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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
27 Jun. - 13 Jul. 2007
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2007
Report Date:
2007

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Principles of method if other than guideline:
plate incorporation assay and preincubation test
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Test material form:
liquid: viscous
Details on test material:
- Manufacturers identification: Novares LA 300
- Substance type: organic
- Test material is 'Oligomerisation and alkylation reaction products of 2-phenylpropene and phenol' (OAPP), EC list number 700-960-7 (assigned manually to validated substances from inquiries by ECHA). Originally the substance phenol, methylstyrenated, CAS No. 68512-30-1, EC No. 270-966-8 was submitted for registration. Subsequent to substance validation, the identity of the substance was changed by ECHA.
- for additional information see respective study record
Specific details on test material used for the study:
- Name of test material (as cited in study report): Novares LA 300 (phenol, methylstyrenated)
- Lot/batch No.: 24087
- Composition of test material: composition is specified in IUCLID Sect. 13 - Assessment reports under Certificate of Analysis_Novares LA 300_phenol, methylstyrenated
- Stability under test conditions: no measured data; based on chemical structure assumed to be stable
- Storage condition of test material: room temperature, exclusion of light

Method

Species / strainopen allclose all
Species / strain / cell type:
other: S. typhimurium TA 1535, TA 1537, TA 98 and TA 102
Species / strain / cell type:
S. typhimurium TA 100
Metabolic activation:
with and without
Metabolic activation system:
Microsomal fraction prepared from induced livers of male Wistar rats, induced with phenobarbital (80 mg/kg bw) and ß-naphthoflavone (100 mg/kg bw) orally (3x)
Test concentrations with justification for top dose:
1st experiment: 10, 31.6, 100, 316, 1000, 2500, and 5000 µg/plate (all strains, +/-S9)
2nd experiment: 62.5, 125, 250, 500, 1000, and 1500 µg/plate (all strains, -S9)
62.5, 125, 250, 500, 1000, 2000, 3500, and 5000 µg/plate (all strains except TA 100, +S9)
250, 500, 1000, 2000, 4000, and 5000 µg/plate (TA 100, +S9)
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: compatible with survival of bacteria and S9 activity
Controls
Untreated negative controls:
yes
Remarks:
dist. water
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
Positive controls:
yes
Positive control substance:
other: without S9: sodium azide (TA100, TA1535); 4-NOPD (Ta98, TA1537); MMS (TA102) // with S9: 2-aminoanthracene (all strains)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (1st and 2nd experiment: plate incorporation)

NUMBER OF REPLICATIONS: 3 per concentration

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth/colony formation


Evaluation criteria:
Considered as mutagenic
- if a clear and dose-related increase in the number of revertants occurs in at least one tester with or without metabolic activation
and/or
- if a biologically relevant positive response for at least one of the dose groups occurs in at least one tester with or without metabolic activation.

An increase is considered relevant
- if in TA 100 and TA 102 mutation rate is at least twice as high as the rate of the solvent control;
- if in TA 98, TA 1535, and TA 1537 the mutation rate is at least 3x higher than that of the solvent control.


Statistics:
According to the OECD guidelines, the biological relevance is the criterion for the interpretation of the results: a statistical evaluation was not considered necessary under this premise (report p. 21).

Results and discussion

Test resultsopen allclose all
Species / strain:
other: S. typhimurium TA 1535, TA 1537, TA 98 and TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Remarks:
reproducible in both tests
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
>= 250 µg/pl., depending on strain and presence of S9
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with
Genotoxicity:
ambiguous
Remarks:
reproducible in both tests at >= 2000 µg/pl.
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
reproducible in both tests at >= 1000 µg/pl.
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid

Any other information on results incl. tables

Summary:

No biologically relevant increases in revertant colony numbers were observed in the four tester strains, TA 1535, TA 1537, TA 98 and TA 102, following treatment with Novares LA 300, either in the presence or in the absence of metabolic activation.

An unremarkable dose-related increase in the mutation rate was found in tester strain TA 100 under metabolic activation: a maximum mutation factor of 2.8 was reached at a dose of 4000 µg/plate. The mutagenic doses were cytotoxic as well.

All reference mutagens induced distinct increases of revertant colonies indicating the validity of the experiment.

Applicant's summary and conclusion

Conclusions:
In one (TA 100) out of five tester strains (other strains TA 1535, TA 1537, TA 98, TA 102), a weak mutagenic dose-related response was observed with metabolic activation. The maximum mutation factor reached was 2.8 at the test substance concentration of 4000 µg/plate. The mutagenic doses were cytotoxic as well.
Overall, the results is considered to be "ambiguous".