N-tert-butylbenzothiazole-2-sulphenamide

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study, comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1985

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 408 (Repeated Dose 90-Day Oral Toxicity Study in Rodents)

Principles of method if other than guideline:

Twenty male and twenty female Sprague-Dawley rates received 0, 100, 300 or 1000 mg N-tert-butylbenzothiazole-2-sulphenamide/kg bw by gavage. The animals were examined for mortality, overt signs of toxicity, body weight and food consumption. Clinical chemistry, haematological and urinalysis determinations were performed. At sacrifice animals were subjected to a complete gross necropsy and a microscopic examination for the control and high dose group performed.

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

N-tert-butylbenzothiazole-2-sulphenamide

EC Number:

202-409-1

EC Name:

N-tert-butylbenzothiazole-2-sulphenamide

Cas Number:

95-31-8

Molecular formula:

C11H14N2S2

IUPAC Name:

N-(1,3-benzothiazol-2-ylsulfanyl)-2-methylpropan-2-amine

Details on test material:

Santocure NS, purity (end of the study): 96.44 %

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

The animals were ca. six week of age and weighed 175-213 g for males and 130-159 g for females at the start.

Administration / exposure

Route of administration:

oral: gavage

Details on route of administration:

Oral gavage is a commonly applied route of administration for studies of this type.

Vehicle:

corn oil

Details on oral exposure:

Dosing suspensions were prepared weekly. All animals were administered a constant volume of suspensions (0.5 ml of suspension per 100 g of body weight).

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Samples were analysed weekly using a liquid chromatographic procedure. Analytical results demonstrated that the test material was stable and appropriate dose suspensions had been prepared.

Duration of treatment / exposure:

90 d

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

0 mg/kg bw/day (nominal)

Remarks:

Vehicle control

Doses / concentrations 2

Dose / conc.:

100 mg/kg bw/day (nominal)

Remarks:

Low

Doses / concentrations 3

Dose / conc.:

300 mg/kg bw/day (nominal)

Remarks:

Mid

Doses / concentrations 4

Dose / conc.:

1 000 mg/kg bw/day (nominal)

Remarks:

High

No. of animals per sex per dose:

20 per dose and sex

Control animals:

yes, concurrent vehicle

Details on study design:

Post-exposure period: no

Positive control:

No.

Examinations

Observations and examinations performed and frequency:

Examinations:

Mortality: twice daily observations

Body weight gain and food consumption: determination weekly

Clinical appearance: checked daily

Hematology (total erythrocyte count (RBC), total leukocyte count (WBC), platelets count (Plt), hematocrit (Hct), level of hemoglobin, red blod cell indices (MCV, MCH, MCHC), determination of the leukocyte differential, reticulocyte count

Clinical Chem.: albumin, total protein, blood urea nitrogen (BUN), total bilirubin, direct bilirubin, glucose, glutamic pyruvic transaminase (D-GPT/ALT), alkaline phosphatase, glutamic oxaloacetate transaminase (D-GOT/AST), gamma glutamyl transpeptidase (Gamma-GT, phosphorous, creatine, cholesterol (Chol), calcium, chloride, sodium, potassium and globulin

Urine analysis: pH, urine protein, blood, glucose, ketone, bilirubin, urobilinogen, urine specific gravity

Sacrifice and pathology:

Organ weights:brain, kidney, liver, testes

Organs prepared: aorta, adrenal glands, bone and bone marrow, brain, epididymis, esophagus, eyes, heart, pancreas, prostate gland, pituitary gland, salivary gland, skin, small intestine, kidneys, colon, lesions, liver, lung, lymph node, mammary gland, muscle, nerve, spinal cord, spleen, stomach, testes, ovaries, thymus, thyroid, parathyroid, trachea, uterus, urinary bladder

Statistics:

Quantitative data were analyzed by Dunnett's t-test to determine significant differences in treatment values versus control at the 95% confidence level. Organ weight/terminal body weights were analyzed by the Mann-Whitney U test and incidence of microscopic abnormalities with the Fischer exact test, both utilizing the Bonferri Inequality procedure.

Results and discussion

Results of examinations

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Animals in all treatment groups salivated after dosing and mid and high dose animals salivated prior to dosing or sometimes resisted dosing. High dose females exhibited urine stained abdomens. Other than the above, no clinical signs or abnormal behavior related to treatment were observed. Due to the lack of gross or microscopic findings of gastrointestinal effects, the behavioral reactions were considered to be related to poor palatability of the test material rather than a toxic response.

Mortality:

no mortality observed

Description (incidence):

All animals survived three months of exposure to the test substance.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Reduced body weights were noted for males at the middle and highest dose groups (300 and 1000 mg/kg bw/day, respectively). Most affected were the high dose group males with 13 weeks reduction at 99% statistical confidence. Middle group males were affected the final 11 out of 13 weeks, primarily at 95% statistical confidence. Body weights were unaffected for the low dose males (100 mg/kg bw/day) and all three female treatment groups as compared to controls.
There was a linear decrease in terminal body weights of males from Groups 1, 2 and 3. There were, however, essentially no changes in weights of females from treated groups as compared to controls.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

Similar amounts of food were consumed by both sexes from all three treatment groups when compared to their control groups. Two exceptions were mid dose group males (300 mg/kg/day) at week 9 and high dose males (1,000 mg/kg/day) at week 1, however, these exceptions appear random and are probably not related to exposure with the test item.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

effects observed, non-treatment-related

Description (incidence and severity):

There were several statistically significant changes in mean values for hematologic or serum chemistry parameters. Statistically significant increases in reticulocyte counts occurred in Group 2 males (300 mg/kg bw/d) and Group 3 (1000 mg/kg bw/d) females. These values remained well within normal limits and were of no toxicological significance.

Clinical biochemistry findings:

effects observed, treatment-related

Description (incidence and severity):

There were several statistically significant changes in mean values for hematologic or serum chemistry parameters. Depressions of SGOT and SGPT at 100 and 1000 mg/kg bw in females were probably artifactual due to high group mean values for these enzymes in control females. When clearly abnormal values for NFOO4 and NFO15 are deleted, a minimal (p 0.05) depression was found affecting only Group 1 females (100 mg/kg bw/d). These changes were not considered to be relevant. Minimal depressions in sodium affected all dose level of females and Group 2 (300 mg/kg bw/d) males. These were within normal range and were not considered to be toxicologically relevant. The only change which may have been biologically relevant was a mild increase in the mean cholesterol value for females from Group 3 (1000 mg/kg bw /d).

Urinalysis findings:

effects observed, treatment-related

Description (incidence and severity):

Urine specific gravity was increased in males from Group 3 (1000 mg/kg bw) and females from Groups 2 and 3 (300 and 1000 mg/kg bw/d). The increased values for males and females from Group 3 were statistically significant. There were increases in the occurrence of mild ketonuria in male Groups 1, 2 and 3.

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

The only significant changes in absolute organ weights were decreased mean brain weight in females from Group 3 (1000 mg/kg bw/d) and increased mean hepatic weight in females from Group 3 (1000 mg/kg bw/d). Organ to body weight ratios were increased in a dose related manner for all male organs for which weights were obtained (statistically significant in most instances) for Groups 2 and 3. Kidneys to body weight ratios were increased in females from Group 3 (statistically significant) as were liver to body weight ratios in females from Groups 2 and 3 (300 and 1000 mg/kg bw/d).

Gross pathological findings:

no effects observed

Description (incidence and severity):

There were no gross lesions associated with chemical administration.

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

There were no microscopic lesions with an apparent association with chemical administration.

Histopathological findings: neoplastic:

not specified

Effect levels

1

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse effects

2

Dose descriptor:

LOAEL

Effect level:

300 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: decreased body weight in males, increase in urine specific gravity in females

Target system / organ toxicity

Key result

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

100 mg/kg bw/day produced no toxicologically significant changes and is therefore set as NOAEL.
The LOAEL is set at 300 mg/kg bw/day, based on decreased body weight in males, an increase in urine specific gravity in females and increase in relative liver and kidney organ weights.

NB. Some parameters (neurobehavioral examinations, endocrine parameters) are not present in the current study when compared to the modern OECD 408 test guideline. A testing proposal is in place for an OECD 443 EOGRTS, which will provide fully up to date data on not only reproduction endpoints but also endocrine and systemic toxicity.

Executive summary:

Study design

The test substance was administered orally by gavage to four groups of 20 male and 20 female Sprague-Dawley rats at doses of 0, 100, 300 or 1000 mg/kg/day for 3 months.

 

Results

All animals survived the treatment period. changes appearently associated with the test substance exposure at 1000 mg/kg7day included increased urine specific gravity and decreased body weights among males. Females at this dose level had stained abdomens, increased urine specific gravity, elevated cholesterol values, and increased liver weights. Middle dose level (300 mg/kg/day) males had decreased body weight and females had increased urine specific gravity. 

 

Conclusion

100 mg/kg bw/day produced no toxicologically significant changes and is therefore set as NOAEL.
The LOAEL is set at 300 mg/kg bw/day, based on decreased body weight in males, an increase in urine specific gravity in females and increase in relative liver and kidney organ weights.