Fatty acids, C6-18, 2,2-dimethyl-1,3-propanediyl esters

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 based on grouping of substances (category approach)

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

GLP - Guideline study. According 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

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

(adopted 2008)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Harlan Laboratories Ltd., ShaThe department of health of the government of the United Kingdom

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

Fatty acids, C8-18 and C18-unsatd., esters with neopentyl glycol

EC Number:

286-072-6

EC Name:

Fatty acids, C8-18 and C18-unsatd., esters with neopentyl glycol

Cas Number:

85186-86-3

IUPAC Name:

85186-86-3

Details on test material:

- Name of test material (as cited in study report): only trade name given
- Physical state: yellow liquid
- Analytical purity: 100 %
- Lot/batch No.: OE10817
- Expiration date of the lot/batch: 21 October 2016

Method

Target gene:

his operon (S. typhimurium strains); trp operon (E. coli strain)

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

Metabolic activation:

with and without

Metabolic activation system:

cofactor supplemented post-mitochondrial fraction (10 % liver S9 mix in standard co-factors), prepared from the livers of rats treated with Phenobarbitone/ß-Naphthoflavone

Test concentrations with justification for top dose:

- Experiment 1 (plate incorporation assay): 50, 150, 500, 1500, and 5000 µg/plate
- Experiment 2 (pre-incubation): 50, 150, 500, 1500, and 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone
- Justification for choice of solvent/vehicle: the test substance was fully miscible in acetone at 100 mg/mL in solubility pre-tests.

Controls

Controls 1

Untreated negative controls:

yes

Remarks:

concurrent untreated control

Negative solvent / vehicle controls:

yes

Remarks:

acetone

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: Without S9: N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG, 2, 3 or 5 µg/plate for WP2uvrA, TA100, or TA1535), 9-Aminoacridine (9AA, 80 µg/plate for TA1537), 4-Nitroquinoline-1-oxide (4NQO, 0.2 µg/plate for TA98)

Controls 2

Untreated negative controls:

yes

Remarks:

concurrent untreated control

Negative solvent / vehicle controls:

yes

Remarks:

acetone

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: with S9: 2-Aminoanthracene (2AA, 1 µg/plate for TA100, 2 µg/plate for TA1535 and TA1537, 10 µg/plate for WP2uvrA), Benz(a)pyrene (BP, 5 µg/plate for TA98)

Details on test system and experimental conditions:

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

DURATION
- Preincubation period: 20 min
- Exposure duration: 48 h

NUMBER OF REPLICATIONS: three replicates/concentration or control

DETERMINATION OF FREQUENCY OF REVERTANT COLONIES: Domino colony counter

DETERMINATION OF CYTOTOXICITY
- Method: bacterial background lawn

Evaluation criteria:

A result was considered as positive, when the following criteria were met:
1. A dose-related increase in mutant frequency over the dose range tested.
2. A reproducible increase at one or more concentrations.
3. Biological relevance against in-house historical control ranges.
4. Statistical analysis of data as determined by UKEMS.
5. Fold increase greater than two times the concurrent solvent control for any tester strain (especially if accompanied by an out-of-historical range response)

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:

other: 1. no cytotoxicity (plate incorparation assay); 2. cytotoxicity at 5000 µg/plate in TA100, TA1535 and TA1537 without S9 (pre-incubation assay)

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

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:

RANGE-FINDING/SCREENING STUDIES:
A preliminary test was carried out to determine the toxicity of the test item using TA100 and WP2uvrA. The concentrations tested were 0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 µg/plate. In addition, a vehicle control (acetone) and a sterility control were run in parallel.
The test item was non-toxic to the strains of bacteria used (TA100 and WP2uvrA).

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 (μg/plate)	Mean number of revertant colonies per plate (average of 3 plates ± standard deviation)
		Base-pair substitution type			Frameshift type
		TA 100	TA1535	WP2uvrA	TA98	TA1537
–	0	84 ± 2.1	14 ± 3.5	22 ± 2.3	30 ± 5.9	19 ± 4.6
–	50	72 ± 14.2	17 ± 3.5	24 ± 2.6	30 ± 6.4	20 ± 5.0
–	150	76 ± 2.3	13 ± 4.4	21 ± 8.1	31 ± 8.5	13 ± 3.5
–	500	82 ± 6.9	18 ± 3.8	23 ± 8.7	23 ± 2.6	13 ± 3.5
–	1500	82 ± 16.5	14 ± 5.9	15 ± 4.0	24 ± 8.3	10 ± 1.5
–	5000	84 ± 2.6	12 ± 1.2	21 ± 7.5	25 ± 6.4	15 ± 2.1
Positive controls, –S9	Name	ENNG	ENNG	ENNG	4NQO	9AA
	Concentrations (μg/plate)	3	5	2	0.2	80
	Mean No. of colonies/plate (average of 3 ± SD)	557 ± 12.1	96 ± 20.6	639 ± 25.1	107 ± 11.5	360 ± 52.9
+	0	88 ± 7.0	13 ± 3.1	33 ± 6.7	17 ± 2.5	14 ± 4.0
+	50	86 ± 3.6	12.± 2.5	29 ± 7.5	14 ± 1.0	14 ± 3.5
+	150	83 ± 7.0	13 ± 3.2	25 ± 10.1	19 ± 1.7	13 ± 3.2
+	500	97 ± 14.8	13 ± 3.5	24 ± 6.7	16 ± 3.5	7 ± 0.6
+	1500	97 ± 27.8	12 ± 2.3	28 ± 5.3	19 ± 7.6	17 ± 2.1
+	5000	87 ± 5.8	12 ± 2.0	29 ± 3.6	22 ± 2.5	13 ± 0.6
Positive controls, +S9	Name	2AA	2AA	2AA	BP	2AA
	Concentrations (μg/plate)	1	2	10	5	2
	Mean No. of colonies/plate (average of 3 ± SD)	554 ± 36.4	412 ± 20.8	206 ± 11.5	114 ± 1.5	268 ± 19.5

 

ENNG = N-ethyl-N-nitro-N-nitrosoguanidine

4NQO = 4-nitroquinoline-N-oxide

9AA = 9-aminoacridine

2AA = 2-Aminoanthracene

BP = Benzo(a)pyrene

P = Precipitate

C = contaminated

 

 

Table 2. Test results of experiment 2 (pre-incubation)

 

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (average of 3 plates ± standard deviation)
		Base-pair substitution type			Frameshift type
		TA 100	TA1535	WP2uvrA	TA98	TA1537
–	0	87 ± 2.1	16 ± 2.6	32 ± 4.6	20 ± 6.8	10 ± 1.7
–	50	84 ± 14.3	17 ± 4.5	24 ± 3.0	23.± 4.4	12 ± 7.1
–	150	78 ± 5.0	15 ± 2.6	22 ± 2.3	21 ± 4.5	10 ± 3.0
–	500	95 ± 12.1	19 ± 5.3	33 ± 7.0	20 ± 2.3	11 ± 2.6
–	1500	83 ± 5.0	17 ± 3.6	33 ± 4.5	22 ± 1.7	11 ± 2.6
–	5000	83 ± 6.0 t	9 ± 1.5 t	34 ± 1.2	14 ± 4.0	12 ± 4.5 t
Positive controls, –S9	Name	ENNG	ENNG	ENNG	4NQO	9AA
	Concentrations (μg/plate)	3	5	2	0.2	80
	Mean No. of colonies/plate (average of 3 ± SD)	532 ± 3.2	170 ± 4.0	769 ± 54.8	127 ± 20.6	1319 ± 210.8
+	0	104 ± 14.5	21 ± 6.1	33 ± 3.2	12 ± 1.2	11 ± 1.2
+	50	99 ± 4.9	19 ± 12.2	28 ± 7.6	12 ± 1.7	9 ± 1.5
+	150	92 ± 1.2	20 ± 8.1	33 ± 3.1	13 ± 3.1	10 ± 2.1
+	500	109 ± 20.4	16 ± 2.6	29 ± 3.8	13 ± 1.7	10 ± 1.0
+	1500	94 ± 12.0	15 ± 2.5	36 ± 6.8	11 ± 0.6	10 ± 2.9
+	5000	100 ± 2.6	11 ± 0.0	34 ± 4.0	10 ± 3.1	7 ± 4.9
Positive controls, +S9	Name	2AA	2AA	2AA	BP	2AA
	Concentrations (μg/plate)	1	2	10	5	2
	Mean No. of colonies/plate (average of 3 ± SD)	1354 ± 153.5	357 ± 6.1	147 ± 5.3	286 ± 1.0	295 ± 1.7

 

ENNG = N-ethyl-N-nitro-N-nitrosoguanidine

4NQO = 4-nitroquinoline-N-oxide

9AA = 9-aminoacridine

2AA = 2-Aminoanthracene

BP = Benzo(a)pyrene

P = Precipitate

C = contaminated

t= citotoxic effects

 

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative