Di(benzothiazol-2-yl) disulphide

Endpoint summary

Description of key information

There are only limited repeated dose toxicity data from MBTS available from an early cancer study in mice; which indicates that MBTS was not carcinogenic under the experimental conditions used.
A read across approach with data from MBT was conducted. There are several repeated dose toxicity studies available for MBT. In a two-generation toxicity study (CMA 1990) a LOAEL of 2500 ppm (ca. 150 mg to 250 mg/kg bw) was observed. In a two year carcinogenicity study with Fischer 344 rats (NTP 1988) a LOAEL of 188 mg/kg bw and day for females and of 375 mg/kg bw and day for males was determined. In a subchronic toxicity study (NTP 1988) a LOAEL of 187 mg/kg bw was noted. In mice a LOAEL of 375 mg/kg bw was suggested in a two-year carcinogenicity study (NTP 1988) and a NOAEL of 188 mg/kg bw in a subchronic toxicity study (NTP 1988). Thus, the NOAEL and LOAEL values from several MBT repeated dose studies are in the same range. The consistent findings in subchronic, chronic and reproduction/ developmental toxicity studies revealed a LOAEL in the range of 150 to 375 mg/kg bw and day. Following the recommendation given in the MAK publication (1999) an overall NOAEL of 50 mg/kg bw and day is suggested. This assumption based on data from the two-generation toxicity study and is supported by the consistent findings in the subchronic and chronic toxicity studies. The overall NOAEL value of 50 mg/kg is used as base for systemic DNEL calculation.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Endpoint conclusion

Dose descriptor:

NOAEL

50 mg/kg bw/day

Species:

rat

Additional information

There are only limited repeated dose toxicity data available for MBTS.

The carcinogenic potential of MBTS was evaluated in an early cancer study in two mice strains (strains B6C3F1 and B6AKF1) (Innes 1969). Even through the study was not performed according to present standards, it should be mentioned that MBTS was not found to be tumorigenic when given orally to mice for 18 months at the maximal tolerated dose, which was fixed at 464 mg/kg MBTS by gavage (dissolved in 0.5 % gelatine) from day 7 to 28 of age and at 1577 ppm in the diet (ca. 237 mg/kg bw/day) from day 28 for additional 17 months.

There are only limited data from an early cancer study available. There are no valid subchronic or chronic repeated dose studies available for MBTS. A read across approach was conducted with repeated dose toxicity data from MBT (benzothiazole-2-thiol) (see discussion endpoint summary toxicokinetics).

Read-across approach with MBT

In a carcinogenicity study (NTP 1988) male and female Fischer 344 rats were administered with MBT for 103 weeks. Groups of 50 male rats were administered 0, 375, or 750 mg/kg 2-mercaptobenzothiazole in corn oil by gavage, 5 days per week for 103 weeks. Groups of 50 female rats were administered 0, 188, or 375 mg/kg 2-mercaptobenzothiazole in corn oil by gavage on the same schedule. All animals were observed two times per day, and clinical signs were recorded once per week. Body weights by cage were recorded once per week for the first 12 week and once per month thereafter. A necropsy was performed on all animals including those found dead. During necropsy, all organs and tissues were examined for grossly visible lesions. Necropsy and histological exams were performed on all animals and all relevant tissues. No haematology and clinical biochemistry were performed. Survival of the low dose group of male rats was significantly lower (p < 0.001) than that of the vehicle controls after week 85 and of the high dose group after week 83 (mortality: low dose 28/50, high dose: 29/50; control: 8/50). Mortality of treated females were not significant different from control. Mean body weights of dosed male rats were similar to or greater than those of the vehicle controls. Mean body weights of dosed female rats were generally greater (up to 11%) than those of the vehicle controls. Rats were lethargic and showed prostration after they were dosed. The principal non-neoplastic lesions seen in these studies were nephropathy and inflammation and ulceration of the forestomach in rats. Nephropathy, characterized by tubular degeneration and regeneration, was present in all male rats and in more than 75% of the female rats; a severity grade from minimal to severe (1- 4) was recorded for each animal. The mean severity of nephropathy was increased in dosed male rats (vehicle control: mild-moderate; low dose and high dose: moderate-severe). However, these changes were later discussed as questionable (TSCA 1991-1993 cited in BG Chemie 2000). Ulcers and inflammation were observed at increased incidences in dosed rats, and epithelial hyperplasia and hyperkeratosis were observed at increased incidences in dosed male and low dose female rats. These effects were later discussed as treatment related effect (bolus-effect) but not as substance-related effect. The occurrence of neoplastic lesions were noted in male and female rats and will be discussed in the chapter carcinogenicity. Overall, no clear carcinogenic activity can be concluded from this study.

Based on the findings of this study a LOAEL for males of 375 mg/kg bw and day (effects: decreased survival rate, lethargic and prostration after dosing) and for females of 188 mg/kg bw and day (effects: lethargic and prostration after dosing) was suggested for repeated dose toxicity.

In an additional NTP carcinogenicity study male and female B6C3F1 mice were administered with MBT for 103 weeks (NTP 1988). Groups of 50 male and 50 female mice were administered 0, 375, or 750 mg/kg 2-mercaptobenzothiazole in corn oil by gavage, 5 days per week for 103 weeks. The study design and schedule were similar to the Fischer 344 rat carcinogenicity study discussed. Survival of the high dose group of female mice was significantly lower than that of the vehicle controls after week 27. The survival rate of the female low dose group and the male treatment groups were comparable to control. Six high dose male and four high dose female mice died on the same day during week 13. Since they were mistakenly dosed twice within a 16-hour period, these mice were censored from the statistical incidence of survival after week 12. Mean body weights of high dose male mice were 6%-14% lower than those of the vehicle controls from week 3 to week 64; the mean body weights of low dose males were 4 to 8 % lower compared to control from week 6 to week 64. In females the mean body weights of the high dose group were within 6 % of the vehicle control, whereas the mean body weights from the low dose group were generally greater than those of the vehicle controls throughout the studies. Similar to the rat study, mice were lethargic and showed prostration after they were dosed; however, no histological alterations of the nerve system were revealed. No substance-related non-neoplastic lesions were indicated in any of the treated mice. The Pathological and histopathological incidences of non-neoplastic lesions were in the range of the vehicle control. Neoplastic lesions noted were within the historical control data range. Based on the findings of this study, the lethargy after dosing, a LOAEL for treated mice of 375 mg/kg bw and day is suggested.

Subchronic studies were conducted with Fischer 344 rats and B6C3F1 mice to evaluate the cumulative toxic effects of repeated administration of MBT and to determine the doses to be used in the 2-year studies discussed above (NTP 1988). In the subchronic toxicity study with Fischer 344 rats groups of 10 animals of each sex were administered 0, 188, 375, 750, or 1500 mg/kg MBT in corn oil by gavage, 5 days per week for 13 weeks. The 13-week study in rats reported in this Technical Report was a second study. In the first study in rats, 3000 mg/kg groups all died during week 1. In the subchronic toxicity study with B6C3F1 mice groups of 10 mice of each sex were administered 0, 94, 188, 375, 750 or 1500 mg/kg MBT. Animals were checked two times per day for clinical signs and mortality. Individual animal weights were recorded weekly. At the end of the 13-week studies, survivors were killed. A necropsy was performed on all animals except those excessively autolysis or cannibalized. Histological exams performed on some animals from all groups. No haematology and clinical biochemistry were done.

No substance-related deaths occurred in Fischer 344 rats treated up to 1500 mg/kg bw and day MBT; whereas in an early study all rats died within 2 days after receiving 3000 mg/kg bw and day. The body weight gain of males from the highest dose group and females of the both highest dose groups were decreased. Treated rats displayed irritable behavior that was more pronounced with increasing dose and was characterized as resistance to gavage. Liver weight and liver weight to body weight ratios were increased in dosed rats with the greatest change occurring at the two highest doses (750 and 1500 mg/kg). No gross or microscopic effects could be related to chemical administration in treated rats. Based on the findings of the study, increased liver weights, decreased body weight gain and irritable behavior after dosing, a LOAEL of 187 mg/kg bw and day is suggested.

Deaths occurred in B6C3F1 mice of the highest dose groups; five of 10 males and 7/10 females that received 1500 mg/kg and 2/10 females dosed with 750 mg/kg bw and day died before the end of the studies. Two of the deaths were related to gavage technique. Chemical administration did not affect body weight gain of treated mice. Liver weights to body weight ratios of dosed groups were higher than those of the vehicle controls. Clonic seizures, lacrimation, and salivation were observed in the 750 and 1500 mg/kg groups. Lethargy and rough coats were observed in the 375 and 750 mg/kg groups. No compound-related gross or microscopic pathologic effects were observed. Based on the findings of the study a NOAEL of 188 mg/kg bw and day is suggested.

The repeated dose toxicity of MBT was evaluated in a chronic feeding study with Slc: ddY mice (Ogawa 1989). Male and female mice (30 per sex treatment groups, 60 per sex control group) were feed with the test substance for 20 months. The animal were treated with 0, 30, 120, 480 and 1920 ppm in the diet (ca. 0, 3.60, 14.69, 57.90, 289.40 mg/kg bw/d in males and 0, 3.61, 13.52, 58.87, 247.98 mg/kg bw/d in females). Haematological parameter, clinical biochemistry and organ weights were examined after 6, 12 and 20 months. For interim dissection 5 animals per treatment group and 10 animals of the control group were sacrificed. No substance-related clinical effects were noted in any of the treated animals. An increase in the mortality rate was seen in males of the high dose group compared to the control group (at 1920 ppm 52.8 % vs. 43.5 % control). The mortality rate in females of the 30 ppm and 1920 ppm groups was higher compared to control, whereas no differences were noted in120 ppm and 480 ppm females. Males from the high dose group gained less weight than control during the whole study. Moreover, males from the 30 ppm and 480 ppm groups gained less weight from week 65 to study termination, whereas males from the 120 ppm treatment group showed a comparable weight gain to the control group. Females from the 120 ppm group showed an increased weight gain from week 35 to study termination. Females from the 30 ppm and 480 ppm groups showed a comparable weight gain to the control group, whereas females from the high dose group showed a transient decrease in weight gain between week 60 and 80. A transient increase in food consumption between week 28 and 50 was noted in high dose males. In males of the high dose group the hematocrit value was significant decreased at study termination. In females of the 480 ppm group a reduction of the hematocrit value and the mean cell volume was noted after 6 months. Moreover, females of this treatment group showed an increase in mean cell hemoglobin and mean cell hemoglobin concentration at study termination (20 months). All other haematological parameters were comparable to the control group. In clinical biochemistry, organ weights and macroscopic evaluations no biologically relevant changes were detected in treated animals compared to control. The histopathological evaluations revealed cell infiltration in the interstitium of the kidney in mid (480 ppm) and high dose males (1920 ppm) at study termination (control: 1/14, 30 ppm: 1/6, 120 ppm: 1/7, 480 ppm: 4/6, 1920 ppm: 5/9). Cell infiltration in the interstitium of the kidney was also noted in females dosed for 20 months with the test substance (control: 9/15, 30 ppm: 2/7, 120 ppm: 5/9, 480 ppm: 3/10, 1920 ppm:1/7). All other histological findings were comparable with the control group. The authors suggested that the NOAEL is 120 ppm (14.6 mg/kg bw/d males, 13.52 mg/kg bw/d females) based on the cell infiltration in the interstitium of the kidney in males. However, according to MAK (1999) the biological relevance of these findings is questionable, because of the low number of animals used in this study, the absence of a clear dose response and the lack of a statistic evaluation of the study.

A two-generation reproductive toxicity study was performed to evaluate the potential effects of MBT on reproduction and development in Sprague-Dawley rats (CMA 1990). The study was designed to determine if MBT has any adverse effects on reproduction functions when fed to F0 and Fl parental animals for a minimum of 70 days prior to mating and continuing until sacrifice. Groups of 28 male and 28 female Sprague-Dawley rats were fed the basal diet or diet containing MBT at concentrations of 2500, 8750, and 15000 ppm. The food was provided ad libitum throughout the study. All F0 and Fl rats were observed daily for signs of overt toxicity, morbidity, or mortality. Body weights were measured weekly for the males throughout the study. For females, body weights were measured weekly prior to confirmation of mating and at specified intervals during gestation and lactation. Food intake was measured on the same days as body weights with the exception of periods of cohabitation when food intake was not measured. F0 and Fl parents were sacrificed and necropsied following weaning of their offspring. Microscopic examination was performed on all tissues collected from the control and 15000 ppm groups. Kidneys from the F0 rats in the other treated groups and liver and kidneys from the Fl rats were also examined microscopically. Liver, kidneys, testes or ovaries from all F0 and Fl parents sacrificed for the scheduled necropsy were weighed. Survival was 100% in all F0 and Fl parental animals. There were no treatment-related clinical signs observed in any of the MBT groups. Food intake, calculated as g/kg/day, was significantly reduced in the 8750 and 15000 ppm groups of the F0 generation during the first week of treatment. Thereafter, food intake was equal to or greater than in the control group. Body weight gain, however, was dose-dependent and significantly reduced in F0 males from all MBT groups and females from mid and high dosage groups during the first week of treatment. Weight gain continued to be slightly reduced for approximately ten weeks for males but not for females. Treatment-related histopathological changes were seen in the kidneys of both F0 and Fl animals. Brown pigment was observed in the lumen and epithelial cells of the proximal convoluted tubules in males and females in the mid and high dosage groups, with a greater incidence in the males than in females. Cortical tubular basophilia and alpha 2 µ-globulin inclusions in the epithelial cells of the proximal convoluted tubules occurred in males from all groups with a higher incidence in the treated groups. In addition, absolute and relative kidney weights were significantly increased for F0 and F1 males in the mid and high dosage groups. Microscopic changes, consisting of hepatocyte hypertrophy, occurred in the livers of the Fl animals from the 15000 and 8750 ppm groups, at a higher incidence in males than in females. The increased workload due to continuous exposure to the test article, apparently led to hepatic hypertrophy. Furthermore, histopathological changes correlated with a significant increase of relative liver weight in the males in the mid and high dosage groups and for females in the high dosage group. Hepatic lesions, noted in the Fl animals are probably related to a greater intake of the test article, since Fl pups started to eat the test diet at approximately 14 days of age, while the F0 animals were administered MBT beginning at seven weeks of age. There were no other treatment-induced changes in any of the organs examined microscopically from rats in the MBT groups of either the F0 or F1 generation. The authors concluded that minimal to mild toxic effects (reduced body weights, increase in absolute and relative kidney and liver weights combined with histopathological changes in the kidneys and liver) were observed in parental animals from all MBT groups. These effects were more prominent in the Fl generation due to a greater intake and longer exposure to the test article. Based on findings of this study (reduced body weight gain) a LOAEL of 2500 ppm (ca. 150 to 250 mg/kg bw MAK 1999) is suggested.

In conclusion:

There are only limited repeated dose toxicity data available for MBTS from an early cancer study in mice; which indicates that MBTS was not carcinogenic under the experimental conditions used. A read across approach with repeated dose toxicity data from MBT was conducted. There are several repeated dose toxicity studies available for MBT. In a two-generation toxicity study (CMA 1990) a LOAEL of 2500 ppm (ca. 150 mg to 250 mg/kg bw) was observed. In a two year carcinogenicity study with Fischer 344 rats (NTP 1988) a LOAEL of 188 mg/kg bw and day for females and of 375 mg/kg bw and day for males was determined. In a subchronic toxicity study (NTP 1988) a LOAEL of 187 mg/kg bw was noted. In mice a LOAEL of 375 mg/kg bw was suggested in a two-year carcinogenicity study (NTP 1988) and a NOAEL of 188 mg/kg bw in a subchronic toxicity study (NTP 1988). Thus, the NOAEL and LOAEL values from several MBT repeated dose studies are in the same range. The consistent findings in subchronic, chronic and reproduction/ developmental toxicity studies revealed a LOAEL in the range of 150 to 375 mg/kg bw and day. Following the recommendation given in the MAK publication (1999) an overall NOAEL of 50 mg/kg bw and day is suggested. This assumption based on data from the two-generation toxicity study and is supported by the consistent findings in the subchronic and chronic toxicity studies.

The overall NOAEL of 50 mg/kg is used for systemic DNEL calculation

Justification for classification or non-classification

No classification is required according to the classification criteria 67/548/EWG and regulation no. 1272/2008 (GHS).