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Genetic toxicity in vivo

Link to relevant study records
Reference
Endpoint:
in vivo mammalian cell study: DNA damage and/or repair
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2013
Reliability:
1 (reliable without restriction)
Qualifier:
according to
Guideline:
OECD Guideline 486 (Unscheduled DNA Synthesis (UDS) Test with Mammalian Liver Cells in vivo)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
unscheduled DNA synthesis
Species:
rat
Strain:
Wistar
Sex:
male/female
Route of administration:
oral: gavage
Frequency of treatment:
The study was performed in two parts, in Experiment 1 perfusion of the livers commenced approximately 16 hours after dosing and in Experiment 2 perfusion was performed approximately 4 hours after dosing.
Remarks:
Doses / Concentrations:
0 mg/kg bw
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
1000 mg/kg bw
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
2000 mg/kg bw
Basis:
nominal conc.
No. of animals per sex per dose:
4
Sex:
male
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Positive controls validity:
valid
Conclusions:
Interpretation of results (migrated information): negative
The test item was considered to be non-genotoxic under the conditions of this study.
Executive summary:

Introduction. A study was performed to assess the potential of the test item to induce DNA repair in isolated rat hepatocytes following in vivo administration. The method used has been designed to be compatible with the OECD Guidelines for Testing of Chemicals No. 486 and follows the recommendations of the UKEMS Sub-Committee on Guidelines for Mutagenicity Testing: Report, Part II revised (Supplementary Mutagenicity Tests: UKEMS recommended procedures, 1993).

Methods. A range-finding test was performed to find suitable dose levels of the test item, route of administration and to determine if the there was any differences in toxicity in male and female rats. As there was no difference in toxicity between sexes the main test was performed using male rats only. The UDS assay was conducted using the oral route of administration using only male animals and with the test item at the maximum dose level of 2000 mg/kg, with 1000 mg/kg as the lower dose level. The study was performed in two parts, in Experiment 1 perfusion of the livers commenced approximately 16 hours after dosing and in Experiment 2 perfusion was performed approximately 4 hours after dosing. Following perfusion the liver hepatocytes were processed to give stained slides which were then scored using a microscope and an automated image analysis system. The method used for scoring the hepatocytes was an area counting method which is compatible with the UKEMS guidelines and OECD test method. Further groups of rats were given a single oral dose of arachis oil, or dosed with 2-acetylaminofluorene (2AAF) at 16 hours or N,Nā€™-dimethylhydrazine dihydrochloride (NDHC) at 4 hours to serve as vehicle and positive controls respectively.

Results. There were no marked increases in the incidence of unscheduled DNA synthesis in animals dosed with the test item at either time point. The positive controls produced marked increases in net nuclear grain counts and in the incidence of cells in repair, and the vehicle control groups gave acceptable values for net nuclear grain counts.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vivo:

AMES- The genotoxicity data on Pigment Red 3 are characterized by a certain inconsistency.Three out of 6 bacterial gene mutation () assays were positive and one test revealed an equivocal result. However there was neither consistency with regard to the requirement of metabolic activation nor with regard to the strains affected. One study revealed mutagenic activity in TA 1537 with and without metabolic activation, whereas the other three studies revealed mutagenic activity with metabolic activation only. In addition different studies revealed effects in different S. typhimurium strains: in one study TA 1537 and TA 1538 were affected, whereas other studies revealed mutagenic activity in TA 1537, TA 100, or in TA 100 and TA 98. A common feature of all positive tests was that mutagenic responses were only weak and only observed at high concentrations characterized by heavy precipitation of the test article. This may lead to the speculation that impurities may have been responsible for the observed weak mutagenic activity, especially as was reported that purification of the test-article results in the loss of mutagenic activity. Still, weak or at least equivocal mutagenic activity was also observed in highly pure (> 99%) preparations. It is however worth to note that the weak mutagenic activity with highly pure PR3 was only observed in a special modification of the Ames test which aims to reduce the azo-groups (Prival test). As an overall conclusion, a weak gene mutation potential of PR3 in bacteria can not be excluded. It might be sensible to check the gene mutation potential in mammalian cells.

Clear and negative studies for mutaginicity have been seen in vitro in OECD 473 and OECD 476. In vivo there were no marked increases in the incidence of unscheduled DNA synthesis in animals dosed with the test item (OECD 486).

Conclusion: the test item is not considered to be mutagenic.


Justification for selection of genetic toxicity endpoint
No adverse effects have been seen in vitro in OECD 473, OECD 476 while ambiguous, possitive and negative test results have been seen in Ames tests with and without metabolic activation (OECD 471). In vivo there were no marked increases in the incidence of unscheduled DNA synthesis in animals dosed with the test item (OECD 486)

Justification for classification or non-classification

Pigment Red 3 does not have to be classified for mutagenicity according to the criteria laid down in the EU Dangerous Substances Directive (67/548/EEC) and in the EU Classification Labelling and Packaging Regulation (1272/2008/EC) because no adverse effects due to the test item have been seen in vitro in OECD 473 and OECD 476 studies while ambiguous, possitive and negative test results have been seen in Ames tests with and without metabolic activation (OECD 471). In vivo there were no marked increases in the incidence of unscheduled DNA synthesis in animals dosed with the test item (OECD 486).