Registration Dossier

Administrative data

Description of key information

Repeated dose toxicity: Oral

In a repeated dose toxicity study, male and female Wistar rats were exposed to the test chemical orally in the concentration of 0, 125, 250 and 500 mg/kg/day body weight. No substance related toxic changes were obsrved in survival, body weight, food consumption,hematology, clinical chemistry,battery test and urinalysis of treated rats. Significant change in liver, kidney,Adrenals and spleen of 250 and 500 mg/kg bw/day treated rats as compared to control. In addition, no gross pathological and histopathological changes were observed in male and female rats. Some changes were observed but they are not test substance related.Therefore, No observed adverse effect level (NOAEL) was considered to be 500 mg/kg/day body weight when rats are exposed to the test chemical orally for 28 days. 

Repeated dose toxicity: inhalation

This end point was considered for waiver since 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 particle size distribution of N-tert-butylacrylamide and that the exposure via the inhalation route is unlikely, this end point was considered for waiver.

 

Repeated dose toxicity: dermal

The acute toxicity value for N-tert-butylacrylamide (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 N-tert-butylacrylamide shall not exhibit 28 day repeated dose toxicity by the dermal route. In addition, there is no data available that suggests that N-tert-butylacrylamide 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:
experimental study
Adequacy of study:
weight of evidence
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Experimental result using OECD recommended guidelines
Justification for type of information:
Data is from study report
Qualifier:
according to
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity in Rodents)
Principles of method if other than guideline:
Repeated Dose 28 days Study of the test chemical in rats.
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Wistar
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: In-House Bred at sa-FORD, Animal Facility
- Age at study initiation: 6 - 8 weeks
- Weight at study initiation: Male : 137-191 g, Female: 137- 165 g
- Fasting period before study: No data available
- Housing: Animals were housed in Polycarbonate cages. Cage rotation was carried out weekly and cleaned at regular intervals. Animals were bedded on sterilized corn cob produced from pure corn, dried and free from dust. Floor of the experimental room and work tops were swept and mopped with disinfectant solution every day or as on requirement.
- Diet (e.g. ad libitum): A conventional laboratory pelleted diet, ad libitum.
- Water (e.g. ad libitum): Aqua guard filtered drinking water in bottles, ad libitum.
- Acclimation period: Male: 6 days, Female: 7 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19.00 to 23.10 °C
- Humidity (%): 49.90 to 69.40%.
- Air changes (per hr): Adequately filtered air 12 times per hour
- Photoperiod (hrs dark / hrs light): 12 hours light and 12 hours dark artificial light.

IN-LIFE DATES: From: September 22, 2014
To: October 20, 2014
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
PREPARATION OF DOSING SOLUTIONS: Substance was weighed and dissolved in a vehicle (corn oil) to achieve desired concentration of test item. Dose formulation was freshly prepared daily. At the time of dosing, dose formulation was kept on the magnetic stirrer to maintain the homogeneity of test item.

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

VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil was used as a vehicle based on the solubility testing.
- Concentration in vehicle: 0,125,250 and 500 mg/kg/day
- Amount of vehicle (if gavage): 1 ml/100g body weight
- Lot/batch no. (if required): MKBQ9948V and MKBG9426V
- Purity: No data available
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Stability and concentration of the substance in dose formulation was analysed by a validated analytical method, at the start of treatment (on day 1) and on day 21.
Duration of treatment / exposure:
28 days
Frequency of treatment:
Daily
Dose / conc.:
0 other: mg/kg/day
Dose / conc.:
125 other: mg/kg/day
Dose / conc.:
250 other: mg/kg/day
Dose / conc.:
500 other: mg/kg/day
No. of animals per sex per dose:
Total : 60
0 mg/kg/day: 5 male, 5 female
125 mg/kg/day: 5 male, 5 female
250 mg/kg/day: 5 male, 5 female
500 mg/kg/day: 5 male, 5 female

Recovery:
0 mg/kg/day: 5 male, 5 female
500 mg/kg/day: 5 male, 5 female
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: The dose levels were selected based on the information provided by Sponsor.
- Rationale for animal assignment (if not random): No data available
- Rationale for selecting satellite groups: No data available
- Post-exposure recovery period in satellite groups: No data available
- Section schedule rationale (if not random): No data available
Observations and examinations performed and frequency:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Twice daily (morning and evening)
- Cage side observations checked in table [No.?] were included: Morbidity and mortality were observed.

DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Once before the first treatment and weekly thereafter.

BODY WEIGHT: Yes
- Time schedule for examinations: At start of treatment and thereafter weekly.

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: No data available
- Compound intake calculated as time-weighted averages from the consumption and body weight gain data: No data available

FOOD EFFICIENCY: No data available
- 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 available

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

OPHTHALMOSCOPIC EXAMINATION: Yes
- Time schedule for examinations: Prior to treatment (During Acclimatization Period) and at the end of the dosing (main groups) and recovery periods (recovery group) were examined.
- Dose groups that were examined: All 60 animals were examined.

HAEMATOLOGY: Yes
- Time schedule for collection of blood: On termination day, just prior to necropsy.
- Anaesthetic used for blood collection: No data available
- Animals fasted: Yes, overnight (approximately 16-18 hr).
- How many animals: All 60 animals were examined.
- Parameters checked in table [No.?] were examined: Total Erythrocyte Count (RBC), Hematocrit (HCT), Mean Corpuscular Volume (MCV), Hemoglobin (HGB), Mean Corpuscular Hemoglobin (MCH), Mean Corpuscular Hemoglobin Concentration (MCHC), Platelet Count (PLT), Total Leukocyte count (WBC), Prothombin Time (PT) and Activated Partial Thromboplastin time (aPTT) were examined.

CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: On termination day, just prior to necropsy.
- Animals fasted: Yes, overnight (approximately 16-18 hr).
- How many animals: All 60 animals were examined.
- Parameters checked in table [No.?] were examined: Glucose (Glu), Cholesterol (Chol), Triglycerides (TRIG), Alanine amino transferase (ALT), Aspartate amino transferase (AST), Gamma-Glutamyl Transpeptidase (GGT), Calcium, Creatine Kinase (CK), Albumin, Total Protein (TP), Creatinine (Crea), Total Bilirubin (T.Bil), Phosphorus, Alkaline phosphatase (ALP), Urea, Lactate Dehydrogenase (LD), Sodium (Na), Potassium (K), Chloride (Cl), Blood urea nitrogen (BUN), Globulin (Glob), A/G and Bile acids were examined.

URINALYSIS: Yes
- Time schedule for collection of urine: No data available
- Metabolism cages used for collection of urine: Yes
- Animals fasted: No data available
- Parameters checked in table [No.?] were examined: Colour, appearance, urine volume, Blood / Blood Cell, Bilirubin, Urobilinogen, Ketone, Protein, Nitrite, Glucose, pH, Specific Gravity, Leucocytes, Microscopical Parameters were examined.

NEUROBEHAVIOURAL EXAMINATION: No data available
- Time schedule for examinations: No data available
- Dose groups that were examined: No data available
- Battery of functions tested: sensory activity / grip strength / motor activity / other: Yes, Sensory reactivity to stimuli, grip strength, hind limb foot splay and motor activity were tested.

OTHER:
Organ weight: Absolute and relative organ weights were examined.

Organ examined: Liver, kidneys, adrenals, testes, epdidymides, Prostate and Seminal vesicle with coagulating glands, uterus with cervix, ovary with oviduct, thymus, spleen, brain and heart were examined.
Bone Marrow Smear were also examined
Sacrifice and pathology:
GROSS PATHOLOGY: Yes
All animals were fasted overnight before necropsy. Animals were sacrificed by using over dose of CO2 and examined externally and macroscopically.

HISTOPATHOLOGY: Yes
All tissues were preserved in 10 % neutral buffered formalin (except eyes and testes; which were fixed using Modified Davidson fluid for 24 hr and then transferred to 10 % neutral buffered formalin (NBF) for preservation) for subsequent histopathological examintion.

Organ examined:
Adrenals, Bone (femur) with joint, Brain (cerebrum,cerebellum,mid brain), Cecum, Colon, Duodenum, Epididymides, Eyes with optic nerve, Gross lesion, Ileum, Jejunum, Kidneys, Liver, Lung, Mammary glands, Mesenteric and Mandibular lymph node, Oesophagus and Ovary with oviduct were examined.
Other examinations:
No data available
Statistics:
Statistical analysis of raw data was performed by using statistical software Sigma Plot 11.0. The mean and standard deviation was calculated using the software and all data was summarized in tabular form. All continuous data (body weight, feed consumption, haematology, clinical chemistry, absolute and relative organ weights) were checked for their homogeneity using Bartlett’s test. All homogenous data was analysed using ANOVA and data showing significance in their variances was subjected to Dunnett’s t-test. All heterogeneous data was analysed using F test and Student’s t-test.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
no effects observed
Food consumption and compound intake (if feeding study):
no effects observed
Food efficiency:
not specified
Water consumption and compound intake (if drinking water study):
not specified
Ophthalmological findings:
no effects observed
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
no effects observed
Behaviour (functional findings):
not specified
Immunological findings:
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Neuropathological findings:
not specified
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
not specified
Details on results:
Mortality:
No mortality and morbidity was observed in treated male and female rat as compare to control.

Clinical signs:
No clinical sign were observed in treatment and recovery dosed male and female rat as compare to control.

Body weight and weight gain: When treated with 500 mg/kg/day body weight, in female rat statistically significant decreased in body weight gain were observed as compare to control.

No change was observed in 125 and 250 mg/kg/day body weight treated male and female rats as compare to control.

Food consumption and compound intake: No change was observed in food consumption of treated male and female rat in treatment and recovery period as compare to control.

Food efficiency: No data available

Water consumption and compound intake: No data available

Opthalmoscopic examination: No abnormalities were observed in treated male and female rat in treatment and recovery period as compare to control.

Haematology: When treated with 500 mg/kg/day body weight, in female rat statistically significant increase neutrophil were observed.

In recovery, statistically significant decrease was observed in MCV and WBC and increase observed in PLT in females and statistically significant decrease in APTT in male rat as compare to control.

Above changes were not related to the test substence and may due to the preanalytical and analytical variables.

Clinical chemistry: When treated with 125, 250 and 500 mg/kg/day body weight, in male rat statistically significant increase were observed in Sodium (Na) and decrease in Lactate Dehydrogenase (LD) and Creatine Kinase (CK) when treated with 125 and 500 mg/kg/day body weight.

When treated with 250 and 500 mg/kg/day body weight, Statistically significant increase was observed in Alkaline phosphatase (ALP) in male rat.

When treated with 500 mg/kg/day body weight, Alanine amino transferase (ALT) increased in male rat as compare to control.

In recovery, Statistically significant increase were observed in Blood urea nitrogen (BUN), Urea, Albumin (ALB), Alkaline phosphatase (ALP), Potassium (K), Chloride (Cl) and Bile acids and statistically significant decrease in Cholesterol (Chol) in male rat and Sodium (Na) in female rat as compare to control.

Urinanalysis: No change was observed in urine analysis of treated animals in treatment and recovery period as compare to control.

Neurobehaviour: No data available

Organ weights: In male rat, relative weight of liver were significantly increase in 250 and 500 mg/kg/day body weight.

Relative organ weight of kidney showed statistically significant increase when treated with in 250 and 500 mg/kg/day body weight.

Absolute and relative organ weight of S.V. with coagulating gland and prostrate as whole was decreased in 250 mg/kg/day body weight in male rat as compare to control.

In female absolute weight of liver showed significantly decrease in 125 mg/kg/day body weight.

In recovery, in female absolute and relative organ weight of Adrenals was increased and absolute organ weight of kidney was decreased when treated with 500 mg/kg/day body weight and absolute and relative organ weight of spleen was decreased in 250 mg/kg/day body weight.

In male, significant difference were observed in absolute & relative weight of liver in male rat when treated with 250 and 500 mg/kg/day body weight.

Gross pathology: No external and internal abnormal changes were observed in treated male and female rats in treatment and recovery period as compare to control.

Histopathology: When treaed with 500 mg/kg/day body weight, hyperplasia of BALT in Lung and MNC infiltration in trachea were observed in male and female this effect was reversed in 14 days recovery period.

These observed changes are inflammatory changes and are the most common observed changes in rodents.
Dose descriptor:
NOAEL
Effect level:
500 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Effect on survival, body weight, food consumption, battery test, hematology, clinical chemistry, organ weight, gross pathology and histopathology.
Critical effects observed:
not specified

Mortality and Morbidity

Group

Treatment

Dose (mg/kg b.wt.)

No. of Animals/

Group

Day of Observations

Observation

G1

Control

0

5

1-29

NMM

G2

Low

125

5

1-29

NMM

G3

Mid

250

5

1-29

NMM

G4

High

500

5

1-29

NMM

G1-R

Control -Recovery

0

5

1-43

NMM

G4-R

High- Recovery

500

5

1-43

NMM

Group

Treatment

Dose (mg/kg b.wt.)

No. of Animals/

Group

Day of Observations

Observation

G1

Control

0

5

1-29

NMM

G2

Low

125

5

1-29

NMM

G3

Mid

250

5

1-29

NMM

G4

High

500

5

1-29

NMM

G1-R

Control -Recovery

0

5

1-43

NMM

G4-R

High- Recovery

500

5

1-43

NMM

Key:NMM = No mortality and morbidity observed.

Mean Absolute Organ Weight (g)

Sex:Male

Organs

 

Group 1

Group 2

Group 3

Group 4

Mean

SD

Mean

SD

Mean

SD

Mean

SD

Body weight (g)

251.80

21.09

239.20

24.78

240.60

18.37

234.60

11.48

Brain

1.910

0.062

1.935

0.053

1.966

0.051

1.900

0.057

Adrenals

0.046

0.013

0.059

0.007

0.053

0.009

0.051

0.010

S.V. With Coagulating gland and prostrate

1.1831

0.22882

1.11382

0.167243

0.8658

0.1214

1.1246

0.12127

Testes

3.003

0.246

2.857

0.296

2.966

0.167

2.810

0.150

Epdidymides

0.902

0.164

1.004

0.135

0.958

0.106

0.954

0.045

Heart

1.044

0.088

1.007

0.078

0.998

0.069

0.911

0.043

Liver

10.450

0.919

11.804

1.553

13.062

1.580

13.492

1.428

Kidneys

2.294

0.219

2.271

0.261

2.575

0.202

2.427

0.192

Spleen

1.356

0.434

1.142

0.332

1.136

0.196

0.905

0.198

Thymus

0.301

0.071

0.311

0.044

0.299

0.071

0.241

0.056

Organs

 

Group 1-R

Group 4-R

Mean

SD

Mean

SD

Body weight (g)

273.40

20.21

280.80

15.39

Brain

1.935

0.120

1.927

0.067

Adrenals

0.056

0.009

0.064

0.008

S.V. With Coagulating gland and prostrate

1.5541

0.4754

1.4925

0.260007

Testes

3.210

0.166

3.168

0.254

Epdidymides

1.145

0.063

1.112

0.153

Heart

1.148

0.080

1.175

0.125

Liver

10.861

0.878

10.788

0.658

Kidneys

2.417

0.304

2.368

0.199

Spleen

1.002

0.106

1.048

0.285

Thymus

0.296

0.106

0.349

0.090

Sex:Female

Organs

Group 1

Group 2

Group 3

Group 4

Mean

SD

Mean

SD

Mean

SD

Mean

SD

Body weight (g)

185.40

15.13

173.00

4.00

178.40

7.37

169.60

5.41

Brain

1.791

0.076

1.692

0.091

1.728

0.040

1.789

0.087

Adrenals

0.059

0.011

0.051

0.011

0.061

0.011

0.064

0.012

Ovary

0.092

0.012

0.106

0.032

0.111

0.024

0.091

0.015

Uterus

0.720

0.120

0.645

0.107

0.561

0.152

0.553

0.115

Heart

0.853

0.118

0.725

0.027

0.731

0.063

0.725

0.079

Liver

8.231

0.754

7.106

0.640

7.743

0.646

8.403

0.586

Kidneys

1.582

0.132

1.412

0.098

1.470

0.097

1.585

0.177

Spleen

0.719

0.279

0.502

0.122

0.485

0.120

0.516

0.225

Thymus

0.300

0.049

0.266

0.081

0.235

0.036

0.242

0.042

Organs

Group 1-R

Group 4-R

Mean

SD

Mean

SD

Body weight (g)

196.60

9.96

188.40

8.50

Brain

1.792

0.087

1.800

0.059

Adrenals

0.077

0.007

0.090

0.006

Ovary

0.164

0.015

0.170

0.018

Uterus

0.777

0.305

0.723

0.120

Heart

0.846

0.054

0.782

0.045

Liver

7.429

0.677

7.394

0.560

Kidneys

1.684

0.058

1.522

0.110

Spleen

0.776

0.126

0.441

0.054

Thymus

0.281

0.062

0.320

0.037

Conclusions:
The No observed adverse effect level (NOAEL) was considered to be 500 mg/kg/day body weight when male and female rat were exposed to the test chemical.
Executive summary:

In a repeated dose toxicity study, male and female Wistar rats were exposed to the test chemical orally in the concentration of 0, 125, 250 and 500 mg/kg/day body weight. No substance related toxic changes were obsrved in survival, body weight, food consumption,hematology, clinical chemistry,battery test and urinalysis of treated rats. Significant change in liver, kidney,Adrenals and spleen of 250 and 500 mg/kg bw/day treated rats as compared to control. In addition, no gross pathological and histopathological changes were observed in male and female rats. Some changes were observed but they are not test substance related.Therefore, No observed adverse effect level (NOAEL) was considered to be 500 mg/kg/day body weight when rats are exposed to the test chemical orally for 28 days. 

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
500 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
K1 level testing data from study report.

Repeated dose toxicity: inhalation - systemic effects

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: inhalation
Data waiving:
exposure considerations
Justification for data waiving:
other:
Critical effects observed:
not specified
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

Link to relevant study records
Reference
Endpoint:
repeated dose toxicity: dermal
Data waiving:
other justification
Justification for data waiving:
other:
Critical effects observed:
not specified
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 chemical was reviewed to determine the toxic nature of the test chemical. The studies are as mentioned below:

Repeated dose toxicity: oral

In a repeated dose toxicity study, male and female Wistar rats were exposed to the test chemical orally in the concentration of 0, 125, 250 and 500 mg/kg/day body weight. No substance related toxic changes were obsrved in survival, body weight, food consumption,hematology, clinical chemistry,battery test and urinalysis of treated rats. Significant change in liver, kidney,Adrenals and spleen of 250 and 500 mg/kg bw/day treated rats as compared to control. In addition, no gross pathological and histopathological changes were observed in male and female rats. Some changes were observed but they are not test substance related.Therefore, No observed adverse effect level (NOAEL) was considered to be 500 mg/kg/day body weight when rats are exposed to the test chemical orally for 28 days. 

 

In another study, repeated dose oral toxicity study was performed, Wistar male rats was treated with the test chemical orally by using drinking water in the concentration of 1907.85 mg/kg/day. Effect on body weight gain was observed in treated rats. No significant effect on clinical sign and Rotarod Performance were observed in treated rats as compared to control. In addition, no significant effect on histopa thology of both tibial and sural nerves and (3H] Colchicine-binding were observed in treated rats as compared to control. Therefore, No observed adverse effect level (NOAEL) was considered to be 1907.85 mg/kg/day when Wistar male rats were treated with the test chemical orally for 90 days.

 

Moreover in the same study by Hashimoto et.al (1981), ddY male mice was treated with the test chemical orally in the concentration of 317.975 mg/kg bw in DMSO for 10 weeks. No significant effect on body weight, testicular weights and neurotoxicity of treated mice were observed as compared to control. Therefore, no observed adverse effect level (NOAEL) was considered to be 317.975 mg/kg bw when ddY male mice were treated with the test chemical orally for 10 weeks.

 

Based on the above mentioned 28 day and 90-day repeated-dose study conducted in experimental animals, no significant toxic effects were seen to occur. The observed effects are gender specific and not species specific and so one cannot arrive to a correct conclusion that the effect is sever and will effect human in long run. If the effects observed in animals are not considered relevant for humans then these should not be used to support classification. Hence, based on the above studies summarized with oral routes it can be observed that NOAEL value of 500 mg/Kg bw/ d in rats should be considered for classification inference.

 

Repeated dose toxicity: inhalation

This end point was considered for waiver since 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 particle size distribution of N-tert-butylacrylamide and that the exposure via the inhalation route is unlikely, this end point was considered for waiver.

 

Repeated dose toxicity: dermal

The acute toxicity value for N-tert-butylacrylamide (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 N-tert-butylacrylamide shall not exhibit 28 day repeated dose toxicity by the dermal route. In addition, there is no data available that suggests that N-tert-butylacrylamide 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 is not likely to be toxic upon repeated dose exposure by oral route. Hence the test chemical is not likely to be toxic 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 is not likely to be toxic upon repeated dose exposure by oral route. Hence the test chemical is not likely to be toxic as per the criteria mentioned in CLP regulation.