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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:
November 1980
Reliability:
2 (reliable with restrictions)
Cross-reference
Reason / purpose:
reference to same study
Reference
Endpoint:
in vitro gene mutation study in bacteria
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
November 1980
Reliability:
2 (reliable with restrictions)
Justification for type of information:
ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
The structures of the target and source substances are identical and differ only with respect to the ratio of enantiomers where the target substance is a single pure L-isomer and the source substance is an equimolar mixture of L and D isomers.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
The target substance, L-Citronellyl nitrile, is a mono-constituent substance (EC No. 695-909-8, CAS no. 35931-93-2).
The source substance, DL-Citronellyl nitrile, is a mono-constituent substance (EC No. 257-288-8, CAS no. 51566-62-2).
The source and target substances are both of high purity with a low concentration of impurities.

3. ANALOGUE APPROACH JUSTIFICATION
The read across hypothesis is based on structural similarity where the only difference between target and source molecules is the enantiomeric ratio. In a non-chiral environment the target and source chemicals will have identical properties but in the chiral environment of living organisms the enantiomers may possess different carcinogenicity and teratogenicity (in a chiral environment, stereoisomers might experience selective absorption, protein binding, transport, enzyme interactions and metabolism, receptor interactions, and DNA binding). Therefore, as a precaution for the developmental toxicity endpoint it is suggested that the NOAEL 250 mg/kg bw/day for L-Citronellyl nitrile is used instead of 500 mg/kg bw/day, as it is not known which form is more potent in vivo. All other endpoints are considered to be acceptable for this substance assuming that 50% of the target compound is available in the test material.

4. DATA MATRIX
Please refer to the data matrix included in the read-across justification document attached in Section 13.2.
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
other: Ames et al. (1975)
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay
Target gene:
Histidine
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
S. typhimurium TA 1538
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
In the preliminary test conducted to assess the toxicity of the test liquid for the bacteria 0.1mg test liquid per plate appeared slightly toxic as revealed by a less dense background lawn of bacterial growth.
Top dose: 0.06mg test liquid per 0.1mL methanol per plate
Vehicle / solvent:
- Vehicle/solvent used: methanol
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
Remarks:
0mg test liquid per 0.1mL methanol per plate
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
other: hycanthone methanesulphonate (1), 2-aminoanthracene (2)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)
- Cell density at seeding: 1 - 4 x 10 9 cells / ml

DURATION
- Exposure duration:three days

NUMBER OF REPLICATIONS: triplicate
Rationale for test conditions:
Based on the test method Ames et al. (1975).
Evaluation criteria:
The colonies (revertants which are histidine independent) are counted and the background lawn of bacterial growth is examined microscopically.
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not valid
Positive controls validity:
not specified
Key result
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Remarks:
0.06mg test liquid per 0.1 mL methanol per plate, with S9 mix: background lawn of bacterial growth less dense than in concomitant control plates
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not applicable
Positive controls validity:
not specified
Key result
Species / strain:
S. typhimurium TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not valid
Positive controls validity:
not specified
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not valid
Positive controls validity:
not specified
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not valid
Positive controls validity:
not specified
Additional information on results:
See attachment.
Conclusions:
Incorporation of the test liquid with the bacteria at levels up to 0.06 mg per plate did not increase the numbers of his+ revertants with any of the five tester strains, either in the presence or in the absence of the S-9 mix.
From the present results it can be concluded that the test item did not reveal any mutagenic activity in the plate incorporation assay with S. typhimurim TA 1535, TA 1537, TA 1538, TA 98 or TA 100 in the presence or absence of the liver microsome activation system under the test conditions employed in this evaluation.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1980
Report Date:
1980

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: Ames et al. (1975)
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid
Details on test material:
Colourless liquid

Method

Target gene:
Histidine
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
S. typhimurium TA 1538
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
In the preliminary test conducted to assess the toxicity of the test liquid for the bacteria 0.1mg test liquid per plate appeared slightly toxic as revealed by a less dense background lawn of bacterial growth.
Top dose: 0.06mg test liquid per 0.1mL methanol per plate
Vehicle / solvent:
- Vehicle/solvent used: methanol
Controls
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
yes
Remarks:
0mg test liquid per 0.1mL methanol per plate
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
other: hycanthone methanesulphonate (1), 2-aminoanthracene (2)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)
- Cell density at seeding: 1 - 4 x 10 9 cells / ml

DURATION
- Exposure duration:three days

NUMBER OF REPLICATIONS: triplicate
Rationale for test conditions:
Based on the test method Ames et al. (1975).
Evaluation criteria:
The colonies (revertants which are histidine independent) are counted and the background lawn of bacterial growth is examined microscopically.

Results and discussion

Test resultsopen allclose all
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not valid
Positive controls validity:
not specified
Key result
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Remarks:
0.06mg test liquid per 0.1 mL methanol per plate, with S9 mix: background lawn of bacterial growth less dense than in concomitant control plates
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not applicable
Positive controls validity:
not specified
Key result
Species / strain:
S. typhimurium TA 1538
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not valid
Positive controls validity:
not specified
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not valid
Positive controls validity:
not specified
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
not specified
Vehicle controls validity:
not specified
Untreated negative controls validity:
not valid
Positive controls validity:
not specified
Additional information on results:
See attachment.

Applicant's summary and conclusion

Conclusions:
Incorporation of the test liquid with the bacteria at levels up to 0.06 mg per plate did not increase the numbers of his+ revertants with any of the five tester strains, either in the presence or in the absence of the S-9 mix.
From the present results it can be concluded that the test item did not reveal any mutagenic activity in the plate incorporation assay with S. typhimurim TA 1535, TA 1537, TA 1538, TA 98 or TA 100 in the presence or absence of the liver microsome activation system under the test conditions employed in this evaluation.