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Toxicological information

Genetic toxicity: in vitro

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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
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
GLP - Guideline study, tested with the source substance bis(2-ethylhexyl)nonanedioate (CAS 103-24-2). In accordance to the ECHA guidance document “Practical guide 6: How to report read-across and categories (March 2010)”, the reliability was changed from RL1 to RL2 to reflect the fact that this study was conducted on a read-across substance.

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2004
Report Date:
2004
Reference Type:
publication
Title:
Reverse Mutation Test of Bis(2-ethylhexyl) azelate in Bacteria
Author:
Miwa, Y.
Year:
2004
Bibliographic source:
Toxicity Testing Reports of Environmental Chemicals, 11, 287-320
Reference Type:
secondary source
Title:
Bis(2-ethylhexyl) azelate
Author:
OECD
Year:
2006
Bibliographic source:
SIDS Initial Assessment Report for SIAM 22, Paris, France, 18-21 April, 2006
Reference Type:
secondary source
Title:
Diesters Category of the Aliphatic Esters Chemicals (Test Plan and Robust Summaries for Substances in the HPV Test Plan)
Author:
US-EPA (American Chemistry Council's Aliphatic Esters Panel)
Year:
2010
Bibliographic source:
High Production Volume (HPV) Chemical Challenge Program (201-16837A and 201-16837B)

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Bis(2-ethylhexyl)nonanedioate
- Analytical purity: 77.2%
- Impurities (identity and concentrations):
Bis(2-ethylhexyl)glutarate 2.2%,
Bis(2-ethylhexyl)adipate 2.4%,
Bis(2-ethylhexyl)pimelate 2.8%,
Bis(2-ethylhexyl)suberate 3.8%,
Bis(2-ethylhexyl)sebacate 3.3%,
Bis(2-ethylhexyl) 1-,9-nonamethylenedicarboxylate 5.3%,
Bis(2-ethylhexyl)1-,10-decamethylenedicarboxylate 0.6%,
Bis(2-ethylhexyl)1-,11-undecamethylenedicarboxylate 0.3%
- Lot/batch No.: N-31101

Method

Target gene:
His operon (S. typhimurium)
Trp operon (E. coli)
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
cofactor supplemented post-mitochondrial fraction (S9 mix), prepared from the livers of rats treated with phenobarbital and 5,6-benzoflavone.
Test concentrations with justification for top dose:
Range finding test:
1.22, 4.88, 19.5, 78.1, 312.5, 625, 1250 and 5000 µg/plate with and without metabolic activation
Main test:
312.5, 625, 1250, 2500 and 5000 µg/plate with and without metabolic activation
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: ethanol
Controls
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: -S9: 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide (AF2; 0.01 µg/plate: TA100, WP2uvrA; 0.1 µg/plate: TA98); sodium azide (SA; 0.5 µg/plate: TA1535); 9-aminoacridine (9AA; 80 µg/plate: TA1537); +S9: 2-aminoanthracene (2AA; up to 10 µg/plate all strains)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 h

NUMBER OF REPLICATIONS: triplicates each in two independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth
Evaluation criteria:
If the number of colonies with revertants was as double as that of control group and dose response was observed, the interpretation of the result was positive.
Statistics:
Mean values and standard deviation were calculated.

Results and discussion

Test resultsopen allclose all
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 nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: There were colorless clear fine oil drop-like precipitations and oil membrane-like precipitations on the surface of agar in any test concentration.

RANGE-FINDING: No cytotoxicity was observed in a range finding study with doses up to 5000 µg/plate.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

 Table 1. Test results of experiment 1 (plate incorporation)

With or without S9-Mix

Test substance concentration

Mean number of revertant colonies per plate

(μg/plate)

(average of 3 plates ± Standard deviation)

 

Base-pair substitution type

cross-linking type

Frameshift type

 

TA 100

TA1535

WP2 uvrA

TA98

TA1537

0 (aqua dest.)

102 ± 8.1

9 ± 2.6

35 ± 2.6

20 ± 3.5

5 ± 0.6

0 (vehicle)

110 ± 14

13 ± 6.2

27 ± 2.1

29 ± 4.2

7 ± 2.3

312.5

100 ± 9.3P

8 ± 2.1P

25 ± 7.0P

23 ± 0.6P

13 ± 3.1P

625

106 ± 2.6P

11 ± 2.6P

31 ± 6.5P

23 ± 4.2P

11 ± 0.6P

1250

112 ± 12.1P

10 ± 1.2P

26 ± 5.5P

29 ± 2.1P

6 ± 0.6P

2500

94 ± 2.5P

9 ± 2.5P

29 ± 2.6P

27 ± 4.0P

10 ± 3.5P

5000

92 ± 11.5P

7 ± 1.0P

24 ± 1.5P

23 ± 5.0P

4 ± 0.6P

Positive controls,
-S9 mix

Name

AF2

SA

AF2

AF2

9AA

Concentrations (μg/plate)

0.01

0.5

0.01

0.1

80

Mean No. of colonies/plate (average of 3 ± SD)

548 ± 7.8

525 ± 7.1

132 ± 6.5

421 ± 11.2

334 ± 87.7

+

0 (medium)

106 ± 12.1

11 ± 0.6

24 ± 3.5

30 ± 3.0

13 ± 1.0

+

0 (vehicle)

105 ± 12

10 ± 4.0

33 ± 7.2

27 ± 3.5

16 ± 3.8

+

312.5

103 ± 8.1P

7 ± 4.4P

30 ± 6.1P

25 ± 4.6P

13 ± 1.7P

+

625

96 ± 8.1P

7 ± 2.6P

31 ± 4.6P

31 ± 8.0P

11 ± 1.5P

+

1250

97 ± 15.6P

9 ± 2.6P

31 ± 5.0P

31 ± 6.4P

14 ± 1.5P

+

2500

96 ± 9.7P

6 ± 3.2P

33 ± 2.6P

25 ± 7.5P

11 ± 2.0P

+

5000

90 ± 3.2P

7 ± 4.2P

32 ± 5.8P

32 ± 5.5P

12 ± 2.9P

Positive controls,
+S9 mix

Name

2AA

2AA

2AA

2AA

2AA

Concentrations (μg/plate)

1

2

10

0.5

2

Mean No. of colonies/plate (average of 3 ± SD)

1036 ± 46.9

327± 18.4

969 ± 142.3

437 ± 30.3

181 ± 29.5

AF2 = 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide

9AA = 9-aminoacridine

2AA = 2-aminoanthracene

SA = sodium azide

P = Precipitate

Table 2. Test results of experiment 2 (plate incorporation)

With or without S9-Mix

Test substance concentration

Mean number of revertant colonies per plate

(μg/plate)

(average of 3 plates ± Standard deviation)

 

Base-pair substitution type

cross-linking type

Frameshift type

TA 100

TA1535

WP2 uvrA

TA98

TA1537

0 (aqua dest.)

132 ± 15.5

8 ± 1.0

39 ± 2.5

23 ± 8.1

9 ± 5.5

0 (vehicle)

131 ± 14.5

9 ± 3.1

41 ± 6.1

31 ± 6.9

9 ± 3.0

312.5

129 ± 17.1P

7 ± 2.1P

38 ± 7.0P

27 ± 4.2P

10 ± 1.5P

625

124 ± 5.2P

7 ± 1.2P

33 ± 7.8P

30 ± 11P

14 ± 5.2P

1250

127 ± 10.4P

8 ± 1.5P

34 ± 4.0P

30 ± 2.1P

13 ± 1.0P

2500

128 ± 16.8P

8 ± 3.8P

38 ± 3.2P

25 ± 1.0P

13 ± 3.6P

5000

124 ± 4.0P

10 ± 2.5P

38 ± 0.0P

27 ± 4.0P

10 ± 2.5P

Positive controls,
- S9

Name

AF2

SA

AF2

AF2

9AA

Concentrations (μg/plate)

0.01

0.5

0.01

0.1

80

Mean No. of colonies/plate (average of 3 ± SD)

569 ± 27.5

589 ± 36.5

137 ± 17.6

446 ± 19.1

411 ± 100.1

+

0 (medium)

137 ± 16.1

7 ± 2.5

33 ± 3.2

31 ± 7.8

12 ± 3.0

+

0 (vehicle)

132 ± 5.2

7 ± 3.2

46 ± 7.2

31 ± 6.1

16 ± 2.1

+

312.5

125 ± 22.4P

7 ± 1.5P

39 ± 2.6P

23 ± 1.2P

12 ± 1.5P

+

625

123 ± 10.0P

8 ± 2.1P

41 ± 5.1P

31 ± 2.5P

14 ± 6.5P

+

1250

125 ± 14.0P

9 ± 3.2P

40 ± 5.7P

23 ± 4.5P

14 ± 4.2P

+

2500

116 ± 3.5P

8 ± 2.1P

39 ± 12.5P

31 ± 1.2P

17 ± 5.1P

+

5000

121 ± 7.6P

11 ± 3.8P

46 ± 4.0P

35 ± 2.5P

13 ± 2.1P

Positive controls,
+ S9

Name

2AA

2AA

2AA

2AA

2AA

Concentrations (μg/plate)

1

2

10

0.5

2

Mean No. of colonies/plate (average of 3 ± SD)

999 ± 117.8

289 ± 6.1

995 ± 77.9

420 ± 33.7

135 ± 13.1

AF2 = 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide

9AA = 9-aminoacridine

2AA = 2-aminoanthracene

SA = sodium azide

P = Precipitate

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative