to 5%. Oral uptake of D4 was evaluated using both closed and open chamber concentration time-course studies after intubation of D4 in corn oil. Conclusions from the oral uptake studies were also verified by comparison with independent data sets for blood concentrations of D4 after oral dosing. The pharmacokinetic (PK) analysis of uptake from the gut and release from blood into chamber air results for oral doses from 10 to 300 mg D4/kg body weight were consistent with a combination of prolonged and slow uptake of D4 from the gastrointestinal tract and of reduced absorption at higher doses, as well as the extrahepatic clearance of D4 in pulmonary tissues. These closed chamber gas uptake studies provide a valuable confirmation of the conclusions reached in other pharmacokinetic studies and have uncovered a situation where closed chamber loss is highly sensitive to respiratory-tract clearance. This sensitivity largely arises from the unusual characteristics of D4: high-affinity metabolic clearance and low blood:air partitioning.

Octamethylcyclotetrasiloxane (D4) has been shown to have effects on the female rat reproductive cycle. This study evaluated the phase of the female rat reproductive cycle affected by D4 using a study design that allowed the complete female reproductive cycle, as well as phases of the cycle, from pre-mating through gestation, to be evaluated. Rats were exposed via whole body vapor inhalation up to 700ppm D4 during the overall phase (28 days prior to mating through gestation day (GD) 19), the ovarian phase (31–3 days prior to mating), the fertilization phase (3 days prior to the start of mating through gestation day 3), and the implantation phase (GD 2–GD 5) of the reproductive cycle. D4 was associated with decreases in implantation sites and litter size in the overall and fertilization phases, but not in the ovarian or implantation phases. In order to further define the sensitive period for D4 exposure, additional groups of rats were exposed on single days. A single 6h exposure to D4 on the day prior to mating resulted in a significant reduction in fertility. These data indicate that there is a very narrow window, around the time of ovulation and fertilization, for D4 to exert effects on the reproductive cycle of the female rat. Subsequent research, reported elsewhere, has elucidated the mode of action and assessed its potential relevance to humans.

In this study, data for human dermal absorption of octamethylcyclotetrasiloxane, D-4, and decamethylcyclopentasiloxane, D-5, through axilla skin in vivo are interpreted using pharmacokinetic models of dermal absorption by adding the dermal exposure route to inhalation physiologically based pharmacokinetics models developed previously. The compartmental model describing dermal absorption of these compounds included volatilization of the applied chemical from the skin surface, diffusion of absorbed chemical back to the skin surface and evaporation of this chemical from the skin surface after the applied dose had cleared from the application site, uptake from the skin compartment into blood, and a storage compartment within the skin. Data from exposures in volunteers (i.e., D-4 and D-5 concentrations in exhaled air and plasma) were used to estimate model parameters. In volunteers exposed to either D-4 or D-5, the maximum concentration of chemical in exhaled air reached a maximum at or prior to 1 h following administration of the test chemical. Based on model calculations, the percent of applied dose of D-4 that was absorbed into systemic circulation for men and women was 0.12 and 0.30%, respectively; for D-5 about 0.05% of the applied dose was absorbed for both men and women. For both D-4 and D-5, model calculations indicate that more than 83% of the chemical that reached systemic circulation was eliminated by exhalation within 24 h. These whole-body pharmacokinetic models for dermal absorption of two semi-volatile compounds provide a valuable tool for understanding factors controlling their dermal absorption through axilla skin and for applying results from these studies in consumer product risk assessments.

ABSTRACT A scientific workshop was held in 2006 to discuss the use of in vitro Absorption, Distribution, Metabolism, and Excretion (ADME) data in chemical bioaccumulation assessments for fish. Computer-based (in silico) modeling tools are widely used to estimate chemical bioaccumulation. These in silico methods have inherent limitations that result in inaccurate estimates for many compounds. Based on a review of the science, workshop participants concluded that two factors, absorption and metabolism, represent the greatest sources of uncertainty in current bioaccumulation models. Both factors can be investigated experimentally using in vitro test systems. A variety of abiotic and biotic systems have been used to predict chemical accumulation by invertebrates, and dietary absorption of drugs and xenobiotics by mammals. Research is needed to determine whether these or similar methods can be used to better predict chemical absorption across the gills and gut of fish. Scientists studying mammals have developed a stepwise approach to extrapolate in vitro hepatic metabolism data to the whole animal. A series of demonstration projects was proposed to investigate the utility of these in vitro–in vivo extrapolation procedures in bioaccumulation assessments for fish and delineate the applicability domain of different in vitro test systems. Anticipating research progress on these topics, participants developed a “decision tree” to show how in vitro information for individual compounds could be used in a tiered approach to improve bioaccumulation assessments for fish and inform the possible need for whole-animal testing.

This study evaluated the potential toxicity of whole-body vapor inhalation of octamethylcyclotetrasiloxane (D4) on reproductive capabilities in exposed F0 and F1 parental animals and the potential effects on neonatal survival, growth, and development of the F1 and F2 offspring. F0 male and female Sprague–Dawley rats (30/sex/group) were exposed to D4 vapor at concentrations of 0, 70, 300, 500 or 700 ppm 6 h per day for at least 70 consecutive days prior to mating and lasted through weaning of the pups on postnatal day (PND) 21. Female exposures were suspended from gestation day (GD) 21 through PND 4 to allow for parturition and permit continuous maternal care for the early neonates. Starting on PND 22, F1 weanlings were exposed to D4 as described for the F0 generation. The F2 pups were not directly exposed to D4. F0 animals were mated once to produce the F1 generation; F1 parental animals were mated twice to produce two F2 litters. In addition, the F1 males were mated with unexposed females. Prolonged estrous cycles, decreased mating and fertility indices were observed in the F1 generation exposed to D4 for the first and second matings. Significant reductions in the mean number of pups born and mean live litter size were observed in the 500 and 700 ppm groups for both the F0 and F1 generations. Implantation sites were also reduced at 700 ppm for both F0 and F1 generations. No adverse effects were observed at any exposure level on anogenital distance, vaginal patency and preputial separation. No adverse effects were seen on male functional reproductive parameters, spermatogenic endpoints, microscopic evaluation of male reproductive tissue, or when the D4-exposed F1 males were mated with the unexposed females, demonstrating that the reproductive toxicity observed was due to D4 exposure to the females. Based on the lack of effect on reproduction when the D4-exposed males were mated to näive females, the NOAEL for male reproductive toxicity was considered to be 700 ppm. Based on the statistically significant effects on fertility and litter size, NOAEL for female reproductive toxicity was considered to be 300 ppm. The findings observed in this study are consistent with suppression or delaying of LH surge as well as acceleration of the onset of female reproductive senescence in the rat. While analogous pathways control ovulation in both rats and humans, there are significant differences in the mechanism for timing and release of LH and resulting changes in the control of ovulation and mating behavior between the two species. If D4 delays rather than causes a prolonged suppression or ablation of the LH surge, the reproductive mode of action of D4 would not likely be relevant for humans.

The objectives of this study were to assess the potential for D4 to suppress the pre-ovulatory lutenizing hormone (LH) surge, to block or delay ovulation, and to evaluate potential effects on reproductive hormones in rats. Female Sprague–Dawley Crl:CD® (SD) IGS BR rats received whole-body vapor inhalation exposure to D4 (0, 700, or 900 ppm) 6 h per day for 3 days. Trunk blood obtained on proestrus at 10 a.m. was evaluated for levels of follicle stimulating hormone (FSH), estradiol (E2), estrone (E1), and progesterone (P4). Other rats had serial blood samples collected via cannula at 2, 4, 6, 8, and 10 p.m. on the day of proestrus and plasma evaluated for LH and prolactin (PRL). Trunk blood was collected at 8 a.m. of estrus and plasma evaluated for FSH, E2, E1, and P4. At 10 a.m. on proestrus, significant increases in E1 levels in the 700 and 900 ppm groups and significant increases in P4 levels in the 900 ppm group were noted. At 8 a.m. on estrus, significant increases in E1, E2, in the E1/E2 ratio and decreases in FSH were noted in the 700 and 900 ppm groups. The major effect on the LH profile was observed most clearly when the rats were grouped by ovulatory status, animals that did or did not ovulate. Regardless of treatment, suppression of the LH surge correlated with blocked ovulation. The percentage of rats that ovulated was (700 ppm, 42%; 900 ppm, 31%) compared to controls (79%). Overall, the data indicate that high exposures to D4 attenuated the pre-ovulatory LH surge and significantly decreased the portion of female rats that ovulated.

A scientific workshop was held in 2006 to discuss the use of in vitro Absorption, Distribution, Metabolism, and Excretion (ADME) data in chemical bioaccumulation assessments for fish. Computer-based (in silico) modeling tools are widely used to estimate chemical bioaccumulation. These in silico methods have inherent limitations that result in inaccurate estimates for many compounds. Based on a review of the science, workshop participants concluded that two factors, absorption and metabolism, represent the greatest sources of uncertainty in current bioaccumulation models. Both factors can be investigated experimentally using in vitro test systems. A variety