10-undecenyl 2-cyano-3,3-diphenylpropenoate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

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

Adequacy of study:

key study

Study period:

11 August 2009 and 2 September 2009

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study conducted in accordance with generally accepted scientific principles, possibly with incomplete or methodological deficiencies, which do not affect the quality of relevant results.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2009

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Remarks:

Date of inspection 19th August 2008, Date of signature 4th March 2009

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

947753-66-4

Cas Number:

947753-66-4

IUPAC Name:

947753-66-4

Constituent 2

Reference substance name:

2-Propenoic acid, 2-cyano-3-(4-methoxyphenyl)-3-phenyl-, 2-ethylhexyl ester

IUPAC Name:

2-Propenoic acid, 2-cyano-3-(4-methoxyphenyl)-3-phenyl-, 2-ethylhexyl ester

Constituent 3

Reference substance name:

700-213-5

EC Number:

700-213-5

IUPAC Name:

700-213-5

Constituent 4

Reference substance name:

Ethyl hexyl methoxycrylene

IUPAC Name:

Ethyl hexyl methoxycrylene

Constituent 5

Reference substance name:

2-Propenoic acid, 2-cyano-3-(4-methoxyphenyl)-3-phenyl, 2-ethylhexyl ester

IUPAC Name:

2-Propenoic acid, 2-cyano-3-(4-methoxyphenyl)-3-phenyl, 2-ethylhexyl ester

Details on test material:

Details on surrogate material:
- Name of test material: Ethyl hexyl methoxycrylene
- Physical state: light amber coloured viscous liquid
- Analytical purity: not stated
- Lot/batch No.: LLR 711
- Date received: 16 April 2009
- Storage condition of test material: room temperature in the dark
- Other: information related to the identity, purity and stability of the test material was the responsibility of the Sponsor

Method

Target gene:

Salmonella typhimurium:
TA 1535, TA 100: hisG46
TA 1537: hisC3076
TA 98: hisD3052

Escherichia coli CM891 (WP2uvrA/pKM101):
contains ochre mutation; deficient in DNA repair system 9uvrA); contains the pKM101 plasmid

Species / strain

Species / strain 1

Species / strain / cell type:

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

Species / strain 2

Species / strain / cell type:

E. coli WP2 uvr A pKM 101

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9

Test concentrations with justification for top dose:

Preliminary toxicity test: 0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 micrograms/plate.
Experiments 1 and 2: 50, 150, 500, 1500 and 5000 micrograms/plate.

Vehicle / solvent:

The chosen vehicle solvent was dimethyl sulfoxide. The test material was miscible in this vehicle.

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

Plates without S9 mix

Migrated to IUCLID6: at 2 micrograms/plate, WP2uvrA-; 3 micrograms/plate, TA100; 5 micrograms/plate, TA1535

Controls 2

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

Plates without S9 mix

Migrated to IUCLID6: at 80 micrograms/plate, TA1537

Controls 3

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

Plates without S9 mix

Migrated to IUCLID6: at 0.2 micrograms/plate, TA98

Controls 4

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene at 1 microgram/plate, TA100; 2 microgram/plate, TA1535 and TA1537; and 10 microgram/plate for WP2uvrA-

Remarks:

Plates with S9 mix

Controls 5

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Remarks:

Migrated to IUCLID6: at 5 microgram/plate, TA98

Details on test system and experimental conditions:

METHOD OF APPLICATION:
A standard plate incorporation method was employed.

Preliminary Toxicity Test
A preliminary toxicity test was conducted over a series of 10 dose levels ranging from 0.15 to 5000 microgram/plate (and control). The test was performed by mixing 0.1 ml of 10-hour bacterial cultures of TA100 or WP2uvrA-, 2 ml of molten, trace histidine or tryptophan supplemented top agar, 0.1 ml of test material formulation and 0.5 ml of S9-mix or phosphate buffer and overlaying onto sterile plates of Vogel-Bonner Minimal agar (30 ml/plate). After approximately 48 hours incubation at 37 deg C, the plates were assessed for the numbers of revertant colonies and examined for effects on the growth of the bacterial background lawn. Manual counts had to be performed at 5000 microgram/plate due to excessive test material precipitation and the presence of an opaque film.

Test 1: (Without Pre-Incubation)
Tests were conducted in the presence or absence of S9. Five dose levels and control were tested up to and including 5,000 micro g/plate. All tests were performed in triplicate.
- The test was performed by mixing in test tubes 0.1 ml of 10-hour bacterial cultures, 2 ml of molten, trace histidine or tryptophan supplemented top agar, 0.1 ml of test material formulation, vehicle or positive control and either 0.5 ml of S9-mix or phosphate buffer. The contents of each tube were mixed overlaid onto sterile plates of Vogel-Bonner Minimal agar (one tube/plate). The plates were incubated at 37 deg C for 48 hours. The frequency of revertant colonies was assessed using a Domino colony counter.

Test 2: (With Pre-Incubation)
A second assay was performed with pre-incubation at 27 deg C for 20 minutes before the addition of the agar overlay. The same 5 concentrations were used in this second test as in the first.

NUMBER OF REPLICATIONS:
3 replicates/strain

DETERMINATION OF CYTOTOXICITY
Any toxic effects of the test substance would be detected by a substantial reduction in revertant colony counts or by the absence of a complete bacterial lawn.

Evaluation criteria:

The criteria for determining a positive response are a dose-related increase in revertant frequency over the dose range tested and/or a reproducible increase at one of more concentrations in at least one bacterial strain with or without metabolic activation. Statistical significance is not the only determining factor for a positive response and the biological relevance of the results should be considered first. A test material is considered non-mutagenic (negative) in the test system if the above criteria are not met.

Statistics:

As reported in: Kirkland DJ, (Ed)(1989) Statistical Evaluation of Mutagenicity Test Data UKEMS sub-committee on Guidelines for Mutagenicity Testing. Report Part III - Cambridge University Press.

Results and discussion

Test results

Test results 1

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:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Test results 2

Species / strain:

E. coli WP2 uvr A pKM 101

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

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

Table 1: Results Test 1 Without S9 Activation

	Revertant colony counts (mean and SD 3 replicates)
Addition (micrograms/plate)	TA98	TA100	TA1535	TA1537	WP2uvrA-
0	18 +/- 2.1	87 +/- 11.2	19 +/- 9.0	9 +/- 4.4	20 +/- 2.5
50	18 +/- 4.0	90 +/- 14.7	20 +/- 4.9	10 +/- 0.6	27 +/- 6.7
150	20 +/- 2.1	86 +/- 13.2	16 +/- 1.7	9 +/- 2.0	21 +/- 6.7
500	22 +/- 5.8	88 +/- 3.2	17 +/- 2.6	9 +/- 1.0	21 +/- 4.7
1500	18 +/- 6.2	70 +/- 6.6	14 +/- 5.5	11 +/- 2.9	17 +/- 3.1
5000	18 +/- 6.1	85 +/- 8.6	12 +/- 4.4	5 +/- 0.0	16 +/- 0.0
2 -Nitroquinoline-N-oxide (0.2)	113 +/- 12.6
ENNG (3)		523 +/- 9.5
ENNG (5)			342 +/- 86.2
9-Aminoacridine (80)				1029 +/- 273.5
ENNG (2)					607 +/- 20.8

Table 2: Results Test 1 With S9 Activation

	Revertant colony counts (mean and SD 3 replicates)
Addition (micrograms/plate)	TA98	TA100	TA1535	TA1537	WP2uvrA-
0	30 +/- 8.0	73 +/- 4.6	12 +/- 2.0	15 +/- 3.6	20 +/- 6.4
50	22 +/- 1.2	78 +/- 7.6	13 +/- 4.6	9 +/- 5.1	21 +/- 4.7
150	24 +/- 4.0	83 +/- 13.6	9 +/- 1.2	12 +/- 1.7	23 +/- 5.9
500	29 +/- 7.1	69 +/- 9.7	10 +/- 0.6	10 +/- 5.0	22 +/- 1.5
1500	27 +/- 3.8	62 +/- 7.9	10 +/- 3.1	11 +/- 1.2	23 +/- 3.5
5000	18 +/- 2.1	61 +/- 17.6	8 +/- 3.5	11 +/- 7.2	26 +/- 6.6
Benzo(a)pyrene (5)	131 +/- 41.6
2 -Aminoanthracene (1)		1578 +/- 52.0
2 -Amionoanthracene (2)			268 +/- 33.5
2 -Aminoanthracene (2)				353 +/- 58.9
2 -Aminoanthracene (10)					311 +/- 15.0

Table 3: Results Test 2 Without S9 Activation (With Pre-Incubation)

	Revertant colony counts (mean and SD 3 replicates)
Addition (micrograms/plate)	TA98	TA100	TA1535	TA1537	WP2uvrA-
0	23 +/- 2.5	110 +/- 3.2	16 +/- 4.0	9 +/- 1.5	19 +/- 1.0
50	22 +/- 3.5	107 +/- 7.8	14 +/- 2.6	11 +/- 1.5	19 +/- 3.1
150	16 +/- 2.0	105 +/- 13.9	14 +/- 3.5	12 +/- 2.3	19 +/- 1.2
500	20 +/- 4.6	108 +/- 10.7	17 +/- 4.6	12 +/- 2.0	18 +/- 3.1
1500	20 +/- 4.6	112 +/- 12.5	18 +/- 2.9	11 +/- 1.7	15 +/- 3.1
5000	19 +/- 5.0	106 +/- 3.8	13 +/- 1.5	9 +/- 0.6	20 +/- 1.0
2 -Nitroquinoline-N-oxide (0.2)	162 +/- 2.5
ENNG (3)		458 +/- 12.7
ENNG (5)			443 +/- 120.2
9-Aminoacridine (80)				694 +/- 281.5
ENNG (2)					757 +/- 84.7

Table 4: Results Test 2 With S9 Activation (With Pre-Incubation)

	Revertant colony counts (mean and SD 3 replicates)
Addition (micrograms/plate)	TA98	TA100	TA1535	TA1537	WP2uvrA-
0	22 +/- 1.5	103 +/- 6.7	13 +/- 1.5	10 +/- 3.1	23 +/- 6.7
50	18 +/- 4.0	93 +/- 5.5	14 +/- 3.2	11 +/- 2.9	21 +/- 3.1
150	21 +/- 3.1	104 +/- 5.5	10 +/- 2.0	10 +/- 3.5	24 +/- 2.1
500	24 +/- 3.2	109 +/- 6.8	12 +/- 0.6	9 +/- 1.0	21 +/- 1.2
1500	22 +/- 3.2	100 +/- 9.5	11 +/- 2.1	10 +/- 1.5	19 +/- 2.6
5000	21 +/- 3.5	107 +/- 6.4	9 +/- 0.6	8 +/- 1.0	21 +/- 2.3
Benzo(a)pyrene (5)	168 +/- 27.1
2 -Aminoanthracene (1)		1630 +/- 71.7
2 -Amionoanthracene (2)			216 +/- 29.5
2 -Aminoanthracene (2)				375 +/- 28.4
2 -Aminoanthracene (10)					255 +/- 19.0

Applicant's summary and conclusion

Conclusions:

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

The test material was considered to be non-mutagenic under the conditions of this test.

Executive summary:

The test material was considered to be non-mutagenic under the conditions of this test. The test was conducted using OECD Guideline 471.