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EC number: 202-411-2 | CAS number: 95-33-0
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Endpoint summary
Administrative data
Key value for chemical safety assessment
Genetic toxicity in vitro
Description of key information
The test substance CBS was negative in bacterial gene mutation assays (Monsanto Co. 1976, MHWJ 1997); also a mouse lymphoma assay was negative (Monsanto Co. 1979). An in vitro chromosomal aberration assay test gave weak evidence for a clastogenic potential in presence of metabolic activation, but the positive findings was assessed to be ambiguous. The only in vivo test (on embryonic mortality) cannot be assessed adequately due to insufficient data reporting. Based on the above discussed findings, it is assessed that CBS is not genotoxic, which is in line with the EU risk assessment.
It is stated in the EU risk assessment (2007) that no relevant evidence for mutagenicity of CBS and its hydrolysis products MBT and CHA was found.
In addition, the genotoxic potential of DCBS (read-across substance of the category approach) was evaluated in an in vivo bone marrow chromosome aberration assay. Male and female Sprague-Dawley CD® rats were administered with 1000 mg/kg bw test substance once via gavage. Groups of 5 males and 5 females (treatment group and vehicle control each) were sacrificed at 6, 24 and 48 hours following test substance administration. The body weight and clinical observations were recorded during the study. The bone marrow cells were prepared and at least 50 mitotic cells per animal were analysed for cytogenetic aberrations. Clinical signs were observed for all 1000 mg/kg treated animals at 24 and 48 hours after application. The clinical signs observed were depression, red stains on nose/eyes, soft faeces, slight depression, urine stains, and rough coat. No animal died during the study. A significant change in body weight was seen for the 48 hours (1000 mg/kg bw) treatment group. No statistically significant differences between the mean mitotic indices of the test groups and the vehicle control were seen. A statistically significant increase in percent aberrant cells and the mean number of aberrations per cell was seen in the positive control group (cyclophosphamide). Results from the 6- and 24 hours sacrifice data show that no statistically significant increase in the frequency of chromosome aberrations compared to control values was seen in the groups treated with DCBS. No statistically significant increase in aberrant cells was noted for the 48 hour treatment group (1 % aberrant cells) compared to historical control data (0.27% aberrant cells), whereas comparison to the current negative control (0% aberrant cells) revealed a statistically relevant difference using the non-parametric analysis. Further statistical analysis was performed on the 48-hours results and comparisons were made with historical control data and data from the current solvent control. The author concluded that the statistical significant increase in aberrations seen at 48 hours is based upon comparison with control value of zero. Based on these findings, the authors concluded that the test substance DCBS is not clastogenic. Moreover, the average number of chromosomes in the examined metaphases was determined for each animal and all treatment groups were statistically compared to the control group. Rats have a normal diploid chromosome number of 42. No statistically significant differences between the mean chromosome numbers of the test group and the vehicle control were noted (Monsanto Co 1985).
Endpoint Conclusion: No adverse effect observed (negative).
Link to relevant study records
- Endpoint:
- in vitro gene mutation study in bacteria
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 471 (Bacterial Reverse Mutation Assay)
- Principles of method if other than guideline:
- N-cyclohexylbenzothiazole-2-sulphenamide was tested in Salmonella typhimurium TA 100, TA 1535, TA 98, TA 1537 and Escherichia coli WP2 uvrA, with or without an exogenous metabolic activation system.
- GLP compliance:
- yes
- Type of assay:
- bacterial reverse mutation assay
- Target gene:
- ames assay
- Species / strain / cell type:
- other: Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537, E. coli WP2 uvrA
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9 mix
- Test concentrations with justification for top dose:
- 6.25 - 5000 µg/plate
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- other: 2-(2-furyl)-3-(5-nitro-2-furyl)acylamide (TA 98; TA 100); sodium azide (TA 1535); 9-aminoacridine (TA 1537). with S9: 2-aminoanthracene (all strains)
- Key result
- Species / strain:
- other: Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537, E. coli WP2 uvrA
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- other: 100 µg/plate (TA1537 without S9); 500 µg/plate (TA 1537 with S9); 1500 µg/plate (TA 100 with S9)
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Conclusions:
- Interpretation of results: negative
- Executive summary:
The test substance CBS was evaluated in a GLP- and guideline study (OECD TG 471) bacterial gene mutation test. Here, the tester strains Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537 and Escherichia coli WP2 uvrA were used. Treatment by the pre-incubation method was done for doses up to 5000 µg/plate. Toxicity effects varied in a strain-specific manner; at high doses from 200 µg/plate upward precipitations were observed. The test substance CBS was negative in presence and absence of metabolic activation (MHWJ 1997).
- Endpoint:
- in vitro cytogenicity / chromosome aberration study in mammalian cells
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Reliability:
- 2 (reliable with restrictions)
- Rationale for reliability incl. deficiencies:
- other: GLP and guideline study, but with restriction concerning cytotoxicity determination and data documentation (study in jananese, abstract and tables in english available, no historical control data documented)
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 473 (In Vitro Mammalian Chromosome Aberration Test)
- Principles of method if other than guideline:
- Genotoxicity of N-cyclohexyl-2-benzothiazolesulfenamide was studied by chromosomal aberration in cultured hamster lung (CHL/IU) cells.
- GLP compliance:
- yes
- Type of assay:
- in vitro mammalian chromosome aberration test
- Target gene:
- chromosomal aberrations
- Species / strain / cell type:
- other: Chinese hamster lung cells (CHL/IU)
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9 mix
- Test concentrations with justification for top dose:
- 10 - 81 µg/ml
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- other: mitomycin C (-S9), CPA (+S9)
- Details on test system and experimental conditions:
- CHL/IU cells.
Continuous treatment (24 and 48 h) without S9
Short term treatment (6 h) with and without S9(18 h sampling time)
Solvent: DMSO
Positive controls:
without S9: Mitomycin C
with S9: Cyclophosphamide
S9 from rat liver induced with phenobarbital and 5,6-benzoflavone.
2 plates per test. - Statistics:
- Fisher's exact test using a Bonferroni's correction for multiple comparisions; Armitage's trend test.
- Key result
- Species / strain:
- other: Chinese hamster lung cells (CHL/IU)
- Metabolic activation:
- with and without
- Genotoxicity:
- ambiguous
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Conclusions:
- Interpretation of results: ambiguous without metabolic activation weak positive with metabolic activation.
- Executive summary:
The test substance CBS was weak positive in an in vitro chromosomal aberration using CHL cells (MHWJ 1997). In the absence of metabolic activation (S-9 mix) no induction of structural chromosomal aberrations was found for doses ranging from 10 to 41 µg/ml; no strong cytotoxicity was induced, but higher doses led to total cytotoxicity; various treatment and sampling times used (24/24 h; 48/48 h and 6/18 h). In the presence of S-9 mix weakly increased frequencies of structural chromosome aberration were found in two experiments after 6 hour treatment and 18 hours sampling. Again, no strong cytotoxicity was induced but higher doses led to total cytotoxicity. The main results were as followed: experiment I: 41 µg/ml, 81 µg/ml results in 3.5 % and 2.0% aberrant cells (excluded gaps); experiment II: 41 mg/ml, 81 µg/ml results in 1.0% and 3.5 % aberrant cells (excluded gaps). Since all solvent controls ranged from 0.5 to 1.0 aberrant cells (excluded gaps), this is interpreted as weak evidence for a clastogenic potential.
However, statistically evaluation revealed no dose dependency for structural chromosomal aberration. In addition, cytotoxcity was indicated in the first experiment as relative methaphase frequency to the solvent control and in the second experiment as cell confluency. No dose dependency in cytotoxicity was seen up to 0.081 mg/ml test substance, whereas higher concentrations could not be evaluated because of very high cytotoxcity. Thus, no dose-depencency in genotoxcity and cytotoxity was noted in the relevant concentration range. The increase in aberrant cells (3.5 %) is rather weak. In addition, the background frequency was low in both experiments (0.5% and 1%). compared to data from literature (XX). In addition, the slight increase in aberrant cells was stated to be significant different from historical solvent control data, but no historical control data are documented. Based on the rather weak increase in aberrant cells, the lack of a dose response relationship in genotoxcity and cytotoxcity and the low background frequency noted, the findings was assumed to be ambiguous.
- 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:
- other: comparable to guideline study with acceptable restriction (e.g. positive controls not consistent with recent guideline, no differentiation on colony-size)
- Principles of method if other than guideline:
- Santocure BO-78-221 (N-cyclohexyl-2-benzothiazolesulfenamide) was tested for mutagenicity in a MLA at the TK locus in L5178Y mouse lymphoma cells with or without S9 metabolic activation according to the protocol of Clive, D., Mutation Res., 31, 17-29, 1975).
- GLP compliance:
- no
- Type of assay:
- mammalian cell gene mutation assay
- Target gene:
- Tk locus
- Species / strain / cell type:
- other: L5178Y mouse lymphoma assay
- Metabolic activation:
- with and without
- Metabolic activation system:
- S9 mix
- Test concentrations with justification for top dose:
- 0.078 µg/ml to 40 µg/ml
- Untreated negative controls:
- yes
- Negative solvent / vehicle controls:
- yes
- True negative controls:
- yes
- Positive controls:
- yes
- Positive control substance:
- other: EMS (-S9), dimethylnitrosamine (+S9)
- Details on test system and experimental conditions:
- Mouse lymphoma assay
- Species / strain:
- mouse lymphoma L5178Y cells
- Metabolic activation:
- with and without
- Genotoxicity:
- negative
- Cytotoxicity / choice of top concentrations:
- cytotoxicity
- Vehicle controls validity:
- valid
- Untreated negative controls validity:
- valid
- Positive controls validity:
- valid
- Conclusions:
- Interpretation of results: negative
- Executive summary:
The test substance CBS was negative in a mouse lymphoma assay with L5178Y cells, with S-9 mix up to a dose of 75 µg /ml and without S9-mix up to a dose of 30 µg/ ml; toxic effects were found with and without S-9 mix in high doses (Monsanto Co 1979).
Referenceopen allclose all
No cytotoxicity was observed in TA 100 without S9 as well as in TA 98 and TA 1535, respectively.
Cytotoxicity: 100 µg/plate (TA1537 without S9); 500 µg/plate (TA 1537 with S9); 1500 µg/plate (TA 100 with S9). Precipitation of the test substance was noted at concentrations of 200 µg/plate and higher.
Cytotoxicity:
80 µg/ml (24 - 48 h treatment -S9)
83 µg/ml (6 h treatment -S9)
160 µg/ml (6 h Treatment +S9).
After 6 h treatment + S9 there were 4.5 % structural chromosomal aberrations
(incl. gaps) induced at 41 µg/ml and 1.88 % polyploidy at 81 µg/ml.
However a trend test showed no dose dependency for the structural chromosomal aberrations. In a confirmation test, 3.5 % structural chromosomal aberrations (incl. gaps) were induced at 81 µg/ml and 1.13 % polyploidy at 41 µg/ml.
However a trend test showed no dose dependency for the structural chromosomal aberrations or the polyploidy.
Experiment I and II with metabolic activation, 6 h treatment
Group | concentration µg/ml | TAG (%) | TA (%) | Polyploid (%) |
experiment I | ||||
Solvent (DMSO) | 0 | 1 (0.5) | 1 (0.5) | 0.38 |
CBS | 41 | 9* (4.5) | 7 (3.5) | 0.88 |
CBS | 81 | 5 (2.5) | 4 (2.0) | 1.88* |
experiment II | ||||
Solvent (DMSO) | 0 | 2 (1.0) | 2 (1.0) | 0.88 |
CBS | 20 | 4 (2.0) | 4 (2.0) | 0.38 |
CBS | 41 | 2 (1.0) | 2 (1.0) | 1.13* |
CBS | 81 | 7* (3.5) | 7 (3.5) | 0.79 |
TAG: total no. of cells with aberrations
TA: total no. of cells with aberrations except gap
*:Significant different from histroical solvent control (p<0.05) by fisher's exact test using a Bonferroni's correction for multiple comparisions.
The statistically significant increase in aberrant cells and polyploidy after treatment with CBS with metabolic activation showed, when trend tested no dose dependency. In addition, according to recommendation given in the current OECD guideline (TG 473), the in vitro chromosome aberration assay is not appropriate method for the detection of polyploidy.
Cytotoxicity:
-S9:
No survivors at 39 µg/ml, 5.7% to 13.3% relative growth for
the concentration range from 15 to 30 µg/ml.
+S9:
No survivors at 80 µg/ml, 75.2% relative growth at 40 µg/ml,
toxicity was moderate to high in 0.313 to 20 µg/ml concentration range.
The test substance CBS did not induce a significant increase in mutations at the TK locus in L5178Y mouse lymphoma cells with or without S9 metabolic activation. A range of concentrations up to the highly toxic levels of 30 µg/ml (without metabolic activation) and 75 µg/ml (with metabolic activation) were assayed. No differentiation on colony-size was made.
The test substance CBS is considered to be inactive in the mouse lymphoma forward mutation assay.
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (negative)
Genetic toxicity in vivo
Description of key information
The only in vivo test (on embryonic mortality) cannot be assessed adequately due to insufficient data reporting. However, the genotoxic potential of DCBS (read-across substance of the category approach) was evaluated in an in vivo bone marrow chromosome aberration assay. DCBS was negative in this assay.
Endpoint conclusion
- Endpoint conclusion:
- no study available
Additional information
In vitro data
The test substance CBS was evaluated in a GLP- and guideline study (OECD TK 471) bacterial gene mutation test. Here, the tester strains Salmonella typhimurium TA 98, TA 100, TA 1535, TA 1537 and Escherichia coli WP2 uvrA were used. Treatment by the pre-incubation method was done for doses up to 5000 µg/plate. Toxicity effects varied in a strain-specific manner; at high doses from 200 µg/plate upward precipitations were observed. The test substance CBS was negative in presence and absence of metabolic activation (MHWJ 1997). These findings were confirmed by another Ames assay, employing Salmonella typhimurium tester strain TA 98, TA 100, TA 1535, TA 1537, TA 1538 and Saccharomyces cerevisiae strain D4 with and without metabolic activation for doses ranging from 0.1 to 500 µg/ plate (Monsanto Co 1976).
The findings from the bacterial mutation assays were confirmed in a mammalian cell mutation assay. The test substance CBS was negative in a mouse lymphoma assay with L5178Y cells, with S-9 mix up to a dose of 75 µg /ml and without S9-mix up to a dose of 30 µg/ ml; toxic effects were found with and without S-9 mix in high doses (Monsanto Co 1979). No differentiation on colony-size was made.
However, the test substance CBS was weak positive in an in vitro chromosomal aberration using CHL cells. In the absence of metabolic activation (S-9 mix) no induction of structural chromosomal aberrations was found for doses ranging from 10 to 41 µg/ml; no strong cytotoxicity was induced, but higher doses led to total cytotoxicity; various treatment and sampling times used (24/24 h; 48/48 h and 6/18 h). In the presence of S-9 mix weakly increased frequencies of structural chromosome aberration were found in two experiments after 6 hour treatment and 18 hours sampling. Again, no strong cytotoxicity was induced but higher doses led to total cytotoxicity.
The main results were as followed: experiment I: 41 µg/ml, 81 µg/ml results in 3.5 % and 2.0% aberrant cells (excluded gaps); experiment II: 41 mg/ml, 81 µg/ml results in 1.0% and 3.5 % aberrant cells (excluded gaps).
Since all solvent controls ranged from 0.5 to 1.0 aberrant cells (excluded gaps), this is interpreted as weak evidence for a clastogenic potential. However, the author stated, that trend tests showed no dose dependency for structural chromosomal aberrations (MHWJ 1997).
In vivo data
Embryonic mortality of CBS was investigated in rats. However, no reliable data on dominant lethal mutations can be derived because test method and data presentation are inadequate (Aleksandrov 1982).
However, in addition the genotoxic potential of DCBS (read-across substance of the category approach) was evaluated in an in vivo bone marrow chromosome aberration assay. Male and female Sprague-Dawley CD® rats were administered with 1000 mg/kg bw test substance once via gavage. Groups of 5 males and 5 females (treatment group and vehicle control each) were sacrificed at 6, 24 and 48 hours following test substance administration. The body weight and clinical observations were recorded during the study. The bone marrow cells were prepared and at least 50 mitotic cells per animal were analysed for cytogenetic aberrations. Clinical signs were observed for all 1000 mg/kg treated animals at 24 and 48 hours after application. The clinical signs observed were depression, red stains on nose/eyes, soft faeces, slight depression, urine stains, and rough coat. No animal died during the study. A significant change in body weight was seen for the 48 hours (1000 mg/kg bw) treatment group. No statistically significant differences between the mean mitotic indices of the test groups and the vehicle control were seen. A statistically significant increase in percent aberrant cells and the mean number of aberrations per cell was seen in the positive control group (cyclophosphamide). Results from the 6- and 24 hours sacrifice data show that no statistically significant increase in the frequency of chromosome aberrations compared to control values was seen in the groups treated with DCBS. No statistically significant increase in aberrant cells was noted for the 48 hour treatment group (1 % aberrant cells) compared to historical control data (0.27% aberrant cells), whereas comparison to the current negative control (0% aberrant cells) revealed a statistically relevant difference using the non-parametric analysis. Further statistical analysis was performed on the 48-hours results and comparisons were made with historical control data and data from the current solvent control. The author concluded that the statistical significant increase in aberrations seen at 48 hours is based upon comparison with control value of zero. Based on these findings, the authors concluded that the test substance DCBS is not clastogenic. Moreover, the average number of chromosomes in the examined metaphases was determined for each animal and all treatment groups were statistically compared to the control group. Rats have a normal diploid chromosome number of 42. No statistically significant differences between the mean chromosome numbers of the test group and the vehicle control were noted (Monsanto Co 1985).
Short description of key information:
The test substance CBS was negative in bacterial gene mutation assays (Monsanto Co. 1976, MHWJ 1997); also a mouse lymphoma assay was negative (Monsanto Co. 1979). An in vitro chromosomal aberration assay test gave weak evidence for a clastogenic potential in presence of metabolic activation, but the positive findings was assessed to be ambiguous. The only in vivo test (on embryonic mortality) cannot be assessed adequately due to insufficient data reporting. Based on the above discussed findings, it is assessed that CBS is not genotoxic, which is in line with the EU risk assessment.
It is stated in the EU risk assessment (2007) that no relevant evidence for mutagenicity of CBS and its hydrolysis products MBT and CHA was found.
Endpoint Conclusion: No adverse effect observed (negative)
Justification for classification or non-classification
The test substance CBS was negative in bacterial gene mutation assays
(Monsanto Co. 1976, MHWJ 1997); also a mouse lymphoma assay was negative
(Monsanto Co. 1979). An in vitro chromosomal aberration assay test gave
weak evidence for a clastogenic potential in presence of metabolic
activation, but the positive findings was assessed to be ambiguous. The
only in vivo test (on embryonic mortality) cannot be assessed adequately
due to insufficient data reporting. Based on the above discussed
findings, it is assessed that CBS is not genotoxic, which is in line
with the EU risk assessment.
It is stated in the EU risk assessment (2007) that no relevant evidence
for mutagenicity of CBS and its hydrolysis products MBT and CHA was
found.
According to CLP classification criteria (Regulation (EC) No 1272/2008) a classification is not justified.
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
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