6,6'-(Ethane-1,2-diylbis(azanediyl))bis(dibenzo[c,e][1,2]oxaphosphinine-6-oxide)“

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Based on the results of the Bacterial Reverse Mutation Test it is concluded that EDA-DOPO is not mutagenic in the Salmonella typhimurium test strains TA97a, TA98, TA100, TA102 and TA1535 in the absence and presence of metabolic activation under the experimental conditions in the pre-sent study.

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2016-09-23 - 2016-10-07

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Species / strain / cell type:

S. typhimurium TA 98

Species / strain / cell type:

S. typhimurium TA 100

Species / strain / cell type:

S. typhimurium TA 102

Species / strain / cell type:

S. typhimurium TA 1535

Species / strain / cell type:

S. typhimurium TA 97

Metabolic activation:

with and without

Metabolic activation system:

S9-mix

Test concentrations with justification for top dose:

DMSO was chosen as vehicle because the best result of a homogeneous and pipettable suspension was achieved with the solvent DMSO. ; tThis solvent does not have any ef-fects on the viability of the bacteria or the number of spontaneous revertants in the tested concentrations.
The following nominal concentrations were prepared for the first experiment:
5000 µg/plate, 1500 µg/plate, 500 µg/plate, 150 µg/plate and 50 µg/plate.
The following nominal concentrations were prepared for the second experiment:
5000 µg/plate, 2500 µg/plate, 1250 µg/plate, 625 µg/plate, 313 µg/plate and 156 µg/plate.

Vehicle / solvent:

DMSO

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

sodium azide

benzo(a)pyrene

other: 4-Nitro-1,2-phenylene diamine, 2-Amino-Anthracene

Evaluation criteria:

The colonies were counted visually and the numbers were recorded. A validated spread-sheet software (Microsoft Excel®) was used to calculate mean values and standard devia-tions of each treatment, solvent control and positive control.
The mean values and standard deviations of each threefold determination was calculated as well as the increase factor f(l) of revertant induction (mean revertants divided by mean spontaneous revertants) of the test item solutions and the positive controls. Additionally, the absolute number of revertants (Rev. Abs.) (mean revertants minus mean spontaneous revertants) was given.
A substance is considered to have mutagenic potential, if a reproducible increase of re-vertant colonies per plate exceeding an increase factor of 2 in at least one strain can be observed. A concentration-related increase over the range tested is also taken as a sign of mutagenic activity.

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 97

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 102

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

First Experiment

Strain		TA97a		TA98		TA100		TA102		TA1535
Induction		-S9	+S9	-S9	+S9	-S9	+S9	-S9	+S9	-S9	+S9
Demin. water	Mean	87	88	19	19	106	113	387	383	24	23
	sd	21.1	20.4	3.6	2.1	18.0	18.5	76.6	18.0	1.0	5.6
DMSO	Mean	73	105	12	17	105	110	356	357	26	24
	sd	1.5	39.0	1.0	4.0	15.3	9.2	6.9	40.3	3.5	6.4
Positive Controls*	Mean	351	388	296	61	647	1176	760	1088	365	145
	sd	175.5	90.6	69.3	9.8	16.2	259.6	28.8	350.9	96.0	23.7
	f(I)	4.81	3.70	24.67	3.59	6.10	10.69	2.13	3.05	15.21	6.04
5000 µg/plate	Mean	104	90	14	12	116	112	291	251	27	28
	sd	11.1	25.7	3.2	1.5	5.3	17.1	28.4	37.2	4.0	8.9
	f(I)	1.42	0.86	1.17	0.71	1.10	1.02	0.82	0.70	1.04	1.17
1500 µg/plate	Mean	86	95	16	13	108	108	231	273	25	26
	sd	23.1	23.0	1.5	2.6	15.5	16.7	34.9	102.2	5.2	6.5
	f(I)	1.18	0.90	1.33	0.76	1.03	0.98	0.65	0.76	0.96	1.08
500 µg/plate	Mean	111	104	16	15	110	113	336	331	24	37
	sd	7.0	30.8	2.6	2.1	16.2	11.1	38.2	67.2	7.4	7.5
	f(I)	1.52	0.99	1.33	0.88	1.05	1.03	0.94	0.93	0.92	1.54
150 µg/plate	Mean	104	108	12	12	105	115	279	235	23	27
	sd	22.1	18.0	1.0	1.0	14.8	6.4	48.4	63.8	5.5	3.6
	f(I)	1.42	1.03	1.00	0.71	1.00	1.05	0.78	0.66	0.88	1.13
50 µg/plate	Mean	91	87	13	13	92	119	335	259	26	22
	sd	18.0	18.1	2.6	1.0	15.9	3.1	6.1	22.0	4.0	4.5
	f(I)	1.25	0.83	1.08	0.76	0.88	1.08	0.94	0.73	1.00	0.92

Second Experiment

Strain		TA97a		TA98		TA100		TA102		TA1535
Induction		-S9	+S9	-S9	+S9	-S9	+S9	-S9	+S9	-S9	+S9
Demin. water	Mean	108	91	9	14	100	96	264	331	19	12
	sd	7.0	8.7	1.5	4.0	2.0	9.2	13.9	22.0	2.9	1.5
DMSO	Mean	91	89	13	10	89	98	257	309	13	14
	sd	14.4	4.9	3.0	1.2	6.0	12.5	6.1	37.8	1.2	2.1
Positive Controls*	Mean	665	448	372	79	444	480	769	1525	291	64
	sd	40.5	150.3	44.0	6.2	40.6	48.0	10.1	113.5	74.4	14.0
	f(I)	7.31	5.03	28.62	7.90	4.44	4.90	2.99	4.94	15.32	4.57
5000 µg/plate	Mean	79	99	16	15	66	78	375	393	14	17
	sd	6.4	15.3	1.0	4.4	8.7	10.4	30.6	93.8	3.2	4.2
	f(I)	0.87	1.11	1.23	1.50	0.74	0.80	1.46	1.27	1.08	1.21
2500 µg/plate	Mean	96	108	12	14	75	88	311	377	13	15
	sd	20.0	2.6	4.4	3.5	10.2	9.6	91.4	26.6	2.3	2.9
	f(I)	1.05	1.21	0.92	1.40	0.84	0.90	1.21	1.22	1.00	1.07
1250 µg/plate	Mean	90	101	11	13	79	84	377	372	12	15
	sd	23.6	16.5	2.0	1.5	4.4	4.2	36.3	50.0	2.1	3.6
	f(I)	0.99	1.13	0.85	1.30	0.89	0.86	1.47	1.20	0.92	1.07
625 µg/plate	Mean	95	91	11	14	70	81	393	445	9	16
	sd	10.6	22.2	1.5	4.6	0.6	15.3	76.4	64.7	0.6	3.5
	f(I)	1.04	1.02	0.85	1.40	0.79	0.83	1.53	1.44	0.69	1.14
312 µg/plate	Mean	81	109	13	15	86	88	299	341	15	15
	sd	9.1	12.1	2.5	3.5	21.2	7.0	39.5	40.1	2.1	3.5
	f(I)	0.89	1.22	1.00	1.50	0.97	0.90	1.16	1.10	1.15	1.07
156 µg/plate	Mean	90	122	15	18	71	88	309	305	11	16
	sd	3.6	6.0	2.0	1.7	5.5	9.6	20.5	14.0	1.2	2.6
	f(I)	0.99	1.37	1.15	1.80	0.80	0.90	1.20	0.99	0.85	1.14

Conclusions:

Based on the results of this study it is concluded that EDA-DOPO is not mutagenic in the Salmonella typhimurium test strains TA97a, TA98, TA100, TA102 and TA1535 in the ab-sence and presence of metabolic activation under the experimental conditions in the pre-sent study.

Executive summary:

Two valid experiments were performed.

 

The study procedures described in this report were based on the most recent OECD and EC guidelines.

The test item EDA-DOPO was tested in theSalmonella typhimuriumreverse mutation assay with five strains ofSalmonella typhimurium(TA97a, TA98, TA100, TA102 and TA1535). The test was performed in triplicates in the presence and absence of S9-mix (rat liver S9-mix induced by Aroclor 1254).

 

In the first experiment, EDA-DOPO (dissolved in DMSO) was tested up to concentrations of 5000 µg/plate in the absence and presence of S9-mix (0.74 % final concentration in the treatment)in the strains TA97a, TA98, TA100, TA102 and TA1535 using the plate incorporation method.

EDA-DOPOshowed no precipitates on the plates at any of the concentrations.

The bacterial background lawn was not reduced at any of the concentrations and no relevant decrease in the number of revertants was observed in all bacteria strains. The test item EDA-DOPO showed no signs of toxicity towards the bacteria strains in both the absence and presence of metabolic activation.

The results of this experiment showed that none of the tested concentrations showed a significant increase in the number of revertants in all tested strains, in the presence and the absence of metabolic activation.

 

On the base of the first experiment, EDA-DOPO was tested up to concentrations of 5000 µg/plate in the absence and presence of S9-mix (0.74% final concentration in the treatment)in all bacteria strain using the pre-incubation method.

EDA-DOPOshowed no precipitates on the plates at any of the concentrations.

The bacterial background lawn was not reduced at any of the concentrations and no relevant decrease in the number of revertants was observed in all bacteria strains. The test item EDA-DOPO showed no signs of toxicity towards the bacteria strains in both the absence and presence of metabolic activation.

The results of this experiments showed that the test item EDA-DOPO caused no increase in the number of revertants in all bacteria strains compared to the solvent control, in both the absence and presence of metabolic activation. The test item EDA-DOPO did not induce a dose-related increase in the number of revertants colonies in all strains, in the presence and absence of metabolic activation.

 

Based on the results of this study it is concluded that EDA-DOPO is not mutagenic in theSalmonella typhimuriumstrains TA97a, TA98, TA100, TA102 and TA1535 in the absence and presence of metabolic activation under the experimental conditions in this study.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the results of the bacterial reverse mutation assay EDA-DOPO has not to be classified for mutagenicity.