Toluene-4-sulphonohydrazide

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:

2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was performed in accordance with the OECD guideline 473, and in compliance with GLP standard.

Cross-reference

Reason / purpose:

reference to same study

Data source

Reference

Reference Type:

publication

Title:

Unnamed

Year:

2006

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)

Deviations:

not specified

Test guideline 2

Qualifier:

according to

Guideline:

JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals

Deviations:

not specified

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Test material

Reference

Name:

Unnamed

Type:

Constituent

Type:

Constituent

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): 4,4'-Oxybis(benzenesulfonyl hydrazide)
- Analytical purity: no data
- Purity test date: 99.3 wt%
- Lot/batch No.: 403650

Method

Target gene:

NA

Species / strain

Species / strain / cell type:

other: Chinese Hamster Lung (CHL/IU) Cells

Details on mammalian cell type (if applicable):

NA

Additional strain / cell type characteristics:

not specified

Metabolic activation:

with and without

Metabolic activation system:

rat, malem, liver, 30% S9 mix induced with phenobarbital and 5,6-benzoflavone

Test concentrations with justification for top dose:

Absence metabolic system; 0, 557, 696, 870, 1,088, 1,360, and 1,700 µg/mL
Presence metabolic system; 0, 357, 446, 557, 696, 870, and 1,088 µg/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: no data

Controls

Untreated negative controls:

yes

Remarks:

solvent

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

mitomycin C

Details on test system and experimental conditions:

Culture establishment: CHL cells in logarithmic were to maintain low aberration frequencies. Subcultured at low density, before over growth occurs. The cells were incubated at 37 C in atmosphere of 5 % (v/v) CO2 and 100 % humidity.

Description of follow up repeat study: Series of seven and ten concentration levels with 1.25-fold intervals were tested in the absence and the presence of metabolic activation system.

Evaluation criteria:

The test was considered as positive in this assay if; 1) Statistically significant increase in the proportion of cells with structural aberrations (excluding gaps) occurs at one or more concentration levels 2) 1) exceed the normal range, 3) If the result of 1) is reproducible.

Statistics:

Fisher’s exact test and Chochran Armitage’s trend test (p-value: 0.05)

Results and discussion

Test results

Species / strain:

other: Chinese Hamster Lung (CHL/IU) Cells

Metabolic activation:

with and without

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

-S9: at 1360 µg/mL and above. +S9: at 696µg/mL and above

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

None

Any other information on results incl. tables

Table 1. Chromosome aberration test on CHL cells treated with 4,4’-oxybis(benzenesulfonylhydrazide) [short-term treatment: -S9 mix]

Compound	Concentration (mg/mL)	Time of exposure (hr)	Relative cell growth (%)	No. of cells analyzed	No. of cells with aberrations –gap (%)	No. of cells analyzed for polyploid	No. of polyploid cells (%)	Final Judgment
DMSO^a)	0	6	100.0	200	0 (0.0)#	200	1 (0.5)
Test substance	557	6	47.7	200	8 (4.0)*	200	1 (0.5)
	696	6	37.7	200	21 (10.5)*	200	4 (2.0)
	870	6	33.1	200	27 (13.5)*	200	0 (0.0)	Positive
	1088	6	18.0	200	6 (3.0)*	200	2 (1.0)
	1360^c)	6	9.1	Toxic
	1700^c)	6	3.0	Toxic
MMC^b)	0.1	6	57.2	200	88 (44.0)*	200	0 (0.0)	Positive

-gap: total number of cells with aberrations except gap

# p≤0.05: Significant difference by trend test (Cochran-Armitage trend test)

* p≤0.05: Significant difference from the negative control group (Fisher’s exact test)

a) Negative control (Dimethyl sulfoxide),                  b) Positive control (Mitomycin C)

c) Visible precipitation was shown at the end of exposure period

Table 2. Chromosome aberration test on CHL cells treated with 4,4’-oxybis(benzenesulfonylhydrazide) [short-term treatment: +S9 mix]

Compound	Concentration (mg/mL)	Time of exposure (hr)	Relative cell growth (%)	No. of cells analyzed	No. of cells with aberrations –gap (%)	No. of cells analyzed for polyploid	No. of polyploid cells (%)	Final Judgment
DMSO^a)	0	6	100.0	200	0 (0.0)#	200	2(1.0)
Test substance	357	6	58.7	200	3 (1.5)*	200	3(1.5)
	446	6	46.9	200	6 (3.0)*	200	2(1.0)
	557	6	18.2	200	29 (14.5)*	200	0(0.0)	Positive
	696	6	4.8	Toxic
	870	6	3.3	Toxic
	1088	6	2.4	Toxic
CP^b)	12.5	6	85.4	200	66 (33.0)*	200	1(0.5)	Positive

-gap: total number of cells with aberrations except gap

# p≤0.05: Significant difference by trend test (Cochran-Armitage trend test)

* p≤0.05: Significant difference from the negative control group (Fisher’s exact test)

a) Negative control (Dimethyl sulfoxide),                   b) Positive control (Cyclophosphamide)

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
positive

The genetic toxicity of OBSH was evaluated in an in vitro Mammalian chromosome aberration test on chinese hamster ling cells. The study was performed in accordance with the OECD guideline 473. OBSH exhibited clastogenic activity on cultured Chinese Hamster Lung cells.

Executive summary:

The genetic toxicity of OBSH was evaluated in an in vitro Mammalian chromosome aberration test on chinese hamster lung cells. The study was performed in accordance with the OECD guideline 473.

Significant dose-dependent increases in the proportion of cells with structural aberrations were observed in cultures both in the absence (557 to 870mg/mL) and presence (446 to 557mg/mL) metabolic activation system. Significant increases in the proportion of cells with numerical aberrations were not observed in cultures at all concentration levels.

In conclusion, OBSH exhibited clastogenic activity on cultured Chinese Hamster Lung cells both with and without S9.