1,1'-(methylenedi-p-phenylene)bismaleimide

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 February 1988 - 18 April 1988

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study report does not cite specific official test guidelines, however the report is thorough and the test conditions are well documented. It may be determined that the methodology is broadly consistent with modern test methods and the report does include a claim that the study was conducted according to the General Principles of Good Laboratory Practice; on this basis the study is considered reliable with restrictions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1988

Report date:

1988

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Test material information

Constituent 1

Reference substance name:

1,1'-(methylenedi-p-phenylene)bismaleimide

EC Number:

237-163-4

EC Name:

1,1'-(methylenedi-p-phenylene)bismaleimide

Cas Number:

13676-54-5

Molecular formula:

C21H14N2O4

IUPAC Name:

1-(4-{[4-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)phenyl]methyl}phenyl)-2,5-dihydro-1H-pyrrole-2,5-dione

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): MB-3000, N,N'-(4,4'-diphenylenemethylene) bismaleimide
- Physical state: Pale yellow powder
- Lot/batch No.: CX-1074
- Storage condition of test material: Room temperature

Method

Species / strain

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Metabolic activation system:

Liver microsomal enzyme fraction

Test concentrations with justification for top dose:

2.5, 5, 10 and 20 μg/ml

Vehicle / solvent:

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

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

cyclophosphamide

ethylmethanesulphonate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium; in agar (plate incorporation); preincubation; in suspension; as impregnation on paper disk

DURATION
- Exposure duration: 24 hours (without S-9) and 6 hours (with S-9) + 18 hours in treatment free media

SPINDLE INHIBITOR (cytogenetic assays): demecolcine (colcemid 0.1μg/mL) 2 hours before required harvest time
STAIN (for cytogenetic assays): 2% Gurrs Giemsa R66

NUMBER OF REPLICATIONS: 2 per treatment with S-9 and 2 without S-9

NUMBER OF CELLS EVALUATED: 100 consecutive well spread metaphases from each culture

DETERMINATION OF CYTOTOXICITY
- Method: count of cells % of control

COUNT OF CELLS with POLYPLOIDY: yes

Evaluation criteria:

KUEMS Guidelines for mutagenicity Testing (1983)

Results and discussion

Test results

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: strain/cell type: CHO-K1 BH4cell line

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

Cytotoxicity

Cell counts for the treatment without S-9 24 hour treatment

	Without S-9
	Culture A		Culture B		Mean %
Concentration of MB-3000 (μg/mL)	No of cells X10^5	% of control	No of cells X10^5	% of control
0	3.75	100	2.86	100	100
2.5	2.96	79	3.06	107	93
5.0	2.35	63	2.69	94	78.5
10.0	1.86	50	1.69	59	54.5
20.0	1.34	36	1.59	56	46.0
EMS 1 mg/mL	1.84	49	1.78	62	55.5

Cell counts for the treatment with S-9 6 hour treatment

	With S-9
	Culture A		Culture B		Mean %
Concentration of MB-3000 (μg/mL)	No of cells X10^5	% of control	No of cells X10^5	% of control
0	1.80	100	1.52	100	100
2.5	1.91	106	1.45	95	100.5
5.0	1.66	92	1.61	106	99
10.0	1.23	68	1.37	90	79
20.0	1.07	59	0.68	45	52
CP 25 ug/mL	1.69	94	1.63	107	100.5

Result of Chromosome aberration assay

Treatment without S-9

Treatment group	Treatment time (hr)	Con-centration	No of diploid cells scored	Cells with polyploidy			No of cells with observed structural aberrations
				N	%	Judge-ment	GAPS	Chromatid		Chromosome		Others		Total No Cells with aberration		Judge-ment
							g	ctb	cte	csb	cse	X	Z	-g	+g
Solvent control	24	0	100	10	(9.1)	-	1	0	0	0	0	0	0	2	3	-
			100	11	(9.9)		2	0	0	0	0	0	0	0	2
			200	21	(9.5)		3 (1.5)	0(0)	0(0)	2(1)	0(0)	0(0)	0(0)	2(1)	5(2.5)
MB-3000	24	2.5	100	12	(10.7)	-	1	2	0	2	1	0	0	5	6	+
			100	8	(7.4)		5	0	0	4	1	0	0	5	10
			200	20	(9.1)		6 (3)	2 (1)	0(0)	6(3)	2(1)	0(0)	0(0)	10(5)	16 (8)
		5.0	100	17	(14.5)	-	4	0	0	1	0	0	0	1	5	+
			100	6	(5.7)		4	0	0	5	0	2	1	7	10
			200	23	(10.3)		8 (4)	0(0)	0(0)	6(3)	0(0)	2(1)	1(0.5)	8(4)	15(7.5)
		10.0	100	11	(9.9)	-	7	0	1	9	0	0	1	10	15	+
			100	8	(7.4)		4	4	1	4	1	0	6	8	12
			200	19	(8.7)		11 (5.5)	4(2)	2(1)	13(6.5)	1(0.5)	0(0)	7(3.5)	18(9)	27(13.5)
		20.0	100	9	(8.3)	-	3	5	2	5	2	0	4	13	15	+
			100	9	(8.3)		6	2	1	5	0	1	5	7	13
			200	18	(8.3)		9 (4.5)	7(3.5)	3(1.5)	10(5)	2(1)	1(0.5)	9(10.5)	20	28(14)
EMS	24	1000	50	6	(10.7)	-	22	17	21	14	3	4	0	43	44	-
			50	3	(5.7)		14	12	15	19	3	3	0	33	39
			100	9	(8.3)		36 (36)	29(29)	36 (36)	33(33)	6(6)	7(7)	0(0)	76(76)	83(83)

 

Treatment with S-9

Treatment group	Treatment time (hr)	Con-centration	No of diploid cells scored	Cells with polyploidy			No of cells with observed structural aberrations
				N	%	Judge-ment	GAPS	Chromatid		Chromosome		Others		Total No Cells with aberration		Judge-ment
							g	ctb	cte	csb	cse	X	Z	-g	+g
Solvent control	6	0	100	14	(12.2)	-	6	1	1	3	0	0	0	5	11	-
			100	2	(2.0)		4	1	1	3	1	0	0	6	10
			200	16	(7.4)		10 (5)	2(1)	2(1)	6(3)	1(0.5)	0(0)	0(0)	11(5.5)	21(10.5)
MB-3000	6	2.5	100	10	(9.1)	-	1	0	0	1	1	0	0	2	3	+
			100	8	(7.4)		1	0	0	0	0	0	0	0	1
			200	18	(8.3)		2 (1)	0(0)	0(0)	1(0.5)	1(0.5)	0(0)	0(0)	2(1)	4 (2)
		5.0	100	6	(5.7)	-	2	0	2	1	0	0	2	3	5	+
			100	6	(5.7)		6	1	1	0	0	0	0	1	6
			200	12	(5.7)		8 (4)	1(0.5)	3(1.5)	1(0.5)	0(0)	0(0)	2(1)	4(2)	11(5.5)
		10.0	100	11	(9.9)	-	7	6	3	2	0	0	4	11	16	+
			100	8	(7.4)		3	2	4	5	1	0	2	9	11
			200	19	(8.7)		10 (5)	8(4)	7(3.5)	7(3.5)	1(0.5)	0(0)	6(3)	20(10)	27(13.5)
		20.0	100	4	(3.8)	-	7	8	2	4	0	0	8	11	17	+
			100	6	(5.7)		6	3	7	4	2	0	9	15	19
			200	10	(4.8)		13 (6.5)	11(3.5)	9(4.5)	8(4)	2(1)	0(0)	17(8.5)	26(13)	36(18)
CP	6	25	50	8	(13.8)	-	9	9	14	29	2	0	0	38	40	-
			50	8	(13.8)		8	11	7	33	1	0	0	42	43
			100	16	(13.8)		17 (17)	20(20)	21(21)	62(62)	3(3)	0(0)	0(0)	80(80)	83(83)

 

Applicant's summary and conclusion

Conclusions:

It was concluded that the test substance showed evidence of cytogenic and clastogenic activities in this in vitro mamalian cell system under the test conditions employed.

Executive summary:

A test was conducted (Safe Pharm Laboratories, 1988) in vitro in Chinese Hamster Ovary cells to assess the clastogenic potential of BMI in mammalian cells. The test was conducted using a 6-hour exposure period in the presence of metabolic activation (achieved by adding induced rat liver homogenate metabolising system) and a 24-hour exposure period in the absence of metabolic activation.

Significant dose-related increases in both toxicity and the frequency of aberrations were observed under both regimes of treatment with the test material. BMI was concluded to be clastogenic to CHO cells in vitro.