β-bromostyrene

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Ames assay:

Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical. The study was performed as per the OECD guideline 471 using Salmonella typhimurium strains TA100, TA1535, TA98, TA1535 and E. coli WP2 uvrA with and without of S9 metabolic activation system. The study was performed at various dose levels and in triplicate plates/dose. Concurrent solvent and positive control chemicals were incorporated in the study. The plates were observed for dose dependent increase in the number of revertants/plate. The test chemical did not induce gene mutation in Salmonella typhimurium strains TA100, TA1535, TA98, TA1535 and E. coli WP2 uvrA in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

In vitro mammalian chromosome aberration study:

Chromosomal aberration study was performed to determine the mutagenic nature of the test chemical. The study was performed using CHL/IU cell line. The duration of treatment was short term from 6-18 hrs and continuous treatment for 24 and 14 hrs. The doses were dissolved in DMSO and the dose levels were Short term treatment (With S9: 0, 0.90, 1.81, 3.61, 7.23 or 14.5µg/mL, Without S9: 0, 7.23, 14.5, 28.9, 57.8 or 116µg/mL), for 24 hrs and 48 hrs treatment (Without S9: 0, 14.5, 28.9, 57.8 or 116µg/mL) and retest dose level was 0, 5.71, 8.56, 12.8, 19.3 or 28.9µg/mL with S9. Concurrent solvent and vehicle control chemicals were included in the study. The test chemical did not induce chromosomal aberrations in CHL/IU cell line in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

Link to relevant study records

Reference

Reference 1

Endpoint:

genetic toxicity in vitro

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from J check

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Principles of method if other than guideline:

Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical

GLP compliance:

not specified

Type of assay:

bacterial reverse mutation assay

Target gene:

Histidine

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98 and TA 100

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Species / strain / cell type:

E. coli WP2 uvr A

Details on mammalian cell type (if applicable):

Not applicable

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

All strains: 0, 1.22, 4.88, 19.5, 78.1, 313, 1250 or 5000 µg/plate

TA100:
Without S9: 0, 2.44, 4.88, 9.77, 19.5, 39.1, 78.1 µg/plate
With S9: 0, 9.77, 19.5, 39.1, 78.1, 156, 313 µg/plate

TA1535: With and without S9: 0, 2.44, 4.88, 9.77, 19.5, 39.1, 78.1 µg/plate

TA98: With and without S9: 0, 9.77, 19.5, 39.1, 78.1, 156, 313 µg/plate

TA1535: With and without S9: 0, 2.44, 4.88, 9.77, 19.5, 39.1, 78.1 µg/plate

E.coli WP2 uvrA: With and without S9: 0, 9.77, 19.5, 39.1, 78.1, 156, 313 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: The chemical was soluble in DMSO

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

sodium azide

other: 2- (2- Furyl)-3-(5-nitro-2-furyl)acrylamide (TA100, WP2uvrA, TA98, -S9), 2-Methoxy-6-chloro-9-[3-(2-chloroethyl)-aminopropylamino]acridine.2HCl (TA1537, -S9), 2-Aminoanthracene (TA1535, WP2uvrA, +S9), Benzo[a]pyrene (TA100, TA1537, TA98, +S9)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar

DURATION
- Preincubation period: No data
- Exposure duration: No data
- Expression time (cells in growth medium): No data
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): No data
SPINDLE INHIBITOR (cytogenetic assays): No data
STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: Triplicate plates were used/dose level and the study was performed in triplicates

NUMBER OF CELLS EVALUATED: No data

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data

OTHER EXAMINATIONS:
- Determination of polyploidy: No data
- Determination of endoreplication: No data
- Other: No data

OTHER: No data

Rationale for test conditions:

No data

Evaluation criteria:

The plates were observed for a dose dependent increase in the number of revertants/plate

Statistics:

No data

Species / strain:

S. typhimurium, other: TA100, TA1535, TA98, TA1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

No data

Remarks on result:

other: No mutagenic potential

Table 1:

Metabolic activation system	Dose (µg/plate)	TA100	TA1535	WP2uvrA	TA98	TA1537
Without S9	DMSO	98	11	18	27	10
	113	11	17	32	7
	4.88	119	1	15	26	7
	19.5	112	5	15	25	6
	78.1	109	7	15	23	8
	313	0	0	0	0	0
	1250	0	0	0	0	0
	5000	0	0	0	0	0
With S9	DMSO	166	16	21	53	12
	1.22	195	14	20	53	7
	4.88	914	17	20	49	8
	19.5	229	10	21	49	5
	78.1	302	14	27	61	9
	313	0	0	0	0	0
	1250	0	0	0	0	0
	5000	0	0	0	0	0
Positive control		AF-2	SAZ	AF-2	AF-2	ICR-191
		535	331	81	436	1550
Positive control		BAP	2AA	2AA	BAP	BAP
		835	299	1153	292	118

 

Table 2:

Metabolic activation system	Dose (µg/plate)	TA100	TA1535	WP2uvrA	TA98	TA1537
Without S9	DMSO	125±17.1	14±3.5	28±1.0	16±3.6	8±2.5
	2.44	119±15.5	7±4.0	NT	NT	7±1.0
	4.88	120±4.6	11±2.5	NT	NT	8±2.5
	9.77	125±2.3	10±5.1	26±6.8	18±2.9	5±1.5
	19.5	117±7.5	8±3.8	26±4.0	20±11.9	6±2.3
	39.1	121±2.1	12±5.1	25±6.0	17±4.0	5±0.6
	78.1	112±10.4	7±0.6	20±8.1	15±2.5	6±2.0
	156	NT	NT	0±0.0	0±0.0	NT
	313	NT	NT	0±0.0	0±0.0	NT

 

 

Table 3:

Metabolic activation system	Dose (µg/plate)	TA100	TA1535	WP2uvrA	TA98	TA1537
Without S9	DMSO	143±9.0	8±2.5	25±7.1	51±11.3	11±2.9
	2.44	NT	13±2.1	NT	NT	14±3.0
	4.88	NT	8±3.1	NT	NT	9±3.5
	9.77	151±5.3	15±2.6	28±13.1	57±8.2	9±3.2
	19.5	166±7.5	9±1.0	28±3.0	44±7.0	10±1.5
	39.1	188±12.9	18±4.2	30±4.5	47±5.7	17±4.0
	78.1	237±29.5	12±3.2	25±5.9	53±10.7	11±3.2
	156	127±10.5	NT	29±6.2	31±2.9	NT
	313	0 0.0	NT	0 0.0	0 0.0	NT

 

Table 4:

Metabolic activation system	Dose (µg/plate)	TA100	TA1535	WP2uvrA	TA98	TA1537
Without S9	DMSO	107±18.0	14±3.8	12±1.7	19±9.5	9±0.6
	2.44	121±5.1	15±1.5	NT	NT	6±1.5
	4.88	118±7.0	9±3.6	NT	NT	4±0.0
	9.77	121±9.9	10±2.3	14±5.2	19±1.7	5±2.5
	19.5	123±12.3	7±1.2	15±1.7	19±3.2	9±2.3
	39.1	112±13.5	8±3.6	21±5.5	19±3.5	5±1.0
	78.1	94±3.5	7±1.0	16±4.5	20±5.2	7±3.1
	156	NT	NT	12±4.6	0±0.0	NT
	313	NT	NT	0 0.0	0±0.0	NT

 

Table 5:

Metabolic activation system	Dose (µg/plate)	TA100	TA1535	WP2uvrA	TA98	TA1537
Without S9	DMSO	124±15.1	9±3.2	19±6.0	46±7.5	11±2.6
	2.44	NT	9±2.0	NT	NT	9±3.5
	4.88	NT	8±1.5	NT	NT	7±1.2
	9.77	142±11.4	9±1.2	15±1.0	39±3.1	8±2.9
	19.5	151±13.3	8±3.8	16±3.0	42±7.6	10±2.5
	39.1	186±12.7	16±5.0	16±4.7	47±13.1	10±2.1
	78.1	212±13.3	11±4.0	16±2.6	44±6.1	11±2.6
	156	115±22.1	NT	17±3.2	27±4.6	NT
	313	0±0.0	NT	0±0.0	0±0.0	NT

Conclusions:

The test chemical did not induce gene mutation in Salmonella typhimurium strains TA100, TA1535, TA98, TA1535 and E. coli WP2 uvrA in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

Executive summary:

Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical. The study was performed as per the OECD guideline 471 using Salmonella typhimurium strains TA100, TA1535, TA98, TA1535 and E. coli WP2 uvrA with and without of S9 metabolic activation system. The study was performed at various dose levels and in triplicate plates/dose. Concurrent solvent and positive control chemicals were incorporated in the study. The plates were observed for dose dependent increase in the number of revertnats/plate. The test chemical did not induce gene mutation in Salmonella typhimurium strains TA100, TA1535, TA98, TA1535 and E. coli WP2 uvrA in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

Reference 2

Endpoint:

in vitro gene mutation study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from J-check

Qualifier:

according to guideline

Guideline:

other: Refer below principle

Principles of method if other than guideline:

Chromosomal aberration study was performed to determine the mutagenic nature of the test chemical

GLP compliance:

not specified

Type of assay:

other: In vitro mammalian chromosome aberration test

Target gene:

No data

Species / strain / cell type:

mammalian cell line, other: CHL/IU

Details on mammalian cell type (if applicable):

MEDIA USED
- Type and identity of media including CO2 concentration if applicable: Minimal essential medium

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

with and without

Metabolic activation system:

S9 mix

Test concentrations with justification for top dose:

0, 14.5, 28.9, 57.8, 116, 231, 463, 925 or 1850 µg/mL

Short term treatment:
With S9: 0, 0.90, 1.81, 3.61, 7.23 or 14.5 µg/mL
Without S9: 0, 7.23, 14.5, 28.9, 57.8 or 116 µg/mL

24 hrs and 48 hrs treatment:
Without S9: 0, 14.5, 28.9, 57.8 or 116 µg/mL

Retest:
With S9: 0, 5.71, 8.56, 12.8, 19.3 or 28.9 µg/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: The chemical was soluble in DMSO

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

cyclophosphamide

mitomycin C

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Preincubation period: No data
- Exposure duration: Short term (6-18 hrs), 24 and 48 hrs
- Expression time (cells in growth medium): Short term (6-18 hrs), 24 and 48 hrs
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): No data
SPINDLE INHIBITOR (cytogenetic assays): No data
STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: No data

NUMBER OF CELLS EVALUATED: No data

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: Yes, cell growth inhibition caused by the test material was noted

OTHER EXAMINATIONS:
- Determination of polyploidy: Yes
- Determination of endoreplication: Yes
- Other: No data

OTHER: No data

Rationale for test conditions:

No data

Evaluation criteria:

The slides were observed for chromatid and chromosome gaps, chromatid and chromosome breaks, chromatid and chromosome exchange and fragmentation

Statistics:

No data

Species / strain:

mammalian cell line, other: CHU/IU

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Additional information on results:

No data

Remarks on result:

other: No mutagenic potential

Table: Cell growth ratio in the cell growth inhibition test in cultured Chinese hamster cells treated with β- bromostyrene (Short term treatment- +S9)

Cell growth inhibition test
Study type		Treatment and conc. (µg/mL)	Cell growth ratio		Observations
S9	Time (hr)		Plate	Mean (%)^b	Condition of cells	Color of medium	Preciptates/Crystals
							1	2
+	6-18	0	100^a	100	-	-	-	-
			87		-	-	-	-
		14.5	49	46	++	-	-	-
			37		++	-	-	-
		28.9	24	33	++	-	-	-
			37		++	-	-	-
		57.8	12	13	+++	-	-	-
			12		+++	-	-	-
		116	0	6	+++	-	-	-
			12		+++	-	-	-
		231	0	0	+++	-	-	-
			0		+++	-	-	+
		463	0	6	g	-	-	+
			12		g	-	+	+
		925	0	6	g	-	+	+
			12		g	-	+	+
		1850	12	13	g	-	+	+
			12		g	-	+	+
Concentration of 50% cell growth inhibition: below 14.5µg/mL

Table: Cell growth ratio in the cell growth inhibition test in cultured Chinese hamster cells treated with β- bromostyrene (Short term treatment- -S9)

Cell growth inhibition test
Study type		Treatment and conc. (µg/mL)	Cell growth ratio		Observations
S9	Time (hr)		Plate	Mean (%)^b	Condition of cells	Color of medium	Preciptates/Crystals
							1	2
-	6-18	0	100^a	100	-	-	-	-
			100		-	-	-	-
		14.5	71	71	+	-	-	-
			71		+	-	-	-
		28.9	57	57	++	-	-	-
			57		++	-	-	-
		57.8	71	57	++	-	-	-
			42		++	-	-	-
		116	14	14	++	-	-	-
			14		++	-	-	-
		231	14	7	+++	-	-	-
			0		+++	-	-	+
		463	0	0	g	-	+	+
			0		g	-	+	+
		925	0	0	g	-	+	+
			0		g	-	+	+
		1850	0	0	g	-	+	+
			0		g	-	+	+
Concentration of 50% cell growth inhibition: belo0w 67.3µg/mL

Table: Cell growth ratio in the cell growth inhibition test in cultured Chinese hamster cells treated with β- bromostyrene (Continuous treatment: 24 hrs)

Cell growth inhibition test
Study type		Treatment and conc. (µg/mL)	Cell growth ratio		Observations
S9	Time (hr)		Plate	Mean (%)^b	Condition of cells	Color of medium	Preciptates/Crystals
							1	2
-	24	0	100^a	100	-	-	-	-
			100		-	-	-	-
		14.5	59	59	+	-	-	-
			59		+	-	-	-
		28.9	59	59	+	-	-	-
			59		+	-	-	-
		57.8	59	59	+	-	-	-
			59		+	-	-	-
		116	0	0	+	-	-	-
			0		+	-	-	-
		231	0	0	TOX	-	-	-
			0		TOX	-	-	+
		463	0	0	g	-	+	+
			0		g	-	+	+
		925	0	0	g	-	+	+
			0		g	-	+	+
		1850	0	0	g	-	+	+
			0		g	-	+	+
Concentration of 50% cell growth inhibition: below 66.7µg/mL

Table: Cell growth ratio in the cell growth inhibition test in cultured Chinese hamster cells treated with β- bromostyrene (Continuous treatment: 48 hrs)

Cell growth inhibition test
Study type		Treatment and conc. (µg/mL)	Cell growth ratio		Observations
S9	Time (hr)		Plate	Mean (%)^b	Condition of cells	Color of medium	Preciptates/Crystals
							1	2
-	48	0	100^a	100	-	-	-	-
			99		-	-	-	-
		14.5	84	80	++	-	-	-
			76		++	-	-	-
		28.9	69	65	++	-	-	-
			61		++	-	-	-
		57.8	69	65	++	-	-	-
			61		++	-	-	-
		116	7	7	++	-	-	-
			7		++	-	-	-
		231	0	0	TOX	-	-	-
			0		TOX	-	-	+
		463	0	0	g	-	+	+
			0		g	-	+	+
		925	0	0	g	-	+	+
			0		g	-	+	+
		1850	0	0	g	-	+	+
			0		g	-	+	+
Concentration of 50% cell growth inhibition: below 72.9µg/mL

Table: Cell growth ratio in the chromosome aberration test in cultured Chinese hamster cells treated with β- bromostyrene (Short term treatment: +S9)

Cell growth inhibition test
Study type		Treatment and conc. (µg/mL)	Cell growth ratio		Observations
S9	Time (hr)		Plate	Mean (%)^b	Condition of cells	Color of medium	Preciptates/Crystals
							1	2
-	6-18	0	100^a	100	-	-	-	-
			100		-	-	-	-
		0.90	100	93	-	-	-	-
			85		-	-	-	-
		1.81	100	100	-	-	-	-
			100		-	-	-	-
		3.61	100	93	-	-	-	-
			85		-	-	-	-
		7.23	85	85	-	-	-	-
			85		-	-	-	-
		14.5	71	73	+	-	-	-
			85		+	-	-	-
		PC	100	100	-	-	-	-
			100		-	-	-	-

a: the plate in the negative control was regarded as 100% growth

b: the mean showed as a growth ration against the negative control value

c: Condition of cells was observed at the end of the treatment. Color of medium was observed immediately after the addition of test solution, precipitates and crystals were observed immediately after the addition of test solution¹and at the end of treatment²

d:         -: most of the cells were attached to the surface of the plates and their shape was normal

+: there was discontinuity amons a small number of surviving cells

++: there was discontinuity among approximately half of the surviving cells

+++: there was discontinuity among most of the surviving cells

e:          - : No changes of color

f:           - : Absence of precipitates /crystals

+: presence of precipitated

g: condition of cells could not be observed due to sever precipitates of the test article

Conclusions:

The test chemical did not induce chromosomal aberrations in CHL/IU cell line in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

Executive summary:

Chromosomal aberration study was performed to determine the mutagenic nature of the test chemical. The study was performed using CHL/IU cell line. The duration of treatment was short term from 6-18 hrs and continuous treatment for 24 and 14 hrs. The doses were dissolved in DMSO and the dose levels were Short term treatment (With S9: 0, 0.90, 1.81, 3.61, 7.23 or 14.5µg/mL, Without S9: 0, 7.23, 14.5, 28.9, 57.8 or 116µg/mL), for 24 hrs and 48 hrs treatment (Without S9: 0, 14.5, 28.9, 57.8 or 116µg/mL) and retest dose level was 0, 5.71, 8.56, 12.8, 19.3 or 28.9µg/mL with S9. Concurrent solvent and vehicle control chemicals were included in the study. The test chemical did not induce chromosomal aberrations in CHL/IU cell line in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Gene mutation in vitro:

Data available for the target chemical was reviewed to determine the mutagenic nature of the test chemical. The studies are as mentioned below:

Gene mutation toxicity study was performed to determine the mutagenic nature of the test chemical. The study was performed as per the OECD guideline 471 using Salmonella typhimurium strains TA100, TA1535, TA98, TA1535 and E. coli WP2 uvrA with and without of S9 metabolic activation system. The study was performed at various dose levels and in triplicate plates/dose. Concurrent solvent and positive control chemicals were incorporated in the study. The plates were observed for dose dependent increase in the number of revertnats/plate. The test chemical did not induce gene mutation in Salmonella typhimurium strains TA100, TA1535, TA98, TA1535 and E. coli WP2 uvrA in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

Chromosomal aberration study was performed to determine the mutagenic nature of the test chemical. The study was performed using CHL/IU cell line. The duration of treatment was short term from 6-18 hrs and continuous treatment for 24 and 14 hrs. The doses were dissolved in DMSO and the dose levels were Short term treatment (With S9: 0, 0.90, 1.81, 3.61, 7.23 or 14.5µg/mL, Without S9: 0, 7.23, 14.5, 28.9, 57.8 or 116µg/mL), for 24 hrs and 48 hrs treatment (Without S9: 0, 14.5, 28.9, 57.8 or 116µg/mL) and retest dose level was 0, 5.71, 8.56, 12.8, 19.3 or 28.9µg/mL with S9. Concurrent solvent and vehicle control chemicals were included in the study. The test chemical did not induce chromosomal aberrations in CHL/IU cell line in the presence and absence of S9 metabolic activation system and hence is not likely to classify as a gene mutant in vitro.

Based on the data available for the target chemical, it does not exhibit gene mutation in vitro. Hence the test chemical is not likely to classify as a gene mutant as per the criteria mentioned in CLP regulation.