Registration Dossier

Administrative data

Description of key information

Repeated dose toxicity: Oral

The No Observed Adverse Effect Level (NOAEL) of the test chemical using rats via oral route, is considered to be 200 mg/kg body weight.

Repeated dose toxicity: Inhalation

According to Annex IX of the REACH regulation, testing by the inhalation route is appropriate only if exposure of humans via inhalation is likely. Taking into account the low vapour pressure of the substance 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione (37052-78-1) which is reported as 5.45E-6mmHg at 25 C. Thus, exposure to inhalable dust, mist and vapour of the 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione is highly unlikely. Therefore this study is considered for waiver.

Repeated dose toxicity: Dermal

The acute toxicity value for 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione (37052-78-1) (as provided in section 7.2.3) is >2000 mg/kg body weight. Also, given the use of the chemical; repeated exposure by the dermal route is unlikely since the use of gloves is common practice in industries. Thus, it is expected that 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione shall not exhibit 28 day repeated dose toxicity by the dermal route. In addition, there is no data available that suggests that 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione shall exhibit repeated dose toxicity by the dermal route. Hence this end point was considered for waiver.

Key value for chemical safety assessment

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records
Reference
Endpoint:
short-term repeated dose toxicity: oral
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
data from handbook or collection of data
Justification for type of information:
data from handbook or collection of data
Reason / purpose:
read-across source
Reason / purpose:
read-across source
Qualifier:
according to
Guideline:
other: Refer below principle
Principles of method if other than guideline:
WoE derived based on the experimental data summerised below
1,To assess toxicological profile of the test chemical to determine target organ of toxicity, its reversibility and No Observed Adverse Effect Level (NOAEL) in the rat after 28 consecutive days of oral administration.
2,Repeated dose oral toxicity study was performed to determine the toxic nature of the test chemical
GLP compliance:
not specified
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Remarks:
1.
Sex:
male/female
Details on test animals and environmental conditions:
1. TEST ANIMALS
- Source:National Institute of Biosciences, Pune
- Females (if applicable) nulliparous and non-pregnant:[yes
- Age at study initiation:6 to 8 weeks
- Weight at study initiation: Male - 173.58 g (= 100 %), Female - 153.64 g (= 100 %)
- Fasting period before study:
- Housing: The rats were housed in polycarbonate cages with paddy as bedding.
After allocation to respective dose groups rats were housed 2/sex/cage.
- Diet (e.g. ad libitum): Rodent feed, ad libitum
- Water (e.g. ad libitum): Water was provided ad libitum from individual bottles attached to the cages. Water was from a local source and passed through the reverse osmosis membrane before use
- Acclimation period: 5 days

DETAILS OF FOOD AND WATER QUALITY: There were no known contaminants, which were reasonably expected to be in the dietary materials and water capable of interfering with the conduct of this study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3°C
- Humidity (%): 30% to 70%
- Air changes (per hr): The animal room was independently provided with at least ten air changes per hour of 100% fresh air that has been passed through the HEPA filters

2. No data
- Photoperiod (hrs dark / hrs light): An artificial light and dark cycle of 12 hours each was provided to the room

IN-LIFE DATES:
From: 12-12-2017
To: 19-03-2018
Route of administration:
oral: gavage
Details on route of administration:
No data
Vehicle:
other: 1. Corn oil; 2. 2% carboxymethylcellulose
Details on oral exposure:
1. PREPARATION OF DOSING SOLUTIONS:The test item was suspended in corn oil for preparation of suspension(s). The suspension(s) of the test chemical were made at volumes suitable for daily use for 28 days. The suspension(s) were prepared at concentrations of 0, 25, 50 and 100 mg/ml such that dosage of 0 (vehicle), 250, 500 and 1000 mg/kg body weight respectively were administered.

DIET PREPARATION
- Rate of preparation of diet (frequency):
- Mixing appropriate amounts with (Type of food):
- Storage temperature of food:

VEHICLE
- Justification for use and choice of vehicle (if other than water): corn oil
- Concentration in vehicle: 0 (vehicle), 250, 500 and 1000 mg/kg body weight
- Amount of vehicle (if gavage): 10 ml/kg body weight.
- Lot/batch no. (if required):
- Purity: No data

2. PREPARATION OF DOSING SOLUTIONS: The test chemical was dissolved in 2% carboxymethylcellulose to give dose level of 0, 12.5, 100, 200 or 400 mg/Kg

DIET PREPARATION
- Rate of preparation of diet (frequency): No data
- Mixing appropriate amounts with (Type of food): No data
- Storage temperature of food: No data

VEHICLE
- Justification for use and choice of vehicle (if other than water): 2% carboxymethylcellulose
- Concentration in vehicle: 0, 12.5, 100, 200 or 400 mg/Kg
- Amount of vehicle (if gavage): No data
- Lot/batch no. (if required): No data
- Purity: No data
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
1. The test item formulation analysis for concentration and stability were conducted
2. No data
Duration of treatment / exposure:
1. 28 days
2. 180 days
Frequency of treatment:
1. Daily
2. 7 days/week
Dose / conc.:
0 mg/kg bw/day (actual dose received)
Remarks:
1
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Remarks:
1
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Remarks:
1
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Remarks:
1
Remarks:
2. Females: 0, 12.5, 100, 200 or 400 mg/Kg

Males: 0, 12.5, 50, 100, 200 or 400 mg/Kg
No. of animals per sex per dose:
1. Total: 72
0 mg/kg bw: 6 male, 6 female
250 mg/kg bw: 6 male, 6 female
500 mg/kg bw: 6 male, 6 female
1000 mg/kg bw: 6 male, 6 female

Reversal group
500 mg/kg bw: 6 male, 6 female
1000 mg/kg bw: 6 male, 6 female

3. Total: 300
0 mg/Kg/day (male/female): 30 males and 30 females
12.5 mg/Kg/day (male/female): 30 males and 30 females
50 mg/Kg/day (males): 30 males
100 mg/Kg/day (female): 30 females
200 mg/Kg/day (male/female): 30 males and 30 females
300 mg/Kg/day (male/female): 30 males and 30 females
Control animals:
yes, concurrent vehicle
Details on study design:
1. - Dose selection rationale: Dose were selected basd on Dose Range Finding study:
1) All the male and female animals from control and all the treated dose groups up to 1000 mg/kg survived throughout the dosing period of 7 days.
2) Male and female animals from control and all the treated dose groups exhibited normal body weight gain at the end of the dosing period of 7 days.
3) Daily clinical observations did not reveal any signs of toxicity in male and female animals from different dose groups during the dosing period of 7 days.
4) Gross pathological examination did not reveal any abnormality attributable to the treatment.
Based on these results, the 28 day study dose levels were finalized as 0 mg/kg, 250 mg/kg, 500 mg/kg and 1000 mg/kg body weight and animals were exposed to the treatment every day for a period of 28 days.

- Rationale for animal assignment (if not random): A total of 72 animals (36 males + 36 females) were selected and randomly distributed into six groups with 6 animals/sex/group for main groups and 6 animals /sex /group for reversal group.
The animals of uniform body weight were selected. The individual body weights of the animals did not exceed ± 20% of group mean body weight. The group means body weights of all the groups were approximately equal..
- Rationale for selecting satellite groups:
- Post-exposure recovery period in satellite groups: 14 days
- Section schedule rationale (if not random):

2. No data
Positive control:
No data
Observations and examinations performed and frequency:
1. CAGE SIDE OBSERVATIONS: Yes
- Time schedule:twice daily
- Cage side observations checked in table [No.?] were included.: Viability observed.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule:once daily

BODY WEIGHT: Yes
- Time schedule for examinations: Body weights were recorded on the day of randomization, day of first dosing, weekly thereafter and a fasting body weight at scheduled sacrifice on day 29 and day 43.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: Yes

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: Not specified

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes / No / Not specified
- Time schedule for examinations:

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations:The eyes of all the animals were examined prior to the initiation of the dosing and at scheduled sacrifice. Eye examination was carried out by using a HEINE mini 2000 ophthalmoscope were employed for evaluation after the induction of mydriasis with 1% solution of tropicamide sulfate.
- Dose groups that were examined: All 72 animals

HAEMATOLOGY: Yes
- Time schedule for collection of blood: At the end of dosing period on day 29 and at termination of recovery period on day 43.
- Anaesthetic used for blood collection: No
- Animals fasted: Yes
- How many animals: All 72 animals
- Parameters checked in table [No.?] were examined.: Hb (Hemoglobin (g/dL)), RBC (Red Blood Corpuscles (x 106 /µL)), HCT (Hematocrit (%)), MCV (Mean Corpuscular Volume (fL)), MCH (Mean Corpuscular Hemoglobin (pg)), MCHC (Mean Corpuscular Hemoglobin Concentration (g/dL)), Platelets (x 103 /µL), WBC (White Blood Corpuscles (x 103 /µL)), Rt.(Reticulocytes (%)), N (Neutrophils (%)), L (Lymphocytes (%)), E (Eosinophils (%)), M (Monocytes (%)), B (Basophil (%)) and Pt. (Prothrombin time (sec.)) were examined.


CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: At the end of dosing period on day 29 and at termination of recovery period on day 43.
- Animals fasted: Yes
- How many animals:All 72 animals
- Parameters checked in table [No.?] were examined.: Total Protein (g/dL), Blood Urea Nitrogen (mg/dL), Urea Nitrogen (mg/dL) Calculated, ALT : Alanine Aminotransferase (U/L), AST, Aspartate Aminotransferase (U/L), ALP : Alkaline Phosphatase (U/L), GGT : Gamma Glutamyl Transferase (U/L), Glucose (mg/dL), Calcium (mmol/L), Phosphorous (mg/dL), Albumin (g/dL), Total Bilirubin (mg/dL), Creatinine (mmol/L), Total Cholesterol (mg/dL), Triglycerides (mg/dL), Globulin (g/dL) Calculated, Sodium (mmol/L), Potassium (mmol/L), Chloride (mmol/L) and
Bile acid (µmol/L) were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: During the last week of dosing period and on reversal group rats at termination of recovery period on day 43.
- Metabolism cages used for collection of urine: Yes
- Animals fasted: Not specified
- Parameters checked in table [No.?] were examined.: Volume , Appearance, Colour, pH , Specific Gravity, Proteins, Glucose, Ketones,Bilirubin, Urobililogen, Occult Blood and Nitrite were examined.

NEUROBEHAVIOURAL EXAMINATION: Yes
- Time schedule for examinations: Towards the end of the exposure period of 28 days and towards the end of the recovery period on day 42, sensory reactivity to stimuli of different types (e.g. auditory, visual and proprioceptive stimuli) assessment of grip strength and motor activity assessment were conducted for all the animals
- Dose groups that were examined: All 72 animals
- Battery of functions tested: sensory activity / grip strength / motor activity / other:Arousal level, Sensory Activity, Visual Placing Response, Air righting response, Grip Strength, Motor Activity,

IMMUNOLOGY: Not specified
- Time schedule for examinations:Not specified
- How many animals:Not specified
- Dose groups that were examined:Not specified
- Parameters checked in table [No.?] were examined.Not specified

OTHER: Organ Weights were examined.

2. Observations and examinations performed & frequency
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Each day at the time of treatment
- Cage side observations checked in table [No.?] were included. general condition, mortality and behavior

DETAILED CLINICAL OBSERVATIONS: No data
- Time schedule: No data

BODY WEIGHT: Yes
- Time schedule for examinations: at weekly intervals

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No data
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data

FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No data

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
- Time schedule for examinations: No data

OPHTHALMOSCOPIC EXAMINATION: No data
- Time schedule for examinations: No data
- Dose groups that were examined: No data

HAEMATOLOGY: Yes
- Time schedule for collection of blood: after 30, 60, 100, and 180 days of treatment
- Anaesthetic used for blood collection: No data
- Animals fasted: No data
- How many animals: No data
- Parameters checked in table [No.?] were examined. No data

CLINICAL CHEMISTRY: No data
- Time schedule for collection of blood: after 30, 60, 100, and 180 days of treatment
- Animals fasted: No data
- How many animals: No data
- Parameters checked in table [No.?] were examined. No data

URINALYSIS: No data
- Time schedule for collection of urine: No data
- Metabolism cages used for collection of urine: No data
- Animals fasted: No data
- Parameters checked in table [No.?] were examined. No data

NEUROBEHAVIOURAL EXAMINATION: No data
- Time schedule for examinations: No data
- Dose groups that were examined: No data
- Battery of functions tested: sensory activity / grip strength / motor activity / other: No data

OTHER: No data
Sacrifice and pathology:
1. GROSS PATHOLOGY: Yes, All the rats surviving at the end of the treatment were sacrificed and gross lesions were noted.

HISTOPATHOLOGY: Yes, From each rat, samples or the whole of the tissues listed below were preserved. All tissues were fixed in 10% neutral buffered formalin except, eyes and testes of all animals were preserved in Davidson’s solution for 24 hours and transferred to 10% neutral buffered formalin.
Procedure for preparation of slides of tissues of various organs from the rats of various dose groups were performed as per the standard operating procedures of Indian Institute of Toxicology, Pune.
Following tissue samples of organs from control and animals treated at different dose groups were preserved and those from control and treated at the highest dose level of 1000 mg/kg were subjected to histopathological examination.

Adrenals, Aorta, Brain (cerebrum, cerebellum and pons), Caecum, Cervix, Colon, Duodenum, Epididymides, Eyes, Heart, Ileum, Jejunum, Kidneys, Liver, Lungs, Mesenteric Lymphnodes, Muscles - Skeletal muscle, Oesophagus, Ovaries, Pancreas, Pharyngeal Lymphnodes, Pituitary, Prostate, Rectum, Sciatic Nerve, Seminal Vesicles with coagulation gland, Skin with Mammary Gland, Spleen, Spinal Cord (Cervical, mid thoracic and lumbar), Sternum with bone marrow, Stomach, Testes, Thymus, Trachea, Thyroid / Parathyroid, Urinary Bladder, Uterus, Vagina.

2. GROSS PATHOLOGY: Yes, 300 animals were sacrificed after 30, 60, 100, and 180 days of treatment tissues for gross examination

HISTOPATHOLOGY: Yes, 300 animals were sacrificed after 30, 60, 100, and 180 days of treatment tissues for microscopic examination
Other examinations:
No data
Statistics:
1. Raw data was processed and analyzed for reporting group means and standard deviations with significance between the controls and treated groups, using SYSTAT 13 validated statistical software supplied by Starcom Information Technology Limited, Bangalore developed by Systat Software, Inc. USA. All the parameters characterized by continuous data such as body weight, feed consumption (calculated as gram per animal), organ weight, relative organ weight, haematological and clinical chemistry data were subjected to Bartlett’s test to meet the homogeneity of variance before conducting Analysis of Variance (ANOVA) and Dunnett’s t-test. Where the data does not meet the homogeneity of variance, Student’s t-test were performed to calculate significance.

Significance was calculated at 5% level and indicated in the summary tables as follows:

* = Significant than control at 95% level of confidence (p<0.05).

2. No data
Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
1. Male -
Group I (Control, 0 mg/kg): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days (animal nos.1 to 6).
Group II (Control, 0 mg/kg, Reversal): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days and during the post-dosing recovery period (animal nos.13 to 18).
Group III (250 mg/kg): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days (animal nos.25 to 30).
Group IV (500 mg/kg): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days (animal nos.37 to 42).
Group V (1000 mg/kg): Polyurea was observed in animals (animal nos. 49, 51 to 54, with onset from day 23) during the dosing period of 28 days.
Group VI (1000 mg/kg, Reversal): Polyurea was observed in animals (animal nos. 61, 63 to 65, with onset from day 23) throughout the dosing period of 28 days and during the post-dosing recovery period (upto day 32).

Female -
Group I (Control, 0 mg/kg): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days (animal nos.7 to 12).
Group II (Control, 0 mg/kg, Reversal): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days and during the post-dosing recovery period (animal nos.19 to 24).
Group III (250 mg/kg): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days (animal nos.31 to 36).
Group IV (500 mg/kg): No clinical signs of toxicity were observed in the animals throughout the dosing period of 28 days (animal nos.43 to 48).
Group V (1000 mg/kg): Polyurea was observed in all animals (animal nos. 55 to 60, with onset from day 22) during the dosing period of 28 days.
Group VI (1000 mg/kg, Reversal): Polyurea was observed in all animals (animal nos.67 to 72, with onset from day 22) throughout the dosing period of 28 days and during the post-dosing recovery period (upto day 32).

Before commencement of treatment:
In home cage observation, rat from different dose groups and control group revealed normal behavior, alterations, vocalization, respiration and palpebral closer.
During handling observation, handling of rats did not reveal any abnormality from different dose groups and control group.
In the open field observation, rat did not reveal any abnormality from different dose groups and control group.
During treatment:
In home cage observation, rat from different dose groups and control group revealed normal behavior, alterations, vocalization, respiration and palpebral closer.
During handling observation, handling of rats did not reveal any abnormality from different dose groups and control group.
In the open field observation, rat did not reveal any abnormality from different dose groups and control group.
Detailed clinical observation did not reveal any abnormality in all groups during the dosing period of 28 days and during the post-dosing recovery period.

2. No untoward signs of toxicity were noted
Mortality:
no mortality observed
Description (incidence):
1. All animals from control and different dose groups survived throughout the dosing period of 28 days and the post-dosing recovery period of 14 days.

2. All animals survived through the treatment period
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
1. Male -
Reduced body weight gain of 14.94% was observed in male animals from 1000 mg/kg dose group.
All male animals from control, 250 mg/kg, 500 mg/kg and 1000 mg/kg reversal dose groups exhibited normal body weight gain throughout the dosing period of 28 days and the recovery period of 14 days.
Female -
Female animals from control, 250 mg/kg, 500 mg/kg and 1000 mg/kg dose groups exhibited normal body weight gain throughout the dosing period of 28 days.
Reduced body weight gain of 11.36% was observed in female animals from 1000 mg/kg reversal group.

2. A daily dose of 200 mg/kg or less caused no depression in rate of weight gain in female rats, and even at the 400 mg/kg level the difference in the weight curves between the controls and treated animals was not striking. In contrast, in male rats, dose levels of 200 mg/kg or more produced some inanition and significantly affected the rate of weight gain.
Food consumption and compound intake (if feeding study):
no effects observed
Description (incidence and severity):
1. Male and Female -
Animals from control and different dose groups exhibited normal feed consumption at the end of the dosing period of 28 days.
Animals from control reversal and high reversal dose groups exhibited normal feed consumption at the end of the recovery period of 14 days.

2. No data
Food efficiency:
not specified
Description (incidence and severity):
2. No data
Water consumption and compound intake (if drinking water study):
not specified
Description (incidence and severity):
2. No data
Ophthalmological findings:
no effects observed
Description (incidence and severity):
1. No ocular abnormalities were observed on ophthalmological examination in the animals during pre-exposure and at the end of the respective termination.

2. No data
Haematological findings:
effects observed, non-treatment-related
Description (incidence and severity):
1. Male :
Total RBC and HCT: Decreased levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05).
Female :
Hb and HCT: Decreased values were obtained for animals from 500 mg/kg dose group, sacrificed on day 29 (p<0.05) and
Total RBC: Decreased values were obtained for animals from 250 mg/kg and 500 mg/kg dose groups, sacrificed on day 29 (p<0.05).

2. No data
Clinical biochemistry findings:
effects observed, non-treatment-related
Description (incidence and severity):
1. Male :
Globulin: Elevated levels were observed in animals from 250 mg/kg dose group, sacrificed on day 29 (p<0.05),
BUN and Urea Nitrogen: Decreased levels were observed in animals from 250 mg/kg dose group, sacrificed on day 29 (p<0.05) and
Calcium : Decreased levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05).
Female :
BUN and Urea Nitrogen: Decreased levels were observed in animals from 500 mg/kg dose group, sacrificed on day 29 (p<0.05),
Alanine Aminotransferase : Decreased levels were observed in animals from 250 mg/kg and 500 mg/kg dose groups, sacrificed on day 29 (p<0.05),
Alkaline Phosphatase: Decreased levels were observed in animals from 250 mg/kg dose group, sacrificed on day 29 (p<0.05) and
Bile Acid : Elevated levels were observed in animals from 1000 mg/kg reversal dose group, sacrificed on day 43 (p<0.05).

2. There were no striking biochemical changes that could be attributed to drug treatment in any of the rats
Urinalysis findings:
effects observed, non-treatment-related
Description (incidence and severity):
1. Urine analysis conducted on male animals during 4th and 6th week of study period (on day 23, 24 and 43), revealed no abnormality attributable to the treatment.
Urine analysis conducted on female animals during 4th week of study period (on day 24 and 26), revealed higher volume of urine from 1000 mg/kg dose group (p<0.05).
Urine analysis conducted on female animals during 6th week of study period (on day 43) revealed no abnormality attributable to the treatment.

2. No data
Behaviour (functional findings):
no effects observed
Description (incidence and severity):
1. Sensory Reactivity Observations:
All animals from control and different dose groups showed normal arousal level, visual response, touch response, auditory response, tail pinch response and visual placing response. Normal air righting reflex was observed in all animals from control and different dose groups in week 4.

Grip Strength:
Grip strength values observed in male and female animals for control and different dose groups were comparable.

Motor Activity:
Motor activity values observed in male and female animals for control and different dose groups were comparable.

2. No data
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
effects observed, non-treatment-related
Description (incidence and severity):
1. Male -
In comparison with controls at the end of dosing on day 29, organ weight data of animals from 1000 mg/kg dose group revealed increased relative weights of brain (p<0.05). Increased relative weights of liver and kidneys (p<0.05) were observed in animals from 500 mg/kg and 1000 mg/kg dose groups. Increased relative weights of spleen (p<0.05) was observed in animals from 250 mg/kg and 1000 mg/kg dose groups. Increased relative weights of thymus (p<0.05) was observed in animals from 250 mg/kg dose group.
In comparison with controls at the end of post-dosing recovery period on day 43, organ weight data of animals from 1000 mg/kg reversal group was found to be comparable.
Female -
In comparison with controls at the end of dosing on day 29, organ weight data of animals from 1000 mg/kg dose group revealed increased relative weights of liver (p<0.05). Increased relative weights of kidneys (p<0.05) was observed in animals from 500 mg/kg and 1000 mg/kg dose groups. In addition, decreased relative weights of heart (p<0.05) was observed in animals from 500 mg/kg dose group.
In comparison with controls at the end of post-dosing recovery period on day 43, organ weight data of animals from 1000 mg/kg reversal group revealed increased relative weights of brain, liver, kidneys, spleen and uterus (p<0.05).
Although significant changes in the values of organ weight were observed in male and female animals from 250 mg/kg and 500 mg/kg dose groups, no related gross pathological and histopathological findings were seen, hence these findings were considered to be of no toxicological importance.

2. Organ weights in treated animals did not differ from those of controls.
Gross pathological findings:
no effects observed
Description (incidence and severity):
1. Gross pathological examination on male and female animals from control and different dose groups did not reveal any abnormality.

2. Gross examination revealed no relevant changes in any of the organs.
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
effects observed, treatment-related
Description (incidence and severity):
1. Histopathological examination revealed focal to diffuse, minimal to moderate tubular dilatation in kidneys in male (1/6) and female (6/6) animals from 1000 mg/kg dose group, in male (1/6) and female (6/6) animals from 500 mg/kg dose group and in male (0/6) and female (5/6) animals from 250 mg/kg dose group and in female (1/6) from 1000 mg/kg reversal dose group.
Focal to multifocal, minimal mononuclear cells infiltration was observed in kidneys in male (1/6) and female (3/6) animals from 1000 mg/kg dose group and in female animals from 250 mg/kg (2/6) and 500 mg/kg (2/6) dose groups.
The incidence of tubular dilatation and mononuclear cells infiltration were observed in male animals were less as compared to female animals, however, effect observed was found reversible in male animals at the end of 14 day reversal period of the study.
Incidental, physiological and congenital histopathological changes which were covered in the background historical data of the pathology from control and high dose groups includes minimal, focal to multifocal periportal mononuclear cells infiltration in liver; minimal, multifocal tubular eosinophilic secretion in the kidneys; minimal, multifocal brown coloured pigmentation in the spleen; minimal, diffuse dilatation of zona reticularis and/or minimal, multifocal vacuolation in zona fasciculata in the adrenals; minimal, luminal seminal coagulum in urinary bladder; minimal, luminal dilatation in the uterus; presence of persistent Rathke’s pouch in the pituitary; diffuse mucification of epithelium in vagina; presence of ultimobranchial cyst and/or presence of Ectopic thymus in the thyroid; in male and female animals from control and high dose group.

2. There was reduction in adipose tissue in rats which failed to gain weight at a normal rate. In rats, a moderate hypoplasia of the bone marrow, with displacement by adipose tissue was evident. However, the cells of the erythrocytic and myelocytic series were found in the usual proportion. The hypoplasia of the bone marrow was not diffuse and resulted in an estimated reduction of the marrow of no more than 20-30s. The vacated marrow spaces were replaced by adipose tissue, suggesting that the loss of marrow had been a slow and gradual process. In addition to the effect on bone marrow, treatment at the higher dose levels resulted in a diminution of colloid in the follicles of the thyroid and in a modest atrophy of lymphoid tissue. The diminution of the colloid content of the thyroid appeared somewhat reminiscent of thyroid stimulation.
Histopathological findings: neoplastic:
not specified
Other effects:
not specified
Details on results:
No data
Dose descriptor:
NOAEL
Remarks:
1
Effect level:
250 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
male
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food consumption and compound intake
ophthalmological examination
haematology
clinical biochemistry
urinalysis
behaviour (functional findings)
organ weights and organ / body weight ratios
gross pathology
histopathology: non-neoplastic
Remarks on result:
other: No effect observed
Dose descriptor:
NOAEL
Remarks:
1
Effect level:
> 250 mg/kg bw/day (actual dose received)
Based on:
test mat.
Sex:
female
Basis for effect level:
clinical signs
mortality
body weight and weight gain
food consumption and compound intake
ophthalmological examination
haematology
clinical biochemistry
urinalysis
behaviour (functional findings)
organ weights and organ / body weight ratios
gross pathology
histopathology: non-neoplastic
Remarks on result:
other: No effect observed
Dose descriptor:
NOAEL
Remarks:
0
Effect level:
200 other: mg/Kg
Based on:
test mat.
Sex:
female
Basis for effect level:
other: No significant effects were noted at the mentioned dose level
Dose descriptor:
NOAEL
Effect level:
100 other: mg/Kg
Based on:
test mat.
Sex:
male
Basis for effect level:
other: No significant effects were noted at the mentioned dose level
Critical effects observed:
not specified
System:
other: not specified
Organ:
not specified
Conclusions:
The No Observed Adverse Effect Level (NOAEL) of the test chemical using rats via oral route, is considered to be 200 mg/kg body weight.
Executive summary:

Data available for the test chemicals was reviewed to determine the toxic nature.The studies are as mentioned below:

In a repeated dose oral toxicity study, Sprague-Dawley male and female rats were treated with the test chemical in the concentration of 0, 250, 500 and 1000 mg/kg bw orally by gavage for 28 days. All the male and female animals from control and different dose groups up to 1000 mg/kg survived throughout the dosing period of 28 days and the recovery period of 14 days. At the end of the dosing period reduced body weight gain of 14.94% was observed in male animals from 1000 mg/kg dose group. All the male animals from control, 250, 500 and 1000 mg/kg reversal dose groups exhibited normal body weight gain throughout the dosing period of 28 days and the recovery period of 14 days. Female animals from control, 250, 500 and 1000 mg/kg dose groups exhibited normal body weight gain throughout the dosing period of 28 days. Reduced body weight gain of 11.36% was observed in female animals from 1000 mg/kg reversal group. Feed intake of animals from control and different dose groups was found to be comparable throughout the dosing period of 28 days and the recovery period of 14 days. Ophthalmoscopic examination, conducted prior to and at the end of dosing period on animals from control and different dose groups did not reveal any abnormality. No signs of toxicity were observed in male and female animals from 250 and 500 mg/kg dose groups during the dosing period of 28 days. Polyurea was observed in male and female animals from 1000 mg/kg and 1000 mg/kg reversal dose groups during the dosing period of 28 days and the recovery period of 14 days. Similarly, Detailed clinical observations conducted at weekly interval (upto 6thweek) did not reveal any abnormality in all male and female animals from control and different dose groups during the dosing period of 28 days and the recovery period of 14 days. Towards the end of the exposure period in week 4, functional observation battery such as sensory reactivity to stimuli of different types (e.g. auditory, visual and proprioceptive stimuli), Grip strength and Motor activity values revealed no abnormalities attributable to the treatment. No statistically significant changes in the values of various Haematological parameters were at the end of the dosing period on day 29. At the end of the recovery period on day 43, statistically significant decrease in the values of Total RBC and HCT at 1000 mg/kg in male. Statistically significant decrease in the values of Hb and HCT at 500 mg/kg and Total RBC at 250 and 500 mg/kg were observed in female rat. At the end of the recovery period on day 43, no statistically significant changes in the values of various parameters in female rats. The decrease in the values of various parameters was marginal and within the normal biological and laboratory limits. Clinical biochemistry analysis at the end of the dosing period on day 29, revealed statistically significant increase in the values of Globulin at 250 mg/kg in male rat. In addition statistically significant decrease was observed in the values of BUN and Urea Nitrogen at 250 mg/kg in male, BUN and Urea Nitrogen at 500 mg/kg in female rat, Alanine Aminotransferase at 250 and 500 mg/kg in female and Alkaline Phosphatase at 250 mg/kg in female rat. At the end of the recovery period on day 43 (Reversal groups) statistically significant increase was observed in the values of Bile Acid at 1000 mg/kg in female and statistically significant decrease was observed in the values of Calcium at 1000 mg/kg in male. The increase/decrease in the values of various parameters was marginal and within the normal biological and laboratory limits. Urine analysis conducted on male animals during 4thand 6thweek of study period (on day 23, 24 and 43), revealed no abnormality attributable to the treatment. Urine analysis conducted on female animals during 4thweek of study period (on day 24 and 26), higher volume of urine was observed in female at 1000 mg/kg dose group. During 6th week of study period (on day 43) revealed no abnormality attributable to the treatment. At termination of dosing on day 29, male animals from 1000 mg/kg dose group revealed increased relative weights of brain. In addition, increased relative weights of liver and kidneys were observed in male at 500 and 1000 mg/kg dose groups when compared with that of controls. Increased relative weights of spleen were observed in male at 250 and 1000 mg/kg as compared to controls. Increased relative weights of thymus were observed in male at 250 mg/kg as compared to controls. No effect on male organ weight sacrificed on day 43 from 1000 mg/kg as comparable to controls. At termination of dosing on day 29, at 1000 mg/kg increased relative weights of liver were observed in female as compared to controls. In addition, increased relative weights of kidneys were observed in female at 500 and 1000 mg/kg as compared to controls. At 500 mg/kg dose decreased relative weights of heart in female rats as compared to controls. Organ weight data of female animals sacrificed on day 43 at 1000 mg/kg increased relative weights of brain, liver, kidneys, spleen and uterus were observed in female as compared to controls. Although significant changes in the values of organ weight of brain, liver, spleen, thymus, heart and uterus were observed in male and female animals at 250 mg/kg and 500 mg/kg dose groups, no related gross pathological and histopathological findings were seen, hence these findings were considered to be of no toxicological importance. Gross pathological examination did not reveal any abnormality attributable to the treatment. Histopathological examination revealed focal to diffuse, minimal to moderate tubular dilatation in kidneys in male (1/6) and female (6/6) animals from 1000 mg/kg dose group, in male (1/6) and female (6/6) animals from 500 mg/kg dose group and in male (0/6) and female (5/6) animals from 250 mg/kg dose group and in female (1/6) from 1000 mg/kg reversal dose group. Focal to multifocal, minimal mononuclear cells infiltration was observed in kidneys in male (1/6) and female (3/6) animals from 1000 mg/kg dose group and in female animals from 250 mg/kg (2/6) and 500 mg/kg (2/6) dose groups. All histopathological the changes observed in male animals were reversible during the recovery period of 14 days. The histopathological changes observed in female animals were evident in reversal group one animal, during the recovery period of 14 days. Therefore, No Observed Adverse Effect Level (NOAEL) of the test chemcal in the Sprague Dawley rat via oral route, over a period of 28 days was found to be 250 mg/kg body weight in male animals and less than 250 mg/kg body weight in female animals.

In another study, repeated dose oral toxicity study was performed to determine the toxic nature of the test chemical. The study was performed using male and female rats. The test chemical was mixed with 2% carboxymethylcellulose and used at dose level of 0, 50 (males), 100 (females), 200 or 400 mg/Kg for 180 days. During the study period, the animals were observed for clinical signs, mortality, body weight changes, changes in hematology and biochemical parameters and subjected to gross and histopathology. No untoward signs of toxicity and no mortality were noted in the treated animals. A daily dose of 200 mg/kg or less caused no depression in rate of weight gain in female rats, and even at the 400 mg/kg level the difference in the weight curves between the controls and treated animals was not striking. In contrast, in male rats, dose levels of 200 mg/kg or more produced some inanition and significantly affected the rate of weight gain. There were no striking biochemical changes that could be attributed to drug treatment in any of the rats. Organ weights in treated animals did not differ from those of controls. Gross examination revealed no relevant changes in any of the organs. There was reduction in adipose tissue in rats which failed to gain weight at a normal rate. In rats, a moderate hypoplasia of the bone marrow, with displacement by adipose tissue was evident. However, the cells of the erythrocytic and myelocytic series were found in the usual proportion. The hypoplasia of the bone marrow was not diffuse and resulted in an estimated reduction of the marrow of no more than 20-30s. The vacated marrow spaces were replaced by adipose tissue, suggesting that the loss of marrow had been a slow and gradual process. In addition to the effect on bone marrow, treatment at the higher dose levels resulted in a diminution of colloid in the follicles of the thyroid and in a modest atrophy of lymphoid tissue. The diminution of the colloid content of the thyroid appeared somewhat reminiscent of thyroid stimulation. Based on the observations made, the No observed adverse effect level (NOAEL) for the test chemical is considered to be 100 mg/Kg for males and 200 mg/Kg for females in a chronic toxicity study.

Based on the data available, The No Observed Adverse Effect Level (NOAEL) of the test chemical 1H-benzimidazole using rats via oral route, is considered to be 200 mg/kg body weight.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
200 mg/kg bw/day
Study duration:
chronic
Species:
rat
Quality of whole database:
Data is from peer reviewed publication

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available
Quality of whole database:
Waiver

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion
Endpoint conclusion:
no study available
Quality of whole database:
Waiver

Repeated dose toxicity: dermal - local effects

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Data available for the test chemicals was reviewed to determine the toxic nature of 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione (37052-78-1) . The studies are as mentioned below:

Repeated dose toxicity: Oral

In a repeated dose oral toxicity study, Sprague-Dawley male and female rats were treated with the test chemical in the concentration of 0, 250, 500 and 1000 mg/kg bw orally by gavage for 28 days. All the male and female animals from control and different dose groups up to 1000 mg/kg survived throughout the dosing period of 28 days and the recovery period of 14 days. At the end of the dosing period reduced body weight gain of 14.94% was observed in male animals from 1000 mg/kg dose group. All the male animals from control, 250, 500 and 1000 mg/kg reversal dose groups exhibited normal body weight gain throughout the dosing period of 28 days and the recovery period of 14 days. Female animals from control, 250, 500 and 1000 mg/kg dose groups exhibited normal body weight gain throughout the dosing period of 28 days. Reduced body weight gain of 11.36% was observed in female animals from 1000 mg/kg reversal group. Feed intake of animals from control and different dose groups was found to be comparable throughout the dosing period of 28 days and the recovery period of 14 days. Ophthalmoscopic examination, conducted prior to and at the end of dosing period on animals from control and different dose groups did not reveal any abnormality. No signs of toxicity were observed in male and female animals from 250 and 500 mg/kg dose groups during the dosing period of 28 days. Polyurea was observed in male and female animals from 1000 mg/kg and 1000 mg/kg reversal dose groups during the dosing period of 28 days and the recovery period of 14 days. Similarly, Detailed clinical observations conducted at weekly interval (upto 6thweek) did not reveal any abnormality in all male and female animals from control and different dose groups during the dosing period of 28 days and the recovery period of 14 days. Towards the end of the exposure period in week 4, functional observation battery such as sensory reactivity to stimuli of different types (e.g. auditory, visual and proprioceptive stimuli), Grip strength and Motor activity values revealed no abnormalities attributable to the treatment. No statistically significant changes in the values of various Haematological parameters were at the end of the dosing period on day 29. At the end of the recovery period on day 43, statistically significant decrease in the values of Total RBC and HCT at 1000 mg/kg in male. Statistically significant decrease in the values of Hb and HCT at 500 mg/kg and Total RBC at 250 and 500 mg/kg were observed in female rat. At the end of the recovery period on day 43, no statistically significant changes in the values of various parameters in female rats. The decrease in the values of various parameters was marginal and within the normal biological and laboratory limits. Clinical biochemistry analysis at the end of the dosing period on day 29, revealed statistically significant increase in the values of Globulin at 250 mg/kg in male rat. In addition statistically significant decrease was observed in the values of BUN and Urea Nitrogen at 250 mg/kg in male, BUN and Urea Nitrogen at 500 mg/kg in female rat, Alanine Aminotransferase at 250 and 500 mg/kg in female and Alkaline Phosphatase at 250 mg/kg in female rat. At the end of the recovery period on day 43 (Reversal groups) statistically significant increase was observed in the values of Bile Acid at 1000 mg/kg in female and statistically significant decrease was observed in the values of Calcium at 1000 mg/kg in male. The increase/decrease in the values of various parameters was marginal and within the normal biological and laboratory limits. Urine analysis conducted on male animals during 4thand 6thweek of study period (on day 23, 24 and 43), revealed no abnormality attributable to the treatment. Urine analysis conducted on female animals during 4thweek of study period (on day 24 and 26), higher volume of urine was observed in female at 1000 mg/kg dose group. During 6th week of study period (on day 43) revealed no abnormality attributable to the treatment. At termination of dosing on day 29, male animals from 1000 mg/kg dose group revealed increased relative weights of brain. In addition, increased relative weights of liver and kidneys were observed in male at 500 and 1000 mg/kg dose groups when compared with that of controls. Increased relative weights of spleen were observed in male at 250 and 1000 mg/kg as compared to controls. Increased relative weights of thymus were observed in male at 250 mg/kg as compared to controls. No effect on male organ weight sacrificed on day 43 from 1000 mg/kg as comparable to controls. At termination of dosing on day 29, at 1000 mg/kg increased relative weights of liver were observed in female as compared to controls. In addition, increased relative weights of kidneys were observed in female at 500 and 1000 mg/kg as compared to controls. At 500 mg/kg dose decreased relative weights of heart in female rats as compared to controls. Organ weight data of female animals sacrificed on day 43 at 1000 mg/kg increased relative weights of brain, liver, kidneys, spleen and uterus were observed in female as compared to controls. Although significant changes in the values of organ weight of brain, liver, spleen, thymus, heart and uterus were observed in male and female animals at 250 mg/kg and 500 mg/kg dose groups, no related gross pathological and histopathological findings were seen, hence these findings were considered to be of no toxicological importance. Gross pathological examination did not reveal any abnormality attributable to the treatment. Histopathological examination revealed focal to diffuse, minimal to moderate tubular dilatation in kidneys in male (1/6) and female (6/6) animals from 1000 mg/kg dose group, in male (1/6) and female (6/6) animals from 500 mg/kg dose group and in male (0/6) and female (5/6) animals from 250 mg/kg dose group and in female (1/6) from 1000 mg/kg reversal dose group. Focal to multifocal, minimal mononuclear cells infiltration was observed in kidneys in male (1/6) and female (3/6) animals from 1000 mg/kg dose group and in female animals from 250 mg/kg (2/6) and 500 mg/kg (2/6) dose groups. All histopathological the changes observed in male animals were reversible during the recovery period of 14 days. The histopathological changes observed in female animals were evident in reversal group one animal, during the recovery period of 14 days. Therefore, No Observed Adverse Effect Level (NOAEL) of the test chemcal in the Sprague Dawley rat via oral route, over a period of 28 days was found to be 250 mg/kg body weight in male animals and less than 250 mg/kg body weight in female animals.

 

In another study, repeated dose oral toxicity study was performed to determine the toxic nature of the test chemical. The study was performed using male and female rats. The test chemical was mixed with 2% carboxymethylcellulose and used at dose level of 0, 50 (males), 100 (females), 200 or 400 mg/Kg for 180 days. During the study period, the animals were observed for clinical signs, mortality, body weight changes, changes in hematology and biochemical parameters and subjected to gross and histopathology. No untoward signs of toxicity and no mortality were noted in the treated animals. A daily dose of 200 mg/kg or less caused no depression in rate of weight gain in female rats, and even at the 400 mg/kg level the difference in the weight curves between the controls and treated animals was not striking. In contrast, in male rats, dose levels of 200 mg/kg or more produced some inanition and significantly affected the rate of weight gain. There were no striking biochemical changes that could be attributed to drug treatment in any of the rats. Organ weights in treated animals did not differ from those of controls. Gross examination revealed no relevant changes in any of the organs. There was reduction in adipose tissue in rats which failed to gain weight at a normal rate. In rats, a moderate hypoplasia of the bone marrow, with displacement by adipose tissue was evident. However, the cells of the erythrocytic and myelocytic series were found in the usual proportion. The hypoplasia of the bone marrow was not diffuse and resulted in an estimated reduction of the marrow of no more than 20-30s. The vacated marrow spaces were replaced by adipose tissue, suggesting that the loss of marrow had been a slow and gradual process. In addition to the effect on bone marrow, treatment at the higher dose levels resulted in a diminution of colloid in the follicles of the thyroid and in a modest atrophy of lymphoid tissue. The diminution of the colloid content of the thyroid appeared somewhat reminiscent of thyroid stimulation. Based on the observations made, the No observed adverse effect level (NOAEL) for the test chemical is considered to be 100 mg/Kg for males and 200 mg/Kg for females in a chronic toxicity study.

Based on the data available, The No Observed Adverse Effect Level (NOAEL) of the test chemical using rats via oral route, is considered to be 200 mg/kg body weight.

Repeated inhalation study:

According to Annex IX of the REACH regulation, testing by the inhalation route is appropriate only if exposure of humans via inhalation is likely. Taking into account the low vapour pressure of the substance 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione (37052-78-1) which is reported as 5.45E-6mmHg at 25 C. Thus, exposure to inhalable dust, mist and vapour of the 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione is highly unlikely. Therefore this study is considered for waiver.

Repeated dermal study;

The acute toxicity value for 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione (37052-78-1) (as provided in section 7.2.3) is >2000 mg/kg body weight. Also, given the use of the chemical; repeated exposure by the dermal route is unlikely since the use of gloves is common practice in industries. Thus, it is expected that 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione shall not exhibit 28 day repeated dose toxicity by the dermal route. In addition, there is no data available that suggests that 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione shall exhibit repeated dose toxicity by the dermal route. Hence this end point was considered for waiver.

Based on the data available and applying the weight of evidence approach, the test chemical 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione (37052-78-1) is not likely to be toxic upon repeated exposure by oral route as per the criteria mentioned in CLP regulation.

Justification for classification or non-classification

Based on the data available and applying the weight of evidence approach, the test chemical 5-methoxy-1,3-dihydro-2H-benzimidazole-2-thione (37052-78-1) is not likely to be toxic upon repeated exposure by oral route as per the criteria mentioned in CLP regulation.