N-tert-Butyl-2-benzothiazolesulfenamide was studied for oral toxicity in rats in an OECD combined repeat dose and reproductive/developmental toxicity screening test at doses of 0, 40, 200 and 1000 mg/kg/day.
With regard to repeat dose toxicity, in the males, temporary salivation after each administration was observed in the 200 mg/kg or more groups as well as suppression of food consumption and body weight gain in the 1000 mg/kg group. At autopsy after 42 days treatment, the number of animals with eosinophilic bodies in the kidney was increased in all treated groups, and increases in the degree of this change and the absolute and relative kidney weights were noted in the 1000 mg/kg group. Although slight hypertrophy of hepatocytes in the central zone was observed in the 200 mg/kg or more groups, increase in the relative liver weight related to the administration was observed only in the 1000 mg/kg group. Decrease in fatty change of hepatocytes in the periportal zone was also observed in the livers of the 1000 mg/kg group. In the 200 mg/kg or more groups, slight increase in the total bilirubin concentration and dose-dependent increase in hemosiderin deposits in the spleen were observed. In addition, hemoglobin and hematocrit value were slightly decreased and hemolytic anemia was induced in the 1000 mg/kg group.
In the females, temporary salivation after each administration was observed in the 200 mg/kg or more groups. Food consumption was decreased in the 1000 mg/kg group prior to mating and along with slight suppression of body weight gain during pregnancy. At autopsy on postpartum day 4, slight vacuolar degeneration in proximal tubules of the kidney was observed in the 200 mg/kg or more groups, as well as slight increase in the relative kidney weight. Slight hypertrophy of hepatocytes in the central zone in the 200 mg/kg or more groups and relative liver weights in the 1000 mg/kg group were found. Deposits of brown pigment in the spleen in the 1000 mg/kg group tended to increase.
With regard to the reproductive and developmental toxicity, no adverse effects on copulation and ovulation were observed in any treated group. Fertility indices in the 40 and 1000 mg/kg groups were lower than the control level. The low index observed in the 40 mg/kg group was seemed to be accidental because all the females in the 200 mg/kg group were found to be fertile. No dose-related abnormalities were observed for parturition and lactation. There were no adverse effects on viability, the sex ratio and body weights of pups in any treated group. N-tert-Butyl-2-benzothiazolesulfenamide at the dose levels applied did not demonstrate teratogenicity.
In conclusion, the NOEL for toxicity of N-tert-butyl-2-benzothiazolesulfenamide was suggested to be less than 40 mg/kg/day in males and 40 mg/kg/day in females, and that for reproductive and developmental toxicity was 200 mg/kg/day in males and females.
N-tert-Butyl-2-benzothiazolesulfenamide was not mutagenic in Salmonella typhimurium TA100, TA1535, TA98, TA1537 and Escherichia coli WP2 uvrA, with or without an exogeneous metabolic activation system.
Genotoxicity of N-tert-butyl-2-benzothiazolesulfenamide was studied by the chromosomal aberration test in cultured Chinese hamster lung (CHL/IU) cells. With short-term treatment, N-tert-butyl-2-benzothiazolesulfenamide induced structural chromosomal aberrations at 0.20 and 0.40 mg/ml (mid and high concentrations) in the presence of an exogenous metabolic activation system. Polyploidy was induced with and without the metabolic activation system at 0.20 and 0.40 mg/ml (mid and high concentrations), and 0.10 and 0.20 mg/ml (mid and high concentrations), respectively. However, the frequency was very low and therefore not considered to be biologically significant.
| Purity | : | 96.4% |
| Test species/strain | : | Rat/Crj:CD (Sprague-Dawley) |
| Test method | : | OECD Combined Repeat Dose and Reproductive/Developmental Toxicity Screening Test |
| Route | : | Oral (Gavage) |
| Doses | : | 0 (vehicle), 40, 200, 1000 mg/kg/day |
| Number of animals/group | : | Males, 13, Females, 13 |
| Vehicle | : | 5% Gum arabic |
| Administration period | : | Males, 42 days, Females, from 14 days prior to mating to Day 3 of lactation |
| Terminal killing | : | Males, Day 43, Females, Day 4 of lactation |
| GLP | : | Yes |
Test results:
b) Females: Temporary salivation after each administration was observed in the 200 mg/kg or more groups. Food consumption was decreased in the 1000 mg/kg group prior to mating and along with slight suppression of body weight gain during pregnancy. At autopsy on postpartum day 4, slight vacuolar degeneration in the proximal tubules of the kidney was observed in the 200 mg/kg or more groups, as well as slight increase in the relative kidney weights. Slight hypertrophy of hepatocytes in the central zone in the 200 mg/kg or more groups and relative liver weights in the 1000 mg/kg group were found. Deposits of brown pigment in the spleens in the 1000 mg/kg group tended to increase. But in many cases were comparable to those in the control group. Thus, it was suggested that hemolysis resulting from the administration did not occur in females. No abnormalities related to the treatment were observed in other organs. N-tert-Butyl-2-benzothiazolesulfenamide at 40 mg/kg did not affect females.
The no effect level (NOEL) for toxicity of N-tert-butyl-2-benzothiazolesulfenamide was suggested to be less than 40 mg/kg/day in males and 40 mg/kg/day in females, and that for reproductive and developmental toxicity was 200 mg/kg/day in males and females.
| Purity | : | 96.0 % |
| Test species/strains | : | Salmonella typhimurium, TA100, TA1535, TA98, TA1537, Escherichia coli WP2 uvrA |
| Test method | : | Guidelines for Screening Mutagenicity Testing of Chemicals (Japan) and OECD Guideline No. 471 and 472 |
| Procedures | : | Pre-incubation method |
| Solvent | : | DMSO |
| Positive controls | : | -S9 mix, 2-(2-Furyl)-3-(5-nitro-2-furyl) acrylamide (TA100, TA98, WP2), Sodium azide (TA1535) and 9-Aminoacridine (TA1537) +S9 mix, 2-Aminoanthracene (five strains) |
| Doses | : | -S9 mix; 0, 1.56, 3.13, 6.25, 12.5, 25.0, 50.0, 100 μg/plate (TA1537) 0, 3.91 - 500 μg/plate (TA98) 0, 15.6 - 500 μg/plate (TA1535) 0, 39.1 - 2500 μg/plate (TA100) 0, 313 - 5000 μg/plate (WP2) +S9 mix; 0, 62.5 - 2000 μg/plate (TA1537) 0, 313 - 5000 μg/plate (TA100, TA1535, TA98, WP2) |
| S9 | : | Rat liver, induced with phenobarbital and 5,6-benzoflavone |
| Plates/test | : | 3 |
| Number of replicates | : | 2 |
| GLP | : | Yes |
Test results:
Genetic effects:
Salmonella typhimurium TA100, TA1535, TA98, TA1537
| + | ? | - | |
| Without metabolic activation: | [ ] | [ ] | [*] |
| With metabolic activation: | [ ] | [ ] | [*] |
Escherichia coli WP2 uvrA
| + | ? | - | |
| Without metabolic activation: | [ ] | [ ] | [*] |
| With metabolic activation: | [ ] | [ ] | [*] |
| Purity | : | 96.0% |
| Type of cell used | : | Chinese hamster lung (CHL/IU) cells |
| Test method | : | Guidelines for Screening Mutagenicity Testing of Chemicals (Japan) and OECD Guideline No. 473 |
| Solvent | : | 0.5% Carboxymethylcellulose sodium solution |
| Positive controls | : | -S9 mix, Mitomycin C +S9 mix, Cyclophosphamide |
| Doses | : | -S9 mix (continuous treatment):0, 0.015, 0.030, 0.060 mg/ml -S9 mix (short-term treatment):0, 0.050, 0.10, 0.20 mg/ml +S9 mix (short-term treatment):0, 0.10, 0.20, 0.40 mg/ml |
| S-9 | : | Rat liver, induced with phenobarbital and 5,6-benzoflavone |
| Plates/test | : | 2 |
| GLP | : | Yes |
Test results:
With short-term treatment with an exogenous metabolic activation system, structural chromosomal aberrations (5.0 and 6.0%, including gap) were induced at 0.20 and 0.40 mg/ml (mid and high concentrations), respectively. Polyploidy (1.13 - 1.63%) was induced with and without the metabolic activation system at 0.20 and 0.40 mg/ml (mid and high concentrations), and 0.10 and 0.20 mg/ml (mid and high concentrations), respectively. However, we concluded that this was not biologically significant since the frequency was low.
Lowest concentration producing cytogenetic effects in vitro:
With metabolic activation (short-term treatment): 0.20 mg/ml (clastogenicity)
| clastogenicity | polyploidy | |||||
| + | ? | - | + | ? | - | |
| Without metabolic activation: | [ ] | [ ] | [*] | [ ] | [ ] | [*] |
| With metabolic activation: | [*] | [ ] | [ ] | [ ] | [ ] | [*] |
| 1) | The tests were performed by the Hatano Research Institute, Food and Drug Safety Center, 729-5 Ochiai, Hadano-shi, Kanagawa, 257, Japan. Tel +81-463-82-4751 Fax +81-463-82-9627 |