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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In the Ames study, no increase in the number of revertant colonies was observed in either strain (S. typhimurium TA98, TA100, TA1535, TA1537, or E. coli WP2 uvrA/pKM101) at any concentrations of p-toluic acid and with or without metabolic activation (S9 mix). These results led to the conclusion that p-toluic acid is not a mutagen in the bacterial reverse mutation assay (Ames test) regardless of metabolic activation.

This chromosome aberration study found p-toluic acid to increase structural chromosomal aberrations.

Link to relevant study records
Reference
Endpoint:
in vitro cytogenicity / chromosome aberration study in mammalian cells
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2004
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
Deviations:
not specified
GLP compliance:
yes
Type of assay:
other: Chromosome aberration study
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material:
- Expiration date of the lot/batch:
- Purity test date:

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material:
- Stability under test conditions:
- Solubility and stability of the test substance in the solvent/vehicle:
- Reactivity of the test substance with the solvent/vehicle of the cell culture medium:

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing:
- Preliminary purification step (if any):
- Final dilution of a dissolved solid, stock liquid or gel:
- Final preparation of a solid:

FORM AS APPLIED IN THE TEST (if different from that of starting material)

TYPE OF BIOCIDE/PESTICIDE FORMULATION (if applicable)

OTHER SPECIFICS:
Species / strain / cell type:
Chinese hamster lung fibroblasts (V79)
Details on mammalian cell type (if applicable):
CELLS USED
- Source of cells: Chinese hamster lung.
- Suitability of cells:
- Cell cycle length, doubling time or proliferation index:
- Sex, age and number of blood donors if applicable:
- Whether whole blood or separated lymphocytes were used if applicable:
- Number of passages if applicable:
- Methods for maintenance in cell culture if applicable:
- Modal number of chromosomes:
- Normal (negative control) cell cycle time:

MEDIA USED
- Type and identity of media including CO2 concentration if applicable:
- Properly maintained: [yes/no]
- Periodically checked for Mycoplasma contamination: [yes/no]
- Periodically checked for karyotype stability: [yes/no)
- Periodically 'cleansed' against high spontaneous background: [yes/no]
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Test concentrations with justification for top dose:
1242 ug/mL without S-9 mix (6 hr short-term treatment)
1005 ug/mL with S-9 mix (6 hr short-term treatment)
775 ug/mL without S-9 mix (24 hr continuous treatment)
Vehicle / solvent:
- Vehicle(s) used:CMC (carboxymethyl cellulose);
- Justification for choice of solvent/vehicle:
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
not specified
True negative controls:
not specified
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
mitomycin C
Details on test system and experimental conditions:
Two plates were used for each concentration. The cells were treated continuously with the test substance for 24 hr in the absence of metabolic activation (continuous treatment) or shortly for 6 hr (short-term treatment) in the presence or absence of metabolic activation. In the short-term treatment, cells were incubated for additional 18 hr in a fresh culture medium without the test substance. The co-factor-supplemented post-mitochondrial fraction (S9 mix) was prepared from the livers of male Sprague-Dawley rats treated with phenobarbital and 5,6-benzoflavon. Mytomycin C or Benzo[a]pyrene was used as a positive control for the assay.
Colcemid (0.1 ug/ml) was added to the culture medium 2 hr before cell harvesting and then chromosome preparations were made. A hundred well-spread metaphase plates were observed under a microscope for each plate. Incidence of polyploid cells and that of cells with structural aberrations such as chromatid breaks and exchanges, chromosome breaks, exchanges and gaps, and others were recorded.
The results were considered negative when the incidence was less than 5%, and considered positive when the incidence was more than 10%.
The results were considered equivocal when the incidence was 5-10%.
Evaluation criteria:
The results were considered negative when the incidence was less than 5%, and considered positive when the incidence was more than 10%.
The results were considered equivocal when the incidence was 5-10%.
Key result
Species / strain:
Chinese hamster lung fibroblasts (V79)
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
not determined
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
In this study, an increase in structural chromosomal aberrations, attributed to a decrease of culture media pH, was observed. Therefore, the confirmation tests were conducted with or without a buffered medium, which was prepared with a double amount of sodium hydrogen carbonate.

In the main and confirmation tests conducted with buffered medium, the number of cells with structural chromosomal aberrations except gaps increased at dose of1000 ug/mL and higher after continuous treatment without metabolic activation (frequency: 5-12 %).
Remarks on result:
mutagenic potential (based on QSAR/QSPR prediction)

Colcemid (0.1 ug/ml) was added to the culture medium 2 hr before cell harvesting and then chromosome preparations were made. A hundred well-spread metaphase plates were observed under a microscope for each plate. Incidence of polyploid cells and that of cells with structural aberrations such as chromatid breaks and exchanges, chromosome breaks, exchanges and gaps, and others were recorded.
The results were considered negative when the incidence was less than 5%, and considered positive when the incidence was more than 10%.
The results were considered equivocal when the incidence was 5-10%.

Conclusions:
This chromosome aberration study found p-toluic acid to increase structural chromosomal aberrations.
Endpoint conclusion
Endpoint conclusion:
adverse effect observed (positive)

Genetic toxicity in vivo

Link to relevant study records
Reference
Endpoint:
in vivo mammalian somatic cell study: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2006
Reliability:
1 (reliable without restriction)
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
not specified
GLP compliance:
yes
Species:
mouse
Strain:
CD-1
Sex:
male
Route of administration:
oral: gavage
Vehicle:
- Vehicle used: DMSO
- Justification for choice of solvent/vehicle:
- Concentration of test material in vehicle:
- Amount of vehicle (if gavage or dermal):
- Type and concentration of dispersant aid (if powder):
- Lot/batch no. (if required):
- Purity:
Duration of treatment / exposure:
Twice with 24 hr interval
Frequency of treatment:
Twice with 24 hr interval
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Dose / conc.:
2 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Tissues and cell types examined:
Micronuclei in red blood cells.from bone marrow.
Key result
Sex:
male
Genotoxicity:
negative
Vehicle controls validity:
valid
Additional information on results:
Although there are no ADME studies, it can be predicted, based on physical chemical considerations and toxicokinetic prediction, that the substance is likely to reach the target tissue, the bone marrow. Based on these results, p-toluic acid is not anticipated to be genotoxic in vivo.
Conclusions:
Although there are no ADME studies, it can be predicted, based on physical chemical considerations and toxicokinetic prediction, that the substance is likely to reach the target tissue, the bone marrow. Based on these results, p-toluic acid is not anticipated to be genotoxic in vivo.
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

No increase in the number of revertant colonies was observed in either strain (S. typhimurium TA98, TA100, TA1535, TA1537, or E. coli WP2 uvrA/pKM101) at any concentrations of p-toluic acid and with or without metabolic activation (S9 mix). These results led to the conclusion that p-toluic acid is not a mutagen in the bacterial reverse mutation assay (Ames test) regardless of metabolic activation.”

Justification for classification or non-classification

On the basis of the mouse micronucleus study, p-toluic acid will not be classified for genetic toxicity.