Triisononanoic acid, triester with 2,2'-[oxybis(methylene)]bis[2-(hydroxymethyl)propane-1,3-diol] tris(2-ethylhexanoate)

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:

8 Dec 1998 - 19 January 1999

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

GLP-Guideline study, tested with the source substance Dipentaerythritol ester of nC5/iC9 acids. 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:

1999

Report date:

1999

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:

other: USA, EPA (TSCA) OPPTS harmonised guidelines

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of Health of the Government of the United Kingdom, UK

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

647028-25-9

Cas Number:

647028-25-9

IUPAC Name:

647028-25-9

Details on test material:

- Name of test material (as cited in study report): only trade name given
- Physical state: straw coloured liquid
- Analytical purity: 100%
- Storage condition of test material: room temperature in the dark

Method

Target gene:

"his operon" (for S. typhimurium strains) and "trp operon" (for E. coli strains)

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 (S9 mix), prepared from the livers of rats treated with Aroclor 1254.

Test concentrations with justification for top dose:

Preliminary toxicity study: 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 µg/plate with and without metabolic activation
First and second experiment: 50, 150, 500, 1500 and 5000 µg/plate with and without metabolic activation

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: acetone

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: -S9: N-ethyl-N`-nitro-N-nitrosoguanidine (3, 5 and 2 µg/plate, respectively) for TA100, TA1535 and WP2uvrA; 9-Aminoacridine (80 µg/plate) for TA1537; 4-Nitroquinolone-1-oxide (0.2 µg/plate) for TA98.

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: +S9: 2-Aminoanthracene (1 µg/plate) for TA100, (2 µg/plate) for TA1535 and TA1537, (10 µg/plate) for WP2uvrA; Benzo(a)pyrene (5 µg/plate) for TA98.

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: inspection of the bacterial background lawn

OTHER: a preliminary toxicity test was carried out in the strains TA100 and WP2uvrA.

Evaluation criteria:

The test material may be considered positive in this test if the following criteria are met: The test material should have induced reproducible and statistically (Dunnett´s method of linear regression) significant increase in the revertant count in at least one strain of bacteria.

Statistics:

Mean values and standard deviation were calculated. Dunnett’s method of linear regression was used.

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, but tested up to precipitating concentrations

Remarks:

oily precipitate was observed at 5000 µg/plate

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, but tested up to precipitating concentrations

Remarks:

oily precipitate was observed at 5000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: An oily precipitate was observed at 5000 µg/plate; however this did not prevent the scoring of revertant colonies.

RANGE-FINDING/SCREENING STUDIES: The dose range for the main test was determined in a preliminary toxicity assay in which the test material was tested at the following concentrations: 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 µg/plate. The test material was non-toxic to the strains of bacteria used (TA100 and WP2uvrA).

COMPARISON WITH HISTORICAL CONTROL DATA: All tester strain cultures exhibited a characteristic number of spontaneous revertants per plate in the vehicle and untreated controls. Furthermore, positive control values were at least two times the respective vehicle control value for each strain. The historical control ranges are presented in Table 1 under "Any other information on results incl. tables".

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1: Historical control ranges

History profile-Vehicle control values
-S9	TA 100	TA 98	TA 1535	TA 1538	TA 1537	WP2uvrA	TA 102
Extreme values	61-196	13-56	10-39	6-41	4-17	12-39	216-339
Mean	117 ± 24.7	28 ± 6.7	24 ± 5.2	21 ± 6.8	10 ± 2.0	20 ± 4.6	264 ± 29.8
History profile-Vehicle control values
+S9	TA 100	TA 98	TA 1535	TA 1538	TA 1537	WP2uvrA	TA 102
Extreme values	63-177	14-52	11-39	11-50	5-20	11-40	205-343
Mean± SD	120 ± 22.2	33 ± 7.1	18 ± 4.0	25 ± 6.2	11 ± 2.3	21 ± 4.8	283 ± 32.7
History profile-Positive control values
-S9	TA 100	TA 98	TA 1535	TA 1538	TA 1537	WP2uvrA	TA 102
Extreme values	277-1126	127-698	163-1005	190-799	161-1149	342-1209	540-1188
Mean± SD	613 ± 175.7	203 ± 57.2	466 ± 208.4	484 ± 146.0	812 ± 201.3	834 ± 197.2	785 ± 172.1
+S9	TA 100	TA 98	TA 1535	TA 1538	TA 1537	WP2uvrA	TA 102
Extreme values	412-1315	198-757	139-516	212-915	123-718	225-1089	511-1090
Mean± SD	949 ± 189.7	420 ± 112.5	291 ± 63.6	454 ± 131.0	266 ± 90.2	734 ± 210.5	705 ± 126.6

Table 2: 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 ± SD)
		Base-pair substitution type			Frameshift type
		TA100	TA1535	WP2uvrA	TA98	TA1537
–	0	81 ± 11.0	16 ± 1.5	25 ± 3.5	19 ± 2.5	7 ± 3.8
–	50	77 ± 2.1	14 ± 0.6	22 ± 0.6	24 ± 0.6	9 ± 3.6
–	150	80 ± 12.2	14 ± 1.0	22 ± 0.6	20 ± 6.7	10 ± 3.8
–	500	83 ± 10.8	13 ± 2.3	23 ± 2.9	21 ± 6.0	7 ± 2.6
–	1500	76 ± 1.5	18 ± 5.2	18 ± 2.6	22 ± 1.2	7 ± 3.2
–	5000	90 ± 11.8 P	18 ± 8.2 P	24 ± 4.0 P	28 ± 4.9 P	5 ± 0.6 P
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)	323 ± 11.7	283 ± 10.0	503 ± 24.2	166 ± 5.3	1027 ± 358.2
		TA100	TA1535	WP2uvrA	TA98	TA1537
+	0	105 ± 7.8	11 ± 1.2	28 ± 3.8	30 ± 3.1	20 ± 3.6
+	50	101 ± 7.0	12 ± 0.6	27 ± 3.8	29 ± 6.6	20 ± 3.2
+	150	102 ± 5.5	16 ± 4.0	32 ± 1.5	27 ± 4.0	23 ± 2.0
+	500	93 ± 16.6	14 ± 4.0	28 ± 6.6	30 ± 6.7	20 ± 1.2
+	1500	86 ± 3.6	15 ± 3.5	26 ± 4.0	31 ± 3.1	17 ± 6.5
+	5000	93 ± 7.8 P	12 ± 1.2 P	24 ± 1.5 P	33 ± 5.7 P	15 ± 2.6 P
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)	920 ± 153.0	198 ± 9.3	555 ± 27.8	551 ± 132.0	196 ± 6.5

Table 3: Test Results of Experiment 2 (plate incorporation)

With or without S9-Mix	Test substance concentration (μg/plate)	Mean number of revertant colonies per plate (average of 3 plates ± SD)
		Base-pair substitution type			Frameshift type
		TA100	TA1535	WP2uvrA	TA98	TA1537
–	0	96 ± 4.5	18 ± 4.2	26 ± 3.8	18 ± 5.3	11 ± 6.5
–	50	97 ± 18.6	17 ± 4.0	29 ± 6.5	16 ± 3.6	12 ± 0.6
–	150	100 ± 9.5	20 ± 9.0	28 ±0.0	16 ± 3.8	11 ± 1.7
–	500	101 ± 8.4	18 ± 6.2	33 ± 6.0	19 ± 4.4	12 ± 1.2
–	1500	87 ± 6.4	17 ± 7.1	29 ± 7.1	22 ± 0.0	12 ± 1.2
–	5000	93 ± 1.0 P	19 ± 3.1 P	33 ± 5.1 P	17 ± 3.2 P	15 ± 6.0 P
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)	880 ± 31.5	257 ± 34.4	1191 ± 59.3	166 ± 4.6	1084 ± 263.4
		TA100	TA1535	WP2uvrA	TA98	TA1537
+	0	85 ± 4.0	14 ± 3.5	35 ± 1.2	31 ± 5.0	21 ± 1.5
+	50	91 ± 7.5	19 ± 0.7	35 ± 10.1	30 ± 8.0	24 ± 1.5
+	150	84 ± 9.5	14 ± 1.0	34 ± 6.1	29 ± 10.7	22 ± 3.5
+	500	101 ± 14.3	10 ± 0.6	42 ± 0.6	28 ± 4.5	22 ± 2.1
+	1500	94 ± 8.7	14 ± 0.6	36 ± 5.3	31 ± 4.7	21 ± 1.0
+	5000	101 ± 6.7 P	14 ± 3.2 P	34 ± 7.0 P	30 ± 5.5 P	22 ± 1.2 P
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)	865 ± 43.2	246 ± 7.5	1051 ± 90.0	611 ± 51.5	182 ± 7.0

ENNG = N-Ethyl-N´-nitro-N-nitrosoguanidine

4NQO = 4-Nitroquinoline-1-oxide

9AA = 9-Aminoacridine

BP = Benzo(a)pyrene

2AA = 2-Aminoanthracence

P = precipitate

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative