# Presentation: 179423

## R E S E A R C H     T R I A N G L E     P A R K ,    N O R T H    C A R O L I N A

**R E S E A R C H     T R I A N G L E     P A R K ,    N O R T H    C A R O L I N A**

## R E S E A R C H     T R I A N G L E     P A R K ,    N O R T H    C A R O L I N A

**R E S E A R C H     T R I A N G L E     P A R K ,    N O R T H    C A R O L I N A**

-   Pre-natal Exposures of Children to Polybrominated Diphenyl Ethers: The Collection of Animal and Human Data Along With the Development and Validation of a PBPK Model James Raymer, Amy Licata, Ed GarnerEPA STAR Grant: R830756July 2004

## Basis for Study

- 	**In-utero exposures and exposures during childhood create potential for adverse health effects.**
**	****	********Abnormal or disturbed development of neurological,  endocrine ****	****	****systems**
**	****	**** ******** ****Some adverse effects later in life could be linked to early ****	****	****exposures**
**	****	**** ******** ****Adult and childhood cancers, reproductive and ****	****	****	****development anomalies, behavioral deficits**
**	****In-utero studies to most environmental chemicals lacking.**
**	**
**	****PBDEs widely occurring; Tetra- and penta- isomers most commonly found in biological media.**

## Study Objectives

******   ****Develop a physiologically based pharmacokinetic (PBPK) animal model for 2,2’,4,4’-tetrabromo- and 2,2’,4,4’,5-pentabromodiphenyl ether that can be used to estimate fetal exposure.  **
******   ****The parameters needed to develop the model will be measured.**

## Study Objectives (cont.)

******   ****Analytical methods for PBDEs in human blood and meconium will be developed and applied to samples to estimate the utility of the model for estimating fetal exposures.**
******  ****Determine if cord blood or meconium are appropriate media for measurement of cumulative exposures of newborn babies to PBDEs.**

## Specific Hypotheses

**A rodent PBPK model for PBDEs can be scaled to be applicable to humans.  **
**The PBDE concentrations in cord blood and meconium from newborns are proportional.  **
**Mother’s blood concentrations of PBDEs are predictive of the cord blood and/or meconium concentrations in newborn babies.  **
**Meconium is a useful medium for assessing cumulative dose to the developing fetus.**

## Potential Benefits

****** ****This work will provide a PBPK model that can be used to estimate exposures of unborn children to the target PBDEs.  **
****** ****The most appropriate matrix for the assessment of *****in-utero***** exposure will be obtained.  **
****** ****The developed PBPK model can be used in future work to study different aspects of exposure, including the impact of chronic and intermittent exposures on time-sensitive, developmental events.  **
****** ****The exposures of a group of mothers and their newborn children to the target analytes will be determined.**

## Approach

******  ****A PBPK model will be developed and validated for the target PBDEs that includes the gestational component.**
******  ****A chronic exposure scenario akin to that anticipated in humans will be used.**
******  ****Necessary partitioning and metabolic parameters will be measured using *****in-vivo***** and *****in-vitro***** experiments.**
******  ****Analytical methods for the PBDEs in all matrices under study will be validated.  (Extraction, fractionation followed by GC/ECD, GC/NCI MS).**
******  ****The model will be scaled to humans and the applicability will be tested using biological samples collected from mothers and newborn infants.**

## PBDE PBPK Pregnancy Model

******  ****Develop PBPK rat pregnancy model for PBDEs that can be used to estimate human fetal exposures to PBDEs **
******  ****At present, no published PBPK pregnancy model for PBDEs**
******  ****RAT model – oral exposure:**
**Animal studies (RTI): PCs; Metabolic rates; Single oral and repeated oral experiments; Single iv experiments; PBDE concentrations will be determined in plasma, urine, feces, and tissues**
******  ****HUMAN model – oral exposure:**
**Human data (RTI via local medical center): mother’s blood (shortly before birth), cord blood, meconium, mother’s and baby’s weight, mother’s age, linkage between mother/baby **

## Schematic for Preliminary PBDE PBPK Pregnancy Model

## PBDE PBPK Pregnancy Model

- RAT pregnancy
- PBPK model
- (oral exposure)

- Model representation
- Model parameterization
- Model simulation
- Model validation

- Model
- Extrapolation

- HUMAN pregnancy
- PBPK model
- (oral exposure)

## Animal  Studies

## Comparison of Rat and Human Hepatic P450 Content

- Shimada et al.** **(1994) *Carcinogenesis*. 15, 2523-2529.
- Geungerich, F.P. “Human Cytochrome P450 Enzymes” in *Cytochrome P450: Structure, Mechanism, and Biochemistry*, Ortiz de Montellano ed., Plenum, NY 1995.

**Differences between rat and human in substrate specificity and isoforms suggest that activities in both species should be studied.**

## Progress to date

- Preliminary PBPK Model Development (Model Representation)
- Synthesis of 2,2’,4,4’- tetrabromodiphenyl ether ( 99.5% pure)
- Synthesis of 2,2’,4,4’,5-pentabromodiphenyl ether ( 65% pure)
- Method development started (solvent extraction/ASE; fractionation)
- In-life animal studies begin July 2004

## Current Tissue Method (Brain)

- Grind tissue with anhydrous sodium sulfate
- Add surrogate (PCB 198)
- Extract 3 times with Hexane (5 mL) – vortex, shake, vortex, centrifuge, decant
- Adjust volume to 15 mL; take 3 mL for lipid determination
- Concentrate remaining 12 mL to 1 mL using KD/micro Snyder
- Fractionate with Florisil
- Reduce Volume (25 mL to 1 mL)
- Add internal standard (PCB 119)
- Analyze by GC/ECD (GC/NCI-MS)

## Spiked Brain Recovery

- Triplicate aliquots of brain (29 mg) were spiked with 500 ng of tetra- and penta- BDE (17 ppm or g/g); Extract, fractionate, and analyze using method.  Analyze all fractions (1-3).
- Recoveries of 80-82% of each target analyte; Majority in fraction 1.
- Responses from blank approximately 2% of targets.
- The background is probably acceptable for dosing studies; Might need to be a bit lower for human samples from environmental exposures (to detect low exposure situations rather than non-detect or no difference from background).

****

## Future Work

- Evaluate method for rat tissues; Modifications are likely, especially in the homogenization step.
- Evaluate potential for necropsy contaminations (dust, etc.)
- Determine partition coefficients
- Perform animals studies; tetra-bromo (begin July); penta-bromo after acquisition of material
- Determine metabolic parameters
- Continue model development and apply as described
- Scale to humans and apply to human samples

## Acknowledgement

*This research is being funded by EPA STAR Grant Number **R830756**.  This work has not been reviewed by EPA.  It does not necessarily represent the views of EPA and no official endorsement should be inferred.*