sodium 4,4-dimethyl-2,5-dioxoimidazolidin-1-ide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

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:

1983-11-01 to 1982-04-08

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study performed according to GLP. Solvent controls for the non-activated portion were lost due to contamination and therefore results were evaluated in comparison with historical solvent control data.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Materials and methods

Test guideline

Test guideline 1

Qualifier:

no guideline available

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Version / remarks:

Predates but essentially complies with

Deviations:

yes

Remarks:

No confirmation of negative result

Test guideline 2

Qualifier:

no guideline available

Guideline:

EU Method B.17 (Mutagenicity - In Vitro Mammalian Cell Gene Mutation Test)

Version / remarks:

predates but essentially complies with

Deviations:

yes

Remarks:

No confirmation of negative result

GLP compliance:

yes

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

5,5-dimethylhydantoin

EC Number:

201-051-3

EC Name:

5,5-dimethylhydantoin

Cas Number:

77-71-4

IUPAC Name:

5,5-dimethylimidazolidine-2,4-dione

Constituent 2

Reference substance name:

Dimethylhydantoin

IUPAC Name:

Dimethylhydantoin

Test material form:

other: crystalline solid

Details on test material:

- Name of test material : Dimethylhydantoin
- Structural formula attached as image file (if other than submission substance): see Data Matrix attachment in Section 13 for DMH structural formula
- Substance type: white crystalline solid
- Physical state: solid
- Analytical purity: no data

Method

Target gene:

Thymidine Kinase (TK) gene

Species / strain

Species / strain / cell type:

mouse lymphoma L5178Y cells

Details on mammalian cell type (if applicable):

Not data

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

S-9 mix

Test concentrations with justification for top dose:

Preliminary toxicity test: 0.01, 0.1, 1.0, 10, 100, 1000 and 5000 µg/mL
Mutagenesis assay: 563, 751, 1001, 1335, 1780, 2373, 3164, 4219, 5625, 7500 and 10000 µg/mL

Vehicle / solvent:

- Solvent used: DMSO and Acetone were inculded as solvents.

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

identity of solvent not stated

True negative controls:

no

Positive controls:

yes

Positive control substance:

ethylmethanesulphonate

Remarks:

in the absence of activation

Migrated to IUCLID6: 1.0 and 0.5 µg/mL

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

identity of solvent not stated

True negative controls:

no

Positive controls:

yes

Positive control substance:

7,12-dimethylbenzanthracene

Remarks:

in the presence of activation

Migrated to IUCLID6: 7.5 and 5.0 µg/mL

Controls 3

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

identity of solvent not stated

True negative controls:

no

Positive controls:

not specified

Positive control substance:

not specified

Details on test system and experimental conditions:

METHOD OF APPLICATION: preincubation

DURATION
- Exposure duration: 4 hours
- Expression time (cells in growth medium): two days (cell population was performed at 24 and 48 hours.) The adjustment for expression time was performed by taking daily cell counts and replacing a volume of cells with fresh medium which yielded a cell population density of 0.3 x 10^6 cells per mL.
- Selection time (if incubation with a selection agent): 10-12 days

SELECTION AGENT (mutation assays): TFT at a final concentration of 3 µg/mL

NUMBER OF REPLICATIONS: 3

NUMBER OF CELLS EVALUATED: 100000 cells per mL

Evaluation criteria:

Positive: if there is a positive dose response and one or more of the three highest dose exhibit a mutant frequency which is two-fold greater than the background level.

Equivocal: if there is no dose response but any one or more doses exhibit a two-fold increase in mutant frequency over background.

Negative: if there is no dose response and none of the test cultures exhibit mutant frequencies which are two-fold greater than background.

Statistics:

Please refer to section Any other information on materials and methods incl. tables under the heading Formulas and Calcualtions.

Results and discussion

Test results

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES:
A preliminary toxicity test with and without S-9 activation was conducted. The solubility of the test article was determined in water. The test article was solubilised and diluted for testing at 5000, 1000, 100, 10, 1.0, 0.1 and 0.01 µg/ml. Test article toxicity was determined by comparing the cell comparison growth at each dose level with that of the solvent controls. Cell population density was determined 24 and 48 hours after the initial exposure to the test article.

ADDITIONAL INFORMATIONS ON MUTAGENICITY ASSAY - COMPARISON WITH HISTORICAL SOLVENT CONTROL DATA:
Solvent controls for the non-activated portion of testing on test article were lost due to contamination and therefore results were evaluated in comparison with historical solvent control data.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Table 1: Cloning Data in the Absence of Activation

 

Concentration	No. of Colonies/TFT plate			Average #/ plate	No. of Colonies/V.C. plate			Average #/ plate	Mutant frequency*	Induced mutant frequency
	1	2	3		1	2	3
10000 µg/mL	46	53	59	53	221	182	+	202	0.5	0.0**
7500 µg/mL	50	73	75	66	208	195	221	208	0.6	0.1
5625 µg/mL	65	64	69	66	190	206	195	197	0.7	0.2
4219 µg/mL	69	55	40	55	178	196	209	194	0.6	0.1
3164 µg/mL	38	44	52	45	203	212	207	207	0.4	-0.1
2373 µg/mL	50	65	55	57	203	178	197	193	0.6	0.1
1780 µg/mL	67	67	57	64	233	248	237	239	0.5	0.0
1335 µg/mL	48	40	51	46	182	194	206	194	0.5	0.0
1001 µg/mL	74	+	+	74	197	187	212	199	0.7	0.2
563 µg/mL	50	42	53	48	208	165	232	202	0.5	0.0
Solvent 1	+	+	+		+	+	+
Solvent 2	+	+	+		214	206	212	211
Ethyl Methanesulfonate
1.0 µL/mL	36	45	31	37	4	4	5	4	17.2
0.5 µL/mL	239	268	237	248	37	38	29	35	14.3
Solvent 1	+	+	+		142	141	163	14.9
Solvent 2	+	+	+		155	161	176	164
* Per 10^4Surviving cells + Culture lost ** Based on historical control values

 

Table 2: Cloning Data in the Presence of Activation

 

Concentration	No. of Colonies/TFT plate			Average #/ plate	No. of Colonies/V.C. plate			Average #/ plate	Mutant frequency*	Induced mutant frequency
	1	2	3		1	2	3
10000 µg/mL	50	55	75	60	210	227	233	223	0.5	-0.3
7500 µg/mL	+	+	+		231	211	224	222	+
5625 µg/mL	81	85	84	83	197	189	+	193	0.9	0.1
4219 µg/mL	88	93	78	86	235	220	218	224	0.8	0.0
3164 µg/mL	64	61	77	67	194	201	197	197	0.7	-0.1
2373 µg/mL	77	69	60	69	234	223	223	227	0.6	-0.2
1780 µg/mL	55	57	42	51	247	251	231	243	0.4	-0.4
1335 µg/mL	67	96	63	75	234	219	231	228	0.7	-0.1
1001 µg/mL	68	66	79	71	227	212	+	220	0.6	-0.2
751 µg/mL	78	51	51	60	214	218	188	207	0.6	-0.2
Solvent 1	64	88	68		+	+	+
Solvent 2	72	85	92	83	209	190	+	200	0.8
7, 12 Dimethylbenz (a) anthracene
7.5 µL/mL	87	105	93	95	40	36	31	36	5.3	4.5
5.0 µL/mL	218	209	214	214	131	11.3	147	130	3.3	2.5
Solvent 1	101	76	68	82	190	201	+	196	0.8
Solvent 2	81	72	68	74	196	201	211	203	0.7 (0.8)
* Per 10^4Surviving cells + Culture lost

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

The test substance DMH was not genotoxic in this assay, both in the presence and absence of metabolic activation.

Executive summary:

This study has been performed on DMH (Dimethyl Hydantoin) and has been used for read-across purposes.

The test substance DMH was not genotoxic in this assay, both in the presence and absence of metabolic activation.