SDBR

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The results of an Ames study and of a chromosome aberration study (both performed according to OECD/EC guidelines (with and without metabolic activation) and under GLP conditions), did not indicate mutagenic properties of SDBR. 

Link to relevant study records

Reference

Reference 1

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

4 October - 18 October 1999

Reliability:

1 (reliable without restriction)

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Version / remarks:

29 December 1992

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

Storage conditions: room temperature

Target gene:

Histindine and tryptophan

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 induced rat liver S9-homogenate

Test concentrations with justification for top dose:

First assay: 0, 62, 185, 556, 1667 and 5000 µg/ plate.
Second assay: 0, 156, 313, 625, 1250 and 2500 µg/ plate.

Vehicle / solvent:

Solvent: DMSO

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

sodium azide

benzo(a)pyrene

other: 2-aminoanthracene

Details on test system and experimental conditions:

The Salmonella typhimurium strains were provided by Dr. B.N. Ames (University of California Berkeley, USA).
Two mutagenicity assays were performed. The plate-incorporation method with the histidine-requiring S. typhimurium mutants TA 1535, TA 1537, TA 98 and TZ 100 and the tryptophan-requiring Escherichia coli mutant WP2 uvrA as indicator strain was applied.

To 2 mL molten top agar containing 0.6% agar, 0.5% NaCl and 0.05 mM L-histidine,HCl/0.05 mM biotin, maintained at 46ºC were added subsequently: 0.1 mL of a fully grown culture of the appropriate strain, 0.1 mL of the appropriate test substance solution or of the negative or positive control substance, and 0.5 mL S9-mix for metabolic activation or 0.5 mL sodium phosphate 100 mM (pH 7.4) for without metabolic activation. The ingredients were thoroughly mixed and the mix was immediately poured onto minimal glucose agar plates (1.5% agar in Vogel and Bonner medium E with 2% glucose). All determinations were made in triplicate. The plates were incubated at ca. 37ºC for three days. Subsequently, the his+ revertants were observed. Cytotoxicity was defined as a reduction in the number of revertant colonies and/or a clearing of the background lawn of bacterial growth.

Rationale for test conditions:

A preliminary test to assess the toxicity of the test substance to the bacteria was not performed, as no cytotoxicity was expected. Instead, the toxicity test was incorporated into the first mutagenicity assay.

Evaluation criteria:

A response is considered to be positive if the mean number of revertant colonies on the test plate was increased two-fold or more compared to that on the vehicle control.
A test substance is considered to be mutagenic if a concentration-related increase or if a positive response reproducible in two independent assays is observed.
A test substance is considered to be not mutagenic in the bacterial gene mutation test if it produces neither a dose-related increase in the mean number if revertant colonies nor a reproducible positive response at any of the test points.

Historical data on the bacterial reverse mutation test (including positive and negative controls) are included in the report.

Statistics:

Not performed.

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity, but tested up to precipitating concentrations

Vehicle controls validity:

valid

Positive controls validity:

valid

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

Detailed results are attached below.
After addition of the test substance to the plates a white precipitation was observed, especially at the highest concentrations.
SDBR was slightly toxic to the TA 1535 and TA 98 strains in the absence of S9 at the highest concentration tested (5000 µg/plate) as evidenced by a decrease in the mean number of revertant colonies.

Conclusions:

The results of an Ames study performed according to EC guidelines (with and without metabolic activation) did not indicate mutagenic properties of SDBR.

Reference 2

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2000

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 473 (In Vitro Mammalian Chromosomal Aberration Test)

Version / remarks:

21 July 1997

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.10 (Mutagenicity - In Vitro Mammalian Chromosome Aberration Test)

Version / remarks:

29 December 1992

Deviations:

no

GLP compliance:

yes

Type of assay:

in vitro mammalian chromosome aberration test

Specific details on test material used for the study:

Storage conditions: ambient temperature

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Remarks:

CHO K1 line

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254-induced rat liver homogenate (S9)

Test concentrations with justification for top dose:

A preliminary solubility study in DMSO was performed. Based on precipitation at higher concentrations, it was decided to use 1000 µg/mL (final concentration in the culture medium) as the highest concentration.
Concentration range in assay 1 (without and with metabolic activation): 0, 50, 75, 100, 250, 500 and 1000 µg/ml
Concentration range in assay 2 (without and with metabolic activation): 0, 50, 100, 300, 500 and 1000 µg/ml

Vehicle / solvent:

DMSO

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

cyclophosphamide

mitomycin C

Details on test system and experimental conditions:

Duplicate cultures were used.
In absence of S9-mix, the treatment time was 18 hours, in the presence of S9-mix the treatment time was 4 hours (after this time the culture medium was replaced with fresh medium). The fixation time was 18 hours after onset of the treatment for all cell cultures.
In the second assay, in the absence of S9-mix treatment/fixation times were 18/18 hours and 32/32 hours. In the presence of metabolic activation the treatment/fixation times were 4/18 hours and 4/32 hours.

For each selected treatment, 200 well-spread metaphases per concentration (100 metaphases per culture), each containing 20-22 centromeres, were analysed by microscopic evaluation for chromatid-type aberrations (gaps, breaks, fragments, interchanges), chromosome-type aberrations (gaps, breaks, minutes, rings, dicentrics) and other abnormalities, such as interstitial deletions, endoreduplication, polyploidy and multiple aberrations (>10 aberrations per cell, excluding gaps), according to the criteria recommended by Savage (1975). If heavily damaged or endoreduplicated cells were observed, these cells were recorded but the cells were not counted and included in the 200 analysed cells. The Vernier readings of all aberrant cells were recorded.

Rationale for test conditions:

The selection of the highest concentration scored was based on toxicity of the test substance to the cells.

Evaluation criteria:

The main criteria for a positive response are a clear dose-related increase in the percentage of cells with structural aberrations over the concurrent control frequencies (in duplicate cultures), or a reproducible single positive dose (in duplicate cultures).
A test substance is considered to be negative if it produces neither a dose-related increase in the number of structural chromosomal aberrations nor a reproducible positive response at any of the test points.
Gaps are recorded separately and not included in the final assessment of clastogenic activity.

Statistics:

Data were analysed statistically by the Fisher's exact probability test (two-sided) for significance of differences between treated and control cultures.

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

In both (independent) chromosome aberration assays, in both the absence and the presence of S9, the substance did not induce reproducible, biologically relevant and statistically significant increase in the number of cells with structural chromosome aberrations at any time point analysed, when compared to the negative control values.
The positive control substances gave the expected statistically significant increases in the number of cells with structural chromosome aberrations.
Detailed results are attached below.

Conclusions:

The results of a reliable chromosome aberration study, performed according to OECD/EC guidelines and GLP principles, do not indicate that SDBR has mutagenic properties.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

The available data on SDBR do not indicate that the SDBR is mutagenic and therefore the substance is not classified for according to Regulation (EC) No 1272/2008.