N,N''-methylenebis[N'-[3-(hydroxymethyl)-2,5-dioxoimidazolidin-4-yl]urea]

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Read-across from Klimisch 1 study performed on a close structural analogue which is manufactured in the same way (reaction of formaldehyde with a modified urea) and reacts with water in the same way (hydrolysis releasing formaldehyde) as the registered substance

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2002

Report date:

2002

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes

Type of assay:

mammalian cell gene mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

1-[1,3-bis(hydroxymethyl)-2,5-dioxoimidazolidin-4-yl]-1,3-bis(hydroxymethyl)urea

EC Number:

278-928-2

EC Name:

1-[1,3-bis(hydroxymethyl)-2,5-dioxoimidazolidin-4-yl]-1,3-bis(hydroxymethyl)urea

Cas Number:

78491-02-8

Molecular formula:

C8H14N4O7

IUPAC Name:

1-[1,3-bis(hydroxymethyl)-2,5-dioxoimidazolidin-4-yl]-1,3-bis(hydroxymethyl)urea

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Method

Target gene:

Thymidine kinase (tk) locus.

Species / strain

Species / strain / cell type:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 induced rat liver S9 mix

Test concentrations with justification for top dose:

5, 10, 20, 40, 60, 80, 100 microg/ml (selected for mutation assessment from a wider range including lower concentrations (and following a preliminary rangefinder test)

Vehicle / solvent:

Water

Controls

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

benzo(a)pyrene

Details on test system and experimental conditions:

At least 1E+07 cells treated for 3h, sampled and plated to measure survival and transferred to growth medium for a 2-day expression time. After the expression period, 1E+04 cells plated in selective medium for mutation assessment. Duplicate cultures treated, quadruplicate plating for mutant colony counting. Small and large mutant colonies distinguished during counting.

Evaluation criteria:

Validity criteria:
- control mutation frequency within normal range
- positive control mutation frequency > solvent control
- plating efficiencies satisfactory.

Statistics:

Log mutation frequency in solvent controls compared to that for each treatment concentration (Dunnett's test for multiple comparisons) plus linear trend test.

Results and discussion

Test results

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

% relative survival and relative total growth clearly reduced at concentrations >20 microg/ml

Vehicle controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

Mutation test results

Test concentration (microg/ml)	Without S9 mix			With S9 mix
	% Relative survival	Relative Total Growth	Mutation Frequency*	% Relative survival	Relative Total Growth	Mutation Frequency*
0 (control)	100	1.00	75.06	100	1.00	62.91
5	95.70	0.90	86.12	107.76	0.97	81.00
10	95.07	0.97	83.77	143.69	0.91	78.83
20	89.41	0.72	115.82	99.49	0.88	84.59
40	68.86	0.58	323.39	84.85	0.60	177.94
60	38.24	0.31	663.01	68.64	0.53	259.73
80	16.74	0.12	1272.72	37.28	0.29	696.65
100	4.92	0.02	1540.42	31.25	0.17	843.74
NQO, 0.1	113.93	0.77	187.60
BP, 3.0				65.05	0.50	528.76

* Per million viable cells

Mutation Frequency result italicised, bold means pairwise comparison and linear trend test significant.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
positive with and without S9 mix

Clear evidence of induced mutation was obtained in this study of a chemical analogue. The increases in both large and small mutant colony numbers suggests a possibility of both gene mutation and larger-scale (chromosome) damage. Based on the close similarity of chemical structure and properties, it is predicted that the registered substance would give a similar result in this test system.