Registration Dossier

Administrative data

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
supporting study
Reliability:
4 (not assignable)
Rationale for reliability incl. deficiencies:
other: Peer reviewed literature
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
publication
Title:
Quinone methode formation from para isomers of methylphenol (cresol), ethylphenol and isopropylphenol: Relationship to toxicity
Author:
Thompson DC, Perera K and London R
Year:
1995
Bibliographic source:
Chem. Res. Toxicol. 8: 55-60

Materials and methods

Objective of study:
metabolism
Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
Test compounds (2 mM each) were dissolved in DMSO and added to pre-incubated rat liver slices (30 min at 37°C in 20 mL glass scintillation vials containing 2.5 mL Krebs-Hepes buffer (pH 7.4)) at a volume of 25 µL (1% v/v) and incubated for up to 6 h. Toxicity was measured as loss of intracellular potassium using a flame photometer ( liver slices removed at appropriate time intervals dried, washed and acidified before centrigugation for 10 minutes and dilution with water).
Microsoms for metabolite formation were prepared from similar rat livers to those used from the slice assays. Incubation at 37°C was performed for various time periods and stopped by acidification. Metabolism was analysed using HPLC, MS and Proton NMR.
GLP compliance:
not specified

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
The test substances are structural analogs of the reference substance. Readacross of data is therefore applicable.
Purity: not specified but indicated to be of highest quality available.
[3H] Glutathione: 44 Ci/mmol
Radiolabelling:
yes

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories, Indianapolis, IN, USA
- Weight at study initiation: 200 - 225 g

Administration / exposure

Route of administration:
other: rats were not dosed, only liver and liver extracts used
Vehicle:
DMSO
Details on exposure:
Rat liver slices or rat liver micrsommes incubated with test material for up to 6 hours
Duration and frequency of treatment / exposure:
Up to 6 hours
Doses / concentrations
Remarks:
Doses / Concentrations:
rat liver slices: 1% v/v
Microsomes: 1 mM
No. of animals per sex per dose:
Not stated.
Control animals:
yes, concurrent vehicle
Positive control:
Not applicable
Details on study design:
Test compounds (2 mM each) were dissolved in DMSO and added to pre-incubated rat liver slices (30 min at 37°C in 20 mL glass scintillation vials containing 2.5 mL Krebs-Hepes buffer (pH 7.4)) at a volume of 25 µL (1% v/v) and incubated for up to 6 h. Toxicity was measured as loss of intracellular potassium using a flame photometer ( liver slices removed at appropriate time intervals dried, washed and acidified before centrigugation for 10 minutes and dilution with water).
Microsoms for metabolite formation were prepared from similar rat livers to those used from the slice assays. Incubation at 37°C was performed for various time periods and stopped by acidification. Metabolism was analysed using HPLC, MS and Proton NMR.
Details on dosing and sampling:
No data
Statistics:
No data

Results and discussion

Preliminary studies:
No data

Toxicokinetic / pharmacokinetic studies

Details on absorption:
No data
Details on distribution in tissues:
No data
Details on excretion:
No data

Metabolite characterisation studies

Metabolites identified:
yes
Details on metabolites:
A single glutathione conjugate peak detected for each compound. Peaks were not observed in absence of glutathione. The rate of formation of the glutathione peak mirrors the toxicity of the parent material. Splitting of the conjugate peak shows two distinct forms suggesting diastereomeric conjugates. MS confirmed that the conjugates are monoglutathione conjugates where the glutathione is attached to the benzylic carbon. This is consistent with the formation of reactive quinone methide intermediates

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): other: possible metabolic pathway in liver identified
Rat liver microsomes initiated the metabolisation of simple alkyl phenols to quinone methides.
Executive summary:

Test Guidance

No Guideline followed

Method and material

Test compounds (2 mM each) were dissolved in DMSO and added to pre-incubated rat liver slices (30 min at 37°C in 20 mL glass scintillation vials containing 2.5 mL Krebs-Hepes buffer (pH 7.4)) at a volume of 25 µL (1% v/v) and incubated for up to 6 h. Toxicity was measured as loss of intracellular potassium using a flame photometer ( liver slices removed at appropriate time intervals dried, washed and acidified before centrigugation for 10 minutes and dilution with water). Microsoms for metabolite formation were prepared from similar rat livers to those used from the slice assays. Incubation at 37°C was performed for various time periods and stopped by acidification. Metabolism was analysed using HPLC, MS and Proton NMR.

Results

A single glutathione conjugate peak was detected for each compound. Peaks were not observed in absence of glutathione. The rate of formation of the glutathione peak mirrors the toxicity of the parent material. Splitting of the conjugate peak shows two distinct forms suggesting diastereomeric conjugates. MS confirmed that the conjugates are monoglutathione conjugates where the glutathione is attached to the benzylic carbon. This is consistent with the formation of reactive quinone methide intermediates

Conclusions

Rat liver microsomes initiated the metabolisation of simple alkyl phenols to quinone methides.