Undec-10-enal

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Ames assay:

The test chemical is not mutagenic in the bacterium Salmonella typhimurium LT-2 strains TA 98, TA 100, TA 1535 and TA 1537 with and without S9 metabolic activation system.

In vitro mammalian chromosome aberration study:

The test chemical did not induce chromosomal aberration in the mammalian cell line used and hence it is not likely to classify as a gene mutant in vitro.

In vitro mammalian cell gene mutation assay:

In a gene toxicity test, Chinese Hamster Ovary (CHO) cells were exposed to the test chemical in the concentration of 0, 0.5, 1.0, 2.5 or 5.0 mM and with S9-induced metabolic activation for 3 hours. The results showed that there was evidence of cytotoxicity after treatment with 2.5 or 5.0 mM. Independently of tested concentration, the results showed no evidence of gene toxicity. Therefore, it is considered that the test chemical in the concentration of 0, 0.5, 1.0, 2.5 or 5.0 mM does not cause genetic mutation(s) when CHO cells are exposed to the test chemical in the presence of metabolic activation.

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

Experimental data from various test chemicals

Justification for type of information:

Data for the target chemical is summarized based on various test chemicals

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

equivalent or similar to guideline

Guideline:

other: Refer below principle

Principles of method if other than guideline:

WoE derived based on the experimental data from various test chemicals

GLP compliance:

not specified

Type of assay:

bacterial gene mutation assay

Target gene:

No data

Species / strain / cell type:

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

Remarks:

1

Details on mammalian cell type (if applicable):

No data

Additional strain / cell type characteristics:

not specified

Species / strain / cell type:

S. typhimurium, other: TA92, TA1535, TA100, TA1537, TA94 and TA98

Remarks:

2

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

with and without

Metabolic activation system:

Liver fraction (S-9) from Aroclor 1254 or methylcholanthrene-induced rats

Test concentrations with justification for top dose:

1. 3 µmole/plate
2. At three different doses with 1000 µg/plate being the maximum dose concentration

Vehicle / solvent:

1. - Vehicle(s)/solvent(s) used: Ethanol
- Justification for choice of solvent/vehicle: No data available

2. - Vehicle(s)/solvent(s) used: Dimethyl sulphoxide
- Justification for choice of solvent/vehicle: No data available

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

other: N-methyl-N'-nitro-N-nitrosoguanidin (without metabolic activation) and 2-aminoanthracene (with activation)

Remarks:

1

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

not specified

Positive control substance:

not specified

Remarks:

2

Details on test system and experimental conditions:

1. METHOD OF APPLICATION: Spot test (in agar)

DURATION
- Preincubation period: No data available
- Exposure duration: No data available
- Expression time (cells in growth medium): No data available
- Selection time (if incubation with a selection agent): No data available
- Fixation time (start of exposure up to fixation or harvest of cells): No data available

SELECTION AGENT (mutation assays): No data available
SPINDLE INHIBITOR (cytogenetic assays): No data available
STAIN (for cytogenetic assays): No data available

NUMBER OF REPLICATIONS: No data available

NUMBER OF CELLS EVALUATED: No data available

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data available

OTHER EXAMINATIONS:
- Determination of polyploidy: No data available
- Determination of endoreplication: No data available
- Other: No data available

OTHER: No data available

2. METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period: 20 mins at 37OC
- Exposure duration: 2 days
- Expression time (cells in growth medium): 2 days
- Selection time (if incubation with a selection agent): No data available
- Fixation time (start of exposure up to fixation or harvest of cells): No data available

SELECTION AGENT (mutation assays): No data available

SPINDLE INHIBITOR (cytogenetic assays): No data available

STAIN (for cytogenetic assays): No data available

NUMBER OF REPLICATIONS: Duplicate

NUMBER OF CELLS EVALUATED: No data available

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data available

OTHER EXAMINATIONS:
- Determination of polyploidy: No data available

- Determination of endoreplication: No data available

- Other: No data available

OTHER: No data available

Rationale for test conditions:

No data

Evaluation criteria:

1. 1. Increase in the number of spontaneous revertants
2. The presence of the rfa-mutation was checked by crystal violet inhibition

2. The result was considered positive if the number of colonies found was twice the number in the control.

3. The plates were observed for number of revertants/plate

Statistics:

No data available

Species / strain:

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

Remarks:

1

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Species / strain:

S. typhimurium, other: TA92, TA1535, TA100, TA1537, TA94 and TA98

Remarks:

2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Additional information on results:

No data

Remarks on result:

other: No mutagenic potential

Conclusions:

The test compound is not mutagenic in the bacterium Salmonella typhimurium LT-2 strains TA 98, TA 100, TA 1535 and TA 1537 with and without S9 metabolic activation system.

Executive summary:

Data available for the test chemicals was reviewed to determine the mutagenic nature of the test chemical. The studies are as mentioned below:

In an in vitro bacterial gene toxicity study, the genotoxic effects of the test chemical was evaluated using Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 in the presence and absence of S9 metabolic activation system. The test chemical was dissolved in ethanol. The bacterial strains were exposed to the test chemical at a concentration of 3 µmole/plate with and without S9 metabolic activation system. The results showed that Undecanal did not induce reversion of mutant strains and hence it is not mutagenic in the bacterium Salmonella typhimurium LT-2 strains TA 98, TA 100, TA 1535 and TA 1537 with and without S9 metabolic activation system.

In another study, Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical. The study was performed using S. typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 with and without S9 metabolic activation system. The test was performed as per the preincubation assay at three different concentrations with 1 mg/plate being the maximum concentration. The chemical was dissolved in DMSO. Preincubation was performed for 20 mins and the exposure duration was for 48 hrs. The result was considered positive if the number of colonies found was twice the number in the control (exposed to the appropriate solvent or untreated). The test chemical did not induce a doubling of revertant colonies over the control using S. typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 in the presence and absence of S9 metabolic activation system and hence the chemical is not likely to classify as a gene mutant in vitro.

Based on the data available, the test chemical is not mutagenic in the bacterium Salmonella typhimurium LT-2 strains TA 98, TA 100, TA 1535 and TA 1537 with and without S9 metabolic activation system.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

Experimental data from various test chemicals

Justification for type of information:

Data for the target chemical is summarized based on the various test chemicals

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Principles of method if other than guideline:

WoE derived based on the experimental data from the various test chemicals

GLP compliance:

not specified

Type of assay:

other: In vitro chromosome aberration assay

Target gene:

No data

Species / strain / cell type:

mammalian cell line, other: Chinese hamster fibroblast cell line CHL

Remarks:

1

Details on mammalian cell type (if applicable):

- Type and identity of media: Minimum
Essential Medium (MEM; GIBCO) supplemented by 10% calf serum
- Properly maintained: yes by 4 day passages
- Periodically checked for Mycoplasma contamination: No data available
- Periodically checked for karyotype stability: No data available
- Periodically "cleansed" against high spontaneous background: No data available

Additional strain / cell type characteristics:

not specified

Species / strain / cell type:

hepatocytes: Primary cultures of rat hepatocytes harvested from female Fischer 344 rats

Details on mammalian cell type (if applicable):

- Type and identity of media: Minimum essential medium (MEM) with Earle's salts and non-essential amino acids, and eventually also with epidermal growth factor.
- Properly maintained: No data available
- Periodically checked for Mycoplasma contamination: No data available
- Periodically checked for karyotype stability: No data available
- Periodically "cleansed" against high spontaneous background: No data available

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

without

Metabolic activation system:

Metabolic activation system

Test concentrations with justification for top dose:

1. At three different doses with 0.125 mg/mL being the maximum dose concentration
2. 0.0, 0.1, 1.0, 10.0 or 100.0 µM

Vehicle / solvent:

1. - Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: The chemical was soluble in DMSO

2. No data

2. No data

Untreated negative controls:

yes

Remarks:

Untreated cells served as negative control

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

not specified

Positive control substance:

not specified

Remarks:

1

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

not specified

Positive control substance:

not specified

Remarks:

2

Details on test system and experimental conditions:

1. METHOD OF APPLICATION: in medium

DURATION
- Preincubation period: No data
- Exposure duration: 48 hrs
- Expression time (cells in growth medium): 48 hrs
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): Giemsa solution (1.5%, pH 6.8)
SPINDLE INHIBITOR (cytogenetic assays): Colcemid
STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: No data

NUMBER OF CELLS EVALUATED: 100 well spread metaphases

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data

OTHER EXAMINATIONS:
- Determination of polyploidy: Yes
- Determination of endoreplication: No data
- Other: No data

OTHER: No data

2. METHOD OF APPLICATION: In medium

DURATION
- Preincubation period: No data available
- Exposure duration: 3 hrs
- Expression time (cells in growth medium): 51 hrs
- Selection time (if incubation with a selection agent): No data available
- Fixation time (start of exposure up to fixation or harvest of cells): No data available

SELECTION AGENT (mutation assays): Hoechst 33258 and Giemsa
SPINDLE INHIBITOR (cytogenetic assays): Colcemid
STAIN (for cytogenetic assays): DAPI

NUMBER OF REPLICATIONS: No data available

NUMBER OF CELLS EVALUATED: 20 well-spread metaphases were scored

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: Mitotic index, 1000 cells were then scored, to determine the percentage of mitotic cells (= mitotic index) and the percentage of cells with micronuclei.

OTHER EXAMINATIONS: No data available
- Determination of polyploidy: No data available
- Determination of endoreplication: No data available
- Other: No data available

OTHER: No data available

Rationale for test conditions:

No data

Evaluation criteria:

1. The incidence of polyploid cells as well as of cells with structural chromosomal aberrations such as chromatid or chromosome gaps, breaks, exchanges, ring formations, fragmentations and others, was recorded on each culture plate. The results were considered to be negative if the incidence was less than 4.9%, equivocal if it was between 5.0 and 9.9%, and positive if it was more than 10.0%.

2. No data

Statistics:

1. No data

2. Student's t-test for independent variables was used to calculate the levels of significance for mitotic index, chromosomal aberrations and sister-chromatid exchange. For statistical analysis of the micronuclei Student's t-test for dependent variables was used.

Species / strain:

mammalian cell line, other: Chinese hamster fibroblast cell line CHL

Remarks:

1

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

not specified

Species / strain:

hepatocytes: Primary cultures of rat hepatocytes harvested from female Fischer 344 rats

Remarks:

2

Metabolic activation:

not specified

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Additional information on results:

1. TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No data
- Effects of osmolality: No data
- Evaporation from medium: No data
- Water solubility: No data
- Precipitation: No data
- Other confounding effects: No data

RANGE-FINDING/SCREENING STUDIES: The maximum dose of each sample was selected by a preliminary test in which the dose needed or 50% cell-growth inhibition was estimated using a cell densitometer

COMPARISON WITH HISTORICAL CONTROL DATA: No data

ADDITIONAL INFORMATION ON CYTOTOXICITY: No data

2. No data

Remarks on result:

other: No mutagenic potential

Conclusions:

The test chemical did not induce chromosomal aberration in the mammalian cell line used and hence it is not likely to classify as a gene mutant in vitro.

Executive summary:

Data available for the test chemicals was reviewed to determine the mutagenic nature of the test chemical. The studies are as mentioned below:

Chromosomal aberration study was performed to determine the mutagenic nature of the test chemical. The cells were exposed to the test material at three different doses with 0.125 mg/mL being the maximum concentration for 48hr. Colcemid (final concn 0.2 µg/ml) was added to the culture 2 hr before cell harvesting. The cells were then trypsinized and suspended in a hypotonic KCI solution (0.075 M) for 13 min at room temperature. After centrifugation the cells were fixed with acetic acid-methanol (1:3, v/v) and spread on clean glass slides. After air-drying, the slides were stained with Giemsa solution for 12-15 min. A hundred well-spread metaphases were observed under the microscope. In the present studies, no metabolic activation systems were applied. The incidence of polyploid cells as well as of cells with structural chromosomal aberrations such as chromatid or chromosome gaps, breaks, exchanges, ring formations, fragmentations and others, was recorded on each culture plate. Untreated cells and solvent-treated cells served as negative controls, in which the incidence of aberrations was usually less than 3.0%. The results were considered to be negative if the incidence was less than 4.9%, equivocal if it was between 5.0 and 9.9%, and positive if it was more than 10.0%. The test chemical did not induce chromosomal aberration in Chinese hamster fibroblast cell line CHL and hence it is not likely to classify as a gene mutant in vitro.

Cytotoxic and mutagenic nature of the test chemicall was tested using female Fischer 344 rat hepatocytes. The experiments was initiated by adding the test chemical at a dose level of 0.0, 0.1, 1.0, 10.0 or 100.0 µM to the cultures. After an incubation time of 3 h, the medium was removed, the dishes were washed twice and then 5 ml final medium with epidermal growth factor (40 ng/ml) and bromodesoxyuridine (10 µM) was added. 48 h later, Colcemid (0.4 µg/ml) was added, the cultures were incubated for a further 3 h and then the cytogenetic analyses was performed. In brief, for chromosomal aberrations, the cells were detached with collagenase solution, the cells were fixed on slides with cold methanol-glacial acetic acid and stained with Hoechst 33258 followed by Giemsa. At least 20 well-spread metaphases were scored. The number of chromosomal aberrations is given per diploid cell (42 chromosomes). For the determination of the mitotic index and the number of micronuclei, the cells were fixed on the culture dishes and stained with DAPI. 1000 cells were then scored, to determine the percentage of mitotic cells (= mitotic index) and the percentage of cells with micronuclei. The test chemical does not induce any genotoxic effects since it failed to induce chromosome aberrations in the rat hepatocytes and hence it is negative for gene mutation in vitro.

Based on the data available, the test chemical did not induce chromosomal aberration in the mammalian cell line used and hence it is not likely to classify as a gene mutant in vitro.

Reference 3

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

30-04-2015 to 03-11-2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

Data is from study report

Qualifier:

according to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Principles of method if other than guideline:

Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical in the presence of S9 metabolic activation system

GLP compliance:

no

Type of assay:

mammalian cell gene mutation assay

Target gene:

Cells deficient in hypoxanthine-guanine phosphoribosyl transferase (HPRT) due to the mutation HPRT+/- to HPRT-/- are resistant to cytotoxic effects of 6-thioguanine (TG). HPRT proficient cells are sensitive to TG (which causes inhibition of cellular metabolism and halts further cell division since HPRT enzyme activity is important for DNA synthesis), so mutant cells can proliferate in the presence of TG, while normal cells, containing hypoxanthine-guanine phosphoribosyl transferase cannot.

This in vitro test is an assay for the detection of forward gene mutations at the in hypoxanthine-guanine phosphoribosyl transferase (HPRT) locus on the X chromosomes of hypodiploid, modal No. 20, CHO cells. Gene and chromosome mutations are considered as an initial step in the carcinogenic process.
The hypodiploid CHO cells are exposed to the test item with and without exogenous metabolic activation. Following an expression time the descendants of the treated cell population are monitored for the loss of functional HPRT enzyme.
HPRT catalyses the transformation of the purine analogues 6-thioguanine (TG) rendering them cytotoxic to normal cells. Hence, cells with mutations in the HPRT gene cannot phosphoribosylate the analogue and survive treatment with TG.

Therefore, mutated cells are able to proliferate in the presence of TG whereas the non-mutated cells die. However, the mutant phenotype requires a certain period of time before it is completely expressed. The phenotypic expression is achieved by allowing exponential growth of the cells for 7 days.

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

- Cell line used: Chinese Hamster Ovary (CHO) cells
- Type and identity of media: Ham's F-12K (Kaighn's) Medium containing 2 mM L-Glutamine supplemented with 10% Fetal Bovine Serum and 1% Penicillin-Streptomycin (10,000 U/mL).
- Properly maintained: Yes
- Periodically checked for Mycoplasma contamination: Not applicable
- Periodically checked for karyotype stability: Not applicable

Additional strain / cell type characteristics:

other: Hypodiploid, modal No. 20

Cytokinesis block (if used):

No data

Metabolic activation:

with

Metabolic activation system:

S9 liver microsomal fraction obtained from Arcolor 1254-induced male Sprague-Dawley rats (Supplier: Molecular Toxicology Inc. via Trinova Biochem GmbH, Giessen, Germany)

Test concentrations with justification for top dose:

0, 0.5, 1.0, 2.5 or 5.0 mM

Vehicle / solvent:

Vehicle(s)/solvent(s) used: DMSO
Justification for choice of solvent/ vehicle: The test chemical was easily dissolved in DMSO

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

7,12-dimethylbenzanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION: In medium with pre-incubation

DURATION
Pre-incubation
One week involving 3 days of incubation with Hypoxanthine-aminopterin-thymidine (HAT) in medium as a mutant cleansing stage, followed by overnight incubation with hypoxanthine-thymidine (HT) in medium prior to a 3-4 days incubation in regular cell medium. After seeding and prior to treatment, the mutant-free cells were incubated for an additional of 24 hours.

Exposure duration
3 hours

Expression time
7 days

Selection time
14 days

Fixation time
7 days (harvest of cells)

SELECTION AGENT (mutation assays): 6-thioguanine (TG)

SPINDLE INHIBITOR (cytogenetic assays): Not applicable

STAIN (for cytogenetic assays): Crystal violet

NUMBER OF REPLICATIONS: A minimum of 2 replicates per dose concentration including negative and positive control.

NUMBER OF CELLS EVALUATED: 5 x 10 E5 cells were plated 7 days after treatment and whatever cells left, after 14 days of incubation with the selection medium, were evaluated.

DETERMINATION OF CYTOTOXICITY
Cytotoxicity test
After being exposed to the test chemical for 3 hours, in the absence or presence of S9, cells were trypsinized and 0.5 x 10 E5 cells per well was seeded in duplicates from two parallel duplicate cultures into 6-well plates in fresh medium. The relative total growth and cytotoxicity was evaluated 24 and 48 hours after seeding.

Rationale for test conditions:

No data

Evaluation criteria:

Chinese Hamster Ovary Cells (CHO) were observed for gene mutation caused by the test compound

Statistics:

No data

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with

Genotoxicity:

negative

Remarks:

<0.05 M

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Effects of pH: No data
- Effects of osmolality: No data
- Evaporation from medium: No data
- Water solubility: No data
- Precipitation: No data
- Definition of acceptable cells for analysis: No data
- Other confounding effects: No data

RANGE-FINDING/SCREENING STUDIES: Completed without S9 metabolic activation. A range of test concentrations (0, 0.0005, 0.001, 0.005, 0.01, 0.05, 0.1, 0.5 or 1.0 mM) was applied 24 hours after seeding to single cultures in fresh medium in 96-well plates. The cell population (control and treated cells) were assessed 24 and 48 hours after treatment using the colorimetric assay MTT and the BCA assay to assess cell viability and total protein concentration, respectively. From the basis of these results, the test concentrations of the chemical was chosen to be included in the gene toxicity test. Since a slight cytotoxicity was evident at the highest tested concentration in this preliminary dose-finding test further testing concentrations were adapted to have a maximum test concentration of 5.0 mM. Since the test chemical was dissolved in DMSO, higher concentrations of the test chemical than the concentration mentioned above would result in a toxic effect of DMSO. The test chemical could only be dissolved in 100% DMSO.

CYTOKINESIS BLOCK (if used)
- Distribution of mono-, bi- and multi-nucleated cells: No data

NUMBER OF CELLS WITH MICRONUCLEI
- Number of cells for each treated and control culture: No data
- Indication whether binucleate or mononucleate where appropriate: No data

HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Positive historical control data: No data
- Negative (solvent/vehicle) historical control data: No data

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Measurement of cytotoxicity used: No data
- Other observations when applicable:No data

Remarks on result:

other: No mutagenic potential

Conclusions:

The test chemical in the concentration of 0, 0.5, 1.0, 2.5 or 5.0 mM did not show any evidence of gene toxicity when CHO cells were exposed to the test chemical.

Executive summary:

In a gene toxicity test, Chinese Hamster Ovary (CHO) cells were exposed to the test chemical in the concentration of 0, 0.5, 1.0, 2.5 or 5.0 mM and S9-induced metabolic activation for 3 hours. The results showed that there was evidence of cytotoxicity after treatment with 2.5 or 5.0 mM. Independently of tested concentration, the results showed no evidence of gene toxicity. Therefore, it is considered that the test chemical in the concentration of 0, 0.5, 1.0, 2.5 or 5.0 mM does not cause genetic mutation(s) when CHO cells are exposed to the test chemical in the presence of metabolic activation.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Gene mutation in vitro:

Data available for the test chemical was reviewed to determine the mutagenic nature of the test chemical. The studies are as mentioned below:

Ames assay:

In an in vitro bacterial gene toxicity study, the genotoxic effects of the test chemical was evaluated using Salmonella typhimurium strains TA98, TA100, TA1535 and TA1537 in the presence and absence of S9 metabolic activation system. The test chemical was dissolved in ethanol. The bacterial strains were exposed to the test chemical at a concentration of 3 µmole/plate with and without S9 metabolic activation system. The results showed that Undecanal did not induce reversion of mutant strains and hence it is not mutagenic in the bacterium Salmonella typhimurium LT-2 strains TA 98, TA 100, TA 1535 and TA 1537 with and without S9 metabolic activation system.

In another study, Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical. The study was performed using S. typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 with and without S9 metabolic activation system. The test was performed as per the preincubation assay at three different concentrations with 1 mg/plate being the maximum concentration. The chemical was dissolved in DMSO. Preincubation was performed for 20 mins and the exposure duration was for 48 hrs. The result was considered positive if the number of colonies found was twice the number in the control (exposed to the appropriate solvent or untreated). The test chemical did not induce a doubling of revertant colonies over the control using S. typhimurium strains TA92, TA1535, TA100, TA1537, TA94 and TA98 in the presence and absence of S9 metabolic activation system and hence the chemical is not likely to classify as a gene mutant in vitro.

Based on the data available, the test chemical is not mutagenic in the bacterium Salmonella typhimurium LT-2 strains TA 98, TA 100, TA 1535 and TA 1537 with and without S9 metabolic activation system.

In vitro mammalian chromosome aberration study:

Chromosomal aberration study was performed to determine the mutagenic nature of the test chemical. The cells were exposed to the test material at three different doses with 0.125 mg/mL being the maximum concentration for 48hr. Colcemid (final concn 0.2 µg/ml) was added to the culture 2 hr before cell harvesting. The cells were then trypsinized and suspended in a hypotonic KCI solution (0.075 M) for 13 min at room temperature. After centrifugation the cells were fixed with acetic acid-methanol (1:3, v/v) and spread on clean glass slides. After air-drying, the slides were stained with Giemsa solution for 12-15 min. A hundred well-spread metaphases were observed under the microscope. In the present studies, no metabolic activation systems were applied. The incidence of polyploid cells as well as of cells with structural chromosomal aberrations such as chromatid or chromosome gaps, breaks, exchanges, ring formations, fragmentations and others, was recorded on each culture plate. Untreated cells and solvent-treated cells served as negative controls, in which the incidence of aberrations was usually less than 3.0%. The results were considered to be negative if the incidence was less than 4.9%, equivocal if it was between 5.0 and 9.9%, and positive if it was more than 10.0%. The test chemical did not induce chromosomal aberration in Chinese hamster fibroblast cell line CHL and hence it is not likely to classify as a gene mutant in vitro.

Cytotoxic and mutagenic nature of the test chemicall was tested using female Fischer 344 rat hepatocytes. The experiments was initiated by adding the test chemical at a dose level of 0.0, 0.1, 1.0, 10.0 or 100.0 µM to the cultures. After an incubation time of 3 h, the medium was removed, the dishes were washed twice and then 5 ml final medium with epidermal growth factor (40 ng/ml) and bromodesoxyuridine (10 µM) was added. 48 h later, Colcemid (0.4 µg/ml) was added, the cultures were incubated for a further 3 h and then the cytogenetic analyses was performed. In brief, for chromosomal aberrations, the cells were detached with collagenase solution, the cells were fixed on slides with cold methanol-glacial acetic acid and stained with Hoechst 33258 followed by Giemsa. At least 20 well-spread metaphases were scored. The number of chromosomal aberrations is given per diploid cell (42 chromosomes). For the determination of the mitotic index and the number of micronuclei, the cells were fixed on the culture dishes and stained with DAPI. 1000 cells were then scored, to determine the percentage of mitotic cells (= mitotic index) and the percentage of cells with micronuclei. The test chemical does not induce any genotoxic effects since it failed to induce chromosome aberrations in the rat hepatocytes and hence it is negative for gene mutation in vitro.

Based on the data available, the test chemical did not induce chromosomal aberration in the mammalian cell line used and hence it is not likely to classify as a gene mutant in vitro.

In vitro mammalian cell gene mutation assay:

In a gene toxicity test, Chinese Hamster Ovary (CHO) cells were exposed to the test chemical in the concentration of 0, 0.5, 1.0, 2.5 or 5.0 mM and with and without S9-induced metabolic activation for 3 hours. The results showed that there was evidence of cytotoxicity after treatment with 2.5 or 5.0 mM. Independently of tested concentration, the results showed no evidence of gene toxicity. Therefore, it is considered that the test chemical in the concentration of 0, 0.5, 1.0, 2.5 or 5.0 mM does not cause genetic mutation(s) when CHO cells are exposed to the test chemical in the presence and absence of metabolic activation.

Based on the data available, the test chemical does not ehxibit gene mutation in vitro. Hence the test chemical is not likely to classify as a gene mutant as per the criteria mentioned in CLP regulation.

Justification for classification or non-classification

Based on the data available, the test chemical does not ehxibit gene mutation in vitro. Hence the test chemical is not likely to classify as a gene mutant as per the criteria mentioned in CLP regulation.