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Toxicological information

Basic toxicokinetics

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Administrative data

Endpoint:
basic toxicokinetics in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Scientifically accceptable and well documented

Data source

Reference
Reference Type:
publication
Title:
In vitro metabolism of the brominated flame retardants 2- ethylhexyl-2,3,4,5-tetrabromobenzoate (TBB) and bis(2-ethylhexyl) 2,3,4,5-tetrabromophthalate (TBPH) in human and rat tissues
Author:
Roberts SC, Macaulay LJ, Stapleton HM
Year:
2012
Bibliographic source:
Chem Res Toxicol 2012; 16;25(7): 1435-1441

Materials and methods

Objective of study:
metabolism
Principles of method if other than guideline:
The in-vitro metabolism of the brominated flame retardant TBPH (bis(2-ethylhexyl) tetrabromophthalate) was examined in human and rat tissues by conducting in vitro experiments with liver and intestinal subcellular fractions.
GLP compliance:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Bis(2-ethylhexyl) tetrabromophthalate
EC Number:
247-426-5
EC Name:
Bis(2-ethylhexyl) tetrabromophthalate
Cas Number:
26040-51-7
Molecular formula:
C24H34Br4O4
IUPAC Name:
1,2-bis(2-ethylhexyl) 3,4,5,6-tetrabromobenzene-1,2-dicarboxylate
Specific details on test material used for the study:
TBPH (99% purity) were purchased as neat solutions from AccuStandard, Inc. (New Haven, CT, USA).
Radiolabelling:
no

Test animals

Species:
other: not applicable - in-vitro test
Strain:
other: not applicable - in-vitro test
Sex:
not specified

Administration / exposure

Route of administration:
other: not applicable - in-vitro test
Vehicle:
other: not applicable - in-vitro test
Duration and frequency of treatment / exposure:
not applicable - in-vitro test
Doses / concentrations
Remarks:
Doses / Concentrations:
not applicable - in-vitro test
No. of animals per sex per dose / concentration:
not applicable - in-vitro test
Control animals:
other: not applicable - in-vitro test

Results and discussion

Preliminary studies:
In experiments with human liver microsomes (HLM), a significant loss of TBPH was not observed, and no metabolites were detected by GC/MS analysis of the sample extracts. An LC/MS-MS method was developed to monitor mono(2-ethylhexyl) tetrabromophthalate (TBMEHP), a potential hydrolysis metabolite of TBPH (Figure 1). After a 6-h incubation with HLM, TBMEHP was not detected as a metabolite of TBPH, and no significant loss of TBPH was observed. However, TBPH was slowly metabolized to form TBMEHP in the presence of 0.1mg/mL of porcine hepatic carboxylesterase (PCE). This reaction was monitored at multiple time points up to 6 h and maintained linearity at an approximate rate of 1.08 pmol/min/mg esterase.

In a previous study (Niino, T., Ishibashi, T., Ishiwata, H., Takeda, K., and Onodera, S. (2003) Characterization of human salivary esterase in enzymatic hydrolysis of phthalate esters. J. Health Sci. 49, 76−81.) with PCE, DEHP (50 μM) was metabolized to form MEHP at a rate of 127 pmol/min/mg protein. This rate was approximately 100 times faster than the hydrolysis of TBPH observed in this study (1.08 pmol/min/mg protein).
The prominent difference between the metabolic hydrolysis of DEHP and TBPH may be a result of steric hindrance by the fully brominated phenyl ring of TBPH.
Main ADME results
Type:
metabolism
Results:
In experiments with human liver microsomes (HLM), a significant loss of TBPH was not observed, and no metabolites were detected by GC/MS analysis of the sample extracts.

Toxicokinetic / pharmacokinetic studies

Details on absorption:
not applicable - in-vitro test
Details on distribution in tissues:
not applicable - in-vitro test
Details on excretion:
not applicable - in-vitro test

Metabolite characterisation studies

Metabolites identified:
no

Bioaccessibility (or Bioavailability)

Bioaccessibility (or Bioavailability) testing results:
not applicable - in-vitro test

Any other information on results incl. tables

In experiments with human liver microsomes (HLM), a significant loss of TBPH was not observed, and no metabolites were detected by GC/MS analysis of the sample extracts. An LC/MS-MS method was developed to monitor mono(2-ethylhexyl) tetrabromophthalate (TBMEHP), a potential hydrolysis metabolite of TBPH (Figure 1). After a 6-h incubation with HLM, TBMEHP was not detected as a metabolite of TBPH, and no significant loss of TBPH was observed. However, TBPH was slowly metabolized to form TBMEHP in the presence of 0.1mg/mL of porcine hepatic carboxylesterase (PCE). This reaction was monitored at multiple time points up to 6 h and maintained linearity at an approximate rate of 1.08 pmol/min/mg esterase.

In a previous study (Niino, T., Ishibashi, T., Ishiwata, H., Takeda, K., and Onodera, S. (2003) Characterization of human salivary esterase in enzymatic hydrolysis of phthalate esters. J. Health Sci. 49, 76−81.) with PCE, DEHP (50 μM) was metabolized to form MEHP at a rate of 127 pmol/min/mg protein. This rate was approximately 100 times faster than the hydrolysis of TBPH observed in this study (1.08 pmol/min/mg protein).

Applicant's summary and conclusion

Conclusions:
Interpretation of results: bioaccumulation potential cannot be judged based on study results
Executive summary:

The in-vitro metabolism of the brominated flame retardant TBPH was examined.

In experiments with human liver microsomes (HLM), a significant loss of TBPH was not observed, and no metabolites were detected by GC/MS analysis of the sample extracts. An LC/MS-MS method was developed to monitor mono(2-ethylhexyl) tetrabromophthalate (TBMEHP), a potential hydrolysis metabolite of TBPH. After a 6-h incubation with HLM, TBMEHP was not detected as a metabolite of TBPH, and no significant loss of TBPH was observed. However, TBPH was slowly metabolized to form TBMEHP in the presence of 0.1mg/mL of porcine hepatic carboxylesterase (PCE). This reaction was monitored at multiple time points up to 6 h and maintained linearity at an approximate rate of 1.08 pmol/min/mg esterase.

In a previous study with PCE, DEHP (50 μM) was metabolized to form MEHP at a rate of 127 pmol/min/mg protein. This rate was approximately 100 times faster than the hydrolysis of TBPH observed in this study (1.08 pmol/min/mg protein).

The prominent difference between the metabolic hydrolysis of DEHP and TBPH may be a result of steric hindrance by the fully brominated phenyl ring of TBPH.

Overall, no metabolites of TBPH were observed with HLM. From this study there is no indication that TBPH is degradated to TBMEHP in vivo. In an in-vitro experiment the hydrolysis rate of TBPH with PCE is by factor 100 slower compared with DEHP.