Pentaerythritol tetrastearate

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

08 Nov - 17 Dec 1999

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

No test substance purity reported

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

Report date:

2000

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

adopted Jul 1997

Deviations:

yes

Remarks:

no purity of test substance

GLP compliance:

yes (incl. QA statement)

Remarks:

The Department of the Government of the United Kingdom

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

2,2-bis[[(1-oxodocosyl)oxy]methyl]propane-1,3-diyl didocosanoate

EC Number:

262-895-6

EC Name:

2,2-bis[[(1-oxodocosyl)oxy]methyl]propane-1,3-diyl didocosanoate

Cas Number:

61682-73-3

Molecular formula:

C93H180O8

IUPAC Name:

3-(docosanoyloxy)-2,2-bis[(docosanoyloxy)methyl]propyl docosanoate (non-preferred name)

Method

Target gene:

his operon, trp operon

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 phenobarbitone and β-naphthoflavone

Test concentrations with justification for top dose:

First experiment (served as range finding study):
5, 15, 50, 150, 500, 1500 and 5000 µg/plate without metabolic activation (TA 1535, TA 1537, TA 98 and TA 100)
50, 150, 500, 1500 and 5000 µg/plate without metabolic activation (WP2uvrA)
50, 150, 500, 1500 and 5000 µg/plate with metabolic activation (all strains)

Second experiment:
50, 150, 500, 1500 and 5000 µg/plate with and without metabolic activation (all strains)

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: none

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

no

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

N-ethyl-N-nitro-N-nitrosoguanidine

benzo(a)pyrene

other: 2-Aminoanthracene: +S9: 1 ,2 or 10 µg/plate for TA 1535, TA 1537, TA 100 and WP2uvrA

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 h

NUMBER OF REPLICATIONS: 3 replicates each in 2 independent experiments

DETERMINATION OF CYTOTOXICITY
- Method: growth of bacterial background lawn

Evaluation criteria:

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

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Key result

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

Vehicle controls validity:

not applicable

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: Precipitation of the test substance was observed at 500 µg/plate and higher for all tester strains with and without metabolic activation

RANGE-FINDING/SCREENING STUDIES: In order to select appropriate dose levels for use in the main study, a preliminary test was carried out to determine the toxicity of the test material. The dose range of the test material was 0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 pg/plate. The test material exhibited toxicity at and above 500 µg/plate to tester strain TA 100 (without S9-mix only). This response however, was not observed in either the range-finding or main studies and was therefore considered artefactual and of no toxicological significance.

HISTORICAL CONTROL DATA (with ranges, means and standard deviation and confidence interval (e.g. 95%)
- Positive historical control data: The positive control values were within the range of the historical control data (please refer to Table 1 under "any other information on material and methods incl. tables") except of all three values for TA 98 and a single value for TA100 and TA 1535, respectively, in Experiment I without metabolic activation, a single value for TA 98 in Experiment I with metabolic activation, and all three values for TA 98 in Experiment II with metabolic activation. These deviations were not considered to invalid the test since a distinct increase of the number of revertants was seen when compared with the control values.
- Negative historical control data: The negative control values were within the range of the historical control data (please refer to Table 1 under "any other information on material and methods incl. tables") except of a single value for TA 1535 in Experiment II with metabolic activation, which was not considered to influence the validity of the test.

Any other information on results incl. tables

Table 2: Summar of Results of Experiment I

Test substance concentration (µg/plate)	Number of revertants (mean number of colonies per plate)					Number of revertants (mean number of colonies per plate)
	TA 100	TA 1535	WP2uvrA	TA 98	TA 1537	TA 100	TA 1535	WP2uvrA	TA 98	TA 1537
Without metabolic activation						With metabolic activation
0	117	24	24	27	13	90	20	15	24	14
	73	15	18	18	8	82	14	11	24	20
	95	12	14	19	6	72	18	15	22	14
5	76	28	nt	28	9	nt
	73	11		15	11
	82	22		24	8
15	86	18	nt	20	12
	63	25		21	11
	79	9		26	10
50	89	16	14	19	10	72	15	15	20	24
	101	21	9	17	9	80	22	20	27	7
	85	15	13	18	18	90	20	23	18	22
150	80	15	10	17	11	82	18	17	23	20
	95	22	11	22	8	101	24	13	21	18
	91	22	13	17	15	102	25	17	16	15
500 P	99	16	14	28	16	95	18	16	21	22
	90	22	10	15	13	81	17	20	23	12
	94	13	17	18	12	88	25	21	24	10
1500 P	104	19	9	30	13	77	35	10	30	33
	115	25	15	21	16	66	13	12	35	12
	96	21	14	26	14	91	21	21	16	15
5000 P	106	17	19	36	7	114	14	16	33	20
	105	15	23	23	14	93	21	13	33	23
	115	14	28	30	11	89	12	13	26	12
Positive controls	ENNG	ENNG	ENNG	4NQO	9AA	2AA	2AA	2AA	BP	2AA
Concentration (µg/plate)	3	5	2	0.2	80	1	2	10	5	2
No. Colonies	307	158	411	98	584	507	401	788	242	333
	297	190	362	101	586	518	292	680	263	373
	274	182	618	93	769	824	308	716	167	284

ENNG: N-ethyl-N`-nitro-N-nitrosoguanidine

9AA: 9-Aminoacridine

4NQO: 4-Nitroquinoline-1-oxide

BP: Benzo(a)pyrene

2AA: 2-Aminoanthracene

P: Precipitation

Table 3: Summary of results of Experiment II

Test substance concentration (µg/plate)	Number of revertants (mean number of colonies per plate)					Number of revertants (mean number of colonies per plate)
	TA 100	TA 1535	WP2uvrA	TA 98	TA 1537	TA 100	TA 1535	WP2uvrA	TA 98	TA 1537
Without metabolic activation						With metabolic activation
0	136	14	23	28	11	108	14	24	28	12
	130	22	27	23	11	96	9	31	33	8
	133	18	27	33	8	109	12	30	39	7
50	142	14	27	26	5	112	18	19	21	13
	140	16	19	23	9	99	16	23	26	9
	130	18	18	20	10	123	12	26	27	23
150	131	13	22	25	6	79	18	26	14	9
	156	21	23	23	14	91	5	24	21	4
	118	20	28	23	6	81	6	24	15	13
500 P	157	20	22	17	9	93	14	16	13	13
	100	17	19	17	15	92	7	42	17	11
	103	19	16	25	7	89	20	28	10	10
1500 P	106	13	23	29	8	100	20	24	17	11
	91	15	27	16	11	78	21	29	15	16
	121	21	22	22	22	114	11	26	15	6
5000 P	150	17	35	25	15	104	17	40	27	11
	116	15	37	20	6	119	20	39	28	11
	107	14	37	20	6	90	9	35	18	15
Positive controls	ENNG	ENNG	ENNG	4NQO	9AA	2AA	2AA	2AA	BP	2AA
Concentration (µg/plate)	3	5	2	0.2	80	1	2	10	5	2
No. Colonies	734	215	878	140	980	832	188	260	156	321
	589	276	727	159	983	917	186	318	161	310
	676	248	891	163	926	909	188	336	158	269

ENNG: N-ethyl-N`-nitro-N-nitrosoguanidine

9AA: 9-Aminoacridine

4NQO: 4-Nitroquinoline-1-oxide

BP: Benzo(a)pyrene

2AA: 2-Aminoanthracene

P: Precipitation

Applicant's summary and conclusion

Conclusions:

CLP/EU GHS criteria not met, no classification required according to Regulation (EC) No 1272/2008.