Octa-1,7-diene

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Octa-1,7-diene

EC Number:

223-054-9

EC Name:

Octa-1,7-diene

Cas Number:

3710-30-3

Molecular formula:

C8H14

IUPAC Name:

octa-1,7-diene

Test material form:

liquid

Method

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

S9 mix (rat)

Test concentrations with justification for top dose:

Pre-Experiment/Experiment I:
3; 10; 33; 100; 333; 1000; 2500; and 5000 μg/plate
Experiment II:
Salmonella strains: 1; 3; 10; 33; 100; 333; 1000; 2500; and 5000 μg/plate
WP2 uvrA: 10; 33; 100; 333; 1000; 2500; and 5000 μg/plate

Vehicle / solvent:

DMSO (>99%)

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Controls 2

Positive controls:

yes

Remarks:

10 μg/plate

Positive control substance:

sodium azide

Remarks:

TA 1535, TA 100 - Without metabolic activation

Controls 3

Positive controls:

yes

Remarks:

10 μg/plate in strain TA 98, 50 μg/plate in strain TA 1537

Positive control substance:

other: 4-nitro-o-phenylene-diamine, 4-NOPD

Remarks:

TA 1537, TA 98 - Without metabolic activation

Controls 4

Positive controls:

yes

Remarks:

2.0 μL/plate

Positive control substance:

methylmethanesulfonate

Remarks:

WP2 uvrA - Without metabolic activation

Controls 5

Positive controls:

yes

Remarks:

2.5 μg/plate (10.0 μg/plate in WP2 uvrA)

Positive control substance:

other: 2-aminoanthracene, 2-AA

Remarks:

TA 1535, TA 1537, TA 98, TA 100, WP2 uvrA - With metabolic activation

Details on test system and experimental conditions:

METHOD OF APPLICATION:
- 1st main experiment: plate incorporation
- 2nd main experiment: preincubation

DURATION:
- 1st main experiment: 48 hours incubation (37°C)
- 2nd main experiment: 30 min preincubation (30°C), 48 hours incubation (37°C)

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY:
Toxicity of the test item can be evident as a reduction in the number of spontaneous revertants or a clearing of the bacterial background lawn.

Evaluation criteria:

VALIDITY CRITERIA:
The Salmonella typhimurium and Escherichia coli reverse mutation assay is considered acceptable if it meets the following criteria:
- regular background growth in the negative and solvent control
- the spontaneous reversion rates in the negative and solvent control are in the range of our historical data
- the positive control substances should produce a significant increase in mutant colony frequencies
- a minimum of five analysable dose levels should be present with at least three dose levels showing no signs of toxic effects, evident as a reduction in the
number of revertants below the indication factor of 0.5.

CRITERIA FOR POSITIVE RESPONSE:
A test item is considered as a mutagen if a biologically relevant increase in the number of revertants exceeding the threshold of twice (strains TA 98, TA 100, and WP2 uvrA) or thrice (strains TA 1535 and TA 1537) the colony count of the corresponding solvent control is observed.
A dose dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration.
An increase exceeding the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.
A dose dependent increase in the number of revertant colonies below the threshold is regarded as an indication of a mutagenic potential if reproduced in an independent second experiment. However, whenever the colony counts remain within the historical range of negative and solvent controls such an increase is not considered biologically relevant.

Statistics:

According to the OECD guideline 471, a statistical analysis of the data is not mandatory.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Test results 5

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

The plates incubated with the test item showed reduced background growth at the following concentrations (μg/plate):

Strain	Experiment I		Experiment II
	without S9 mix	with S9 Mix	without S9 mix	with S9 Mix
TA 1535	333 - 5000	333 - 5000	100 - 5000	333 - 5000
TA 1537	333 - 5000	1000 - 5000	333 - 5000	333 - 5000
TA 98	333 - 5000	1000 - 5000	100 - 5000	333 - 5000
TA 100	333 - 5000	333 - 5000	1000 - 5000	333 - 5000
WP2 uvrA	2500 - 5000	2500 - 5000	5000	2500 - 5000

Toxic effects, evident as a reduction in the number of revertants (below the induction factor of 0.5), were observed at the following concentrations (μg/plate):

Strain	Experiment I		Experiment II
	without S9 mix	with S9 Mix	without S9 mix	with S9 Mix
TA 1535	5000	/	2500 - 5000	1000 - 5000
TA 1537	333 - 5000	1000 - 5000	333 - 5000	333 - 5000
TA 98	2500 - 5000	5000	1000 - 5000	2500 - 5000
TA 100	333 - 5000	333 - 5000	1000 - 5000	333 - 5000
WP2 uvrA	/	/	/	5000

/ = no toxic effects evident as a reduction in the number of revertants (below the induction factor of 0.5)

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative without metabolic activation
negative with metabolic activation

In conclusion, it can be stated that under the experimental conditions reported, the test item did not induce gene mutations by base pair changes or frameshifts in the genome of the bacterial strains used.