1-cyanoallyl acetate

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Gene mutation in bacteria (according to OECD 471, Ames test): negative in S. typhimurium TA 1535, TA 1537, TA 98 and TA 100 with and without metabolic activation

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

adopted May 26, 1983; not tested in TA 102 or E.coli WP2 uvrA

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Metabolic activation system:

s9 liver microsomal fraction from rat

Test concentrations with justification for top dose:

Exp. 1 plate incorporation: 3.3 to 100 ug/plate,
Exp. 2: pre-incubation test: 6.6-333 ug/plate (TA 1535, TA1537), 1.0-100 (TA98, TA100)

Vehicle / solvent:

water

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

water

Positive controls:

yes

Remarks:

positive controls as seen below

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

without S9 mix : TA 1535, TA100

Positive controls:

yes

Positive control substance:

other: 4-Nitro-o-phenylene-diamine

Remarks:

TA1537, TA98, without S9mix

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene

Remarks:

with S9 mix

Details on test system and experimental conditions:

METHOD OF APPLICATION: Experiment 1: in agar (plate incorporation); Experiment 2: preincubation


DETERMINATION OF CYTOTOXICITY
- Method: reduction in the number of revertants

Evaluation criteria:

- negative control in the laboratory historical range for each tester strain
- positive control chemicals produces responses in all tester strains within laboratory historical range
- Test substance is considered as mutagenic if:
- a dose-related and reproducible increase in the number of revertants occurs
- a substantial and reproducible increase for at least one test concentration (TA98, TA100 twofold increase, TA1535, TA1537 threefold increase compared to the corresponding solvent control)
Mean plate count should be at least three times the concurrent vehicle control group mean
- selected dose range should include a clearly toxic concentration or should exhibit limited solubility as demonstrated by the preliminary toxicity range-finding test or should extend to 5 mg/plate

Statistics:

none

Key result

Species / strain:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

in concentrations above 100 ug/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Conclusions:

Interpretation of results:
negative

Under the conditions tested, the test substance did not cause gene mutations in the genome of the tester strains used.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Description of key information

Genotoxicity in somatic cells in vivo (EU Method B.12, mutagenicity, Micronucleus test): negative for clastogenicity in NMRI mice

Link to relevant study records

Reference

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)

Version / remarks:

1992

GLP compliance:

yes

Type of assay:

micronucleus assay

Species:

mouse

Strain:

NMRI

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan-Winkelmann GmbH, Borchen, Germany
- Fasting period before study: 16 hours
- Housing: individually
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 6 days

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: water (aqua ad niectabilia)
- Amount of vehicle (if gavage or dermal): 10 ml/kg bw

Duration of treatment / exposure:

single dose

Frequency of treatment:

single dose

Post exposure period:

sampling time 24 and 48 h post application

Remarks:

Doses / Concentrations:
males: 31.6 mg/kg bw., females 21.5 mg/kg bw.
Basis: maximum tolerated dose as determined in a prexperiment
nominal conc.

No. of animals per sex per dose:

negative control: 12
positive control: 6
test material: 14
Evaluation: 7 test material group, 6 negative control, 5 positive control

Control animals:

other: physiological saline solution (0.9 %)

Positive control(s):

- cyclophosphamide
- Route of administration: oral (gavage)
- Doses / concentrations: 31.6 mg/kg bw

Tissues and cell types examined:

bone marrow smear of the femurs
From each animal 1000 PCEs were scored.

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION:
highest tolerable dose (higher concentrations caused severe toxic symptoms)

METHOD OF ANALYSIS:
scoring under microscope

Evaluation criteria:

test substance is considered mutagenic if it produces a statistically significant (p< 0.05) and reproducible positive response at any of the test points compared to the negative control group

Statistics:

Poisson test

Key result

Sex:

male/female

Genotoxicity:

negative

Toxicity:

yes

Vehicle controls validity:

valid

Negative controls validity:

not examined

Positive controls validity:

valid

Additional information on results:

hypokinesiahypokinesia, tremor, clonic convulsions (females only), decreased muscle tone, loss of righting reflexes. One female animal died.

RESULTS OF RANGE-FINDING STUDY
- Dose range: 31.6 - 38.3 mg/kg bw (males), 21.5-31.6 mg/kg bw (females)
- Solubility: yes
- Clinical signs of toxicity in test animals: severe toxicity symptoms, one female of the high dose died.
- Evidence of cytotoxicity in tissue analyzed: no
- Rationale for exposure: methodical reasons


RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): no
- Ratio of PCE/NCE (for Micronucleus assay): slightly lower in the test group, no clear myelotoxic effect can be deduced from the findings
control: 2.8/1.6 (24 h), 1.6/3.0 (48 h) ACA: 2.0/1.4 (24 h), 2.0/0 (48 h), values are presented as males/females

Conclusions:

Interpretation of results: negative
No significant test material related increase in micronucleated PCEs was observed. Therefore the substance is considered non mutagenic in the mouse micronucleus assay. In the positive control animals, a significant increase in the number of PCEs was observed and thus validated the test.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Mode of Action Analysis / Human Relevance Framework

Genetic toxicity in vitro:

In vitro gene mutation in bacteria:

The test item was investigated for the mutagenic potential in a Bacterial Reverse Mutation Assay (Ames test) performed according to OECD guideline 471 and in compliance with GLP (96 -0070 -DGM). S. typhimurium TA 1535, TA 1537, TA 98 and TA 100 were exposed to the test in item in the presence and absence of metabolic activation (S9 mix) in triplicate.

Based on a plate incorporation pre-experiment with and without metabolic activation, concentrations for the main mutation assay were selected at 1.0 – 333 μg/plate. The pre-experiment was reported as experiment I. A pre-incubation test with the same concentrations with and without metabolic activation served as experiment II.Solvent (water) and appropriate positive controls were included. After incubation, plates were inspected for bacterial background lawn and the number of revertant colonies was determined.

The test item did not induce a biologically relevant increase in the number of revertant colonies in any of the tested strains either with or without metabolic up to the highest tested concentration in the presence and absence of S9 mix. Positive and vehicle controls confirmed the validity of the test system and demonstrated the functionality of the S9 mix.

Thus, under the experimental conditions chosen, the test item is not mutagenic in bacteria with and without metabolic activation.

Genetic toxicity in vivo:

In a guideline study performed according to EU Method B.12 on mutagenicity and in compliance with GLP, the test item was dissolved in water and administered orally to NMRI mice (96 -0069-DGM). Based on the results of a preliminary experiment, dose levels for the mutagenicity study were selected. Groups of 14 mice/sex received an application of 21.5 -31.6 and 31.6 -38.8 mg/kg bw for females and males, respectively. A group of 6 animal/sex served as positive control and received 31.6 mg/kg bw cyclophosphamide. 24 and 48 h after the last injection, bone marrow smears were prepared. For each animal, 1000 polychromatic erythrocytes were screened for the presence of micronuclei.

One female animal died prior to the sampling time. Clinical symptoms were hpokinesiahypokinesia, tremor, clonic convulsions  in females only and decreased muscle tone, loss of righting reflexes both sexes.

There were no treatment-related changes in the frequency of micronucleated PCEs. The test item did not induce a statistically significant increase in the frequency of micronucleated PCEs. Ratio of PCE/NCE was slightly lower in the test groups. In the positive control animals, a significant increase in the number of PCEs was observed and thus validated the test. Under the conditions of the mentioned study, the test item has no genotoxic potential in vivo.

Additional information

The available information on genotoxicity in vitro (Ames test) and in vivo (micronucleus test) do not require classification according to Regulation No. (EC) 1272/2008.

No classification for genetic toxicity is warranted according to the criteria of the Globally Harmonized System of Classification and Labelling of Chemicals (GHS) of the United Nations.