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Toxicological information

Repeated dose toxicity: oral

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Administrative data

Endpoint:
short-term repeated dose toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report date:
2014

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 407 (Repeated Dose 28-Day Oral Toxicity Study in Rodents)
Principles of method if other than guideline:
Repeated Dose 28-day Oral Toxicity Study of Benzyl Propionate in the Rat.
GLP compliance:
yes

Test material

Constituent 1
Chemical structure
Reference substance name:
Benzyl propionate
EC Number:
204-559-3
EC Name:
Benzyl propionate
Cas Number:
122-63-4
Molecular formula:
C10H12O2
IUPAC Name:
benzyl propionate
Details on test material:
- Name of test material (as cited in study report):Benzyl Propionate - Molecular formula (if other than submission substance):C10H12O2 - Molecular weight (if other than submission substance):164.2 g/mole- Substance type:Organic- Physical state:Liquid

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
Details on test animal
TEST ANIMALS
- Source: National Institute of Biosciences, Pune.

- Age at study initiation: 6 to 8 weeks old

- Weight at study initiation:
Male 169.50 gg
Female 152.35 g

- Fasting period before study: No data available

- Housing: Animals were housed in polycarbonate cages. Three rats of same sex were housed together in each cage of size 39 cm X 28 cm X 14 cm. Paddy husk was used as bedding material.

- Diet (e.g. ad libitum): Rodent feed supplied by the Nutrivet Life Sciences, Pune, was provided ad libitum from individual feeders on cage top.

- 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 prior to dosing.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3°C (actual range: 19.5 °C to 22.8 °C)

- Humidity (%):30% to 70% (actual range: 53.3% to 58.1%).

- Air changes (per hr): Ten air changes per hour of 100% fresh air that has been passed through the HEPA filters.

- Photoperiod (hrs dark / hrs light): An artificial light and dark cycle of 12


Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
Details on oral exposure
PREPARATION OF DOSING SOLUTIONS: Benzyl Propionate was diluted with Corn oil for
preparation of dosing solution(s).

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): No data available

- Concentration in vehicle:
The solution(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 bw/day.

- Amount of vehicle (if gavage):
0.00 mg/ml/day, 26.60 mg/ml/day, 52.38 mg/ml/ day and 105.27 mg/ml/day.

- Lot/batch no. (if required): No data available

- Purity: No data available
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis for concentration and stability of Benzyl Propionate were conducted at Subcontracted Laboratory. Test item formulation samples prepared day 1 (pre-dosing) were sent to Subcontracted Laboratory.
Duration of treatment / exposure:
28 days consecutively
Frequency of treatment:
Once daily
Doses / concentrations
Remarks:
Doses / Concentrations:
0 (vehicle), 250, 500 and 1000 mg/kg bw/day.
Basis:
actual ingested
No. of animals per sex per dose:
Control: 6 male, 6 female
250 mg/kg bw/day: 6 male, 6 female
500 mg/kg bw/day: 6 male, 6 female
100 0mg/kg bw/day: 6 male, 6 female
Control animals:
yes, concurrent vehicle
Details on study design:
Details on study design
- Dose selection rationale:
Based on results from a preliminary 14-day study there was no chenge in the survivel, body weight, Daily clinical observations and Gross pathological examination of 1000 mg/kg/bw/day group. 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.

- Rationale for animal assignment (if not random): Animals were randomized by sex and body weight

- 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

Examinations

Observations and examinations performed and frequency:
Observations and examinations performed & frequency
CAGE SIDE OBSERVATIONS: Yes

- Time schedule: No data available

- Cage side observations checked in table [No.?] were included. : Rats were observed for Behavior, Alterations, Vocalizations, Respiration and Palpebral closure.

DETAILED CLINICAL OBSERVATIONS: Yes

- Time schedule: Once daily, At least once a week there after until

BODY WEIGHT: Yes

- Time schedule for examinations: On the day of randomization, first day of dosing, weekly thereafter and a fasting body weight at scheduled sacrifice on day 29.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): No data available

- 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:
- 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: Yes / No / No data

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: Once a day

- Dose groups that were examined: 0 mg/kg, 250 mg/kg, 500 mg/kg and 1000 mg/kg bw/day.

HAEMATOLOGY: Yes, By using Beckman Coulter haematology analyzer.
- Time schedule for collection of blood:
At termination.

- Anaesthetic used for blood collection: No data available

- Animals fasted: Yes, overnight fasted prior to sampling.

- How many animals: Blood collected from all rats of 0 mg/kg, 250 mg/kg, 500 mg/kg and 1000 mg/kg bw/day group at termination.

- Parameters checked in table [No.?] were examined. : Hemoglobin, Red Blood Corpuscles, Hematocrit, Mean Corpuscular Volume, Mean Corpuscular Hemoglobin, Mean Corpuscular Hemoglobin Concentration, Platelets, White Blood Corpuscles, Neutrophils, Lymphocytes, Eosinophils, Monocytes, Basophil and Prothrombin time were checked, Table No.H; Appendix No.VII.

CLINICAL CHEMISTRY: Yes, By using Dimension XpandPlus and Acculyte 5P.

- Time schedule for collection of blood: At termination.

- Animals fasted: Yes, overnight fasted prior to sampling.

- How many animals: Blood collected from all rats of 0 mg/kg, 250 mg/kg, 500 mg/kg and 1000 mg/kg bw/day group at termination.

- Parameters checked in table [No.?] were examined. : Total Protein, Blood Urea Nitrogen, Urea Nitrogen, Alanine Aminotransferase, Aspartate Aminotransferase, Alkaline Phosphatase, Gamma Glutamyl Transferase, Glucose, Calcium, Phosphorous, Albumin, Total Bilirubin, Creatinine , Total Cholesterol, Triglycerides, Globulin Calculated
Sodium, Potassium, Chloride were checked, Table No.I; Appendix No.VIII.

URINALYSIS: No data available

- Time schedule for collection of urine: No data available

- Metabolism cages used for collection of urine: No data available

- Animals fasted: No data available

- Parameters checked in table [No.?] were examined. : No data available

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

OTHER: Viability, Behavior in Home cage, Vocalizations, Respiration, Palpebral closure, Handling Observations, Urination, Defecation, Prominence of Eye, Lacrimation, Salivation, Piloerection, Examination of Skin / Fur, Stereotype Behaviour, Rearing (Rears), Clonic and Tonic Movements and Severity of Gait were observed.
Sacrifice and pathology:
Sacrifice and pathology
GROSS PATHOLOGY: Yes
Gross necropsy was conducted. All the animals of 0 mg/kg, 250 mg/kg, 500 mg/kg and 1000 mg/kg/bw/day group were sacrificed and gross lesions were noted.

HISTOPATHOLOGY: Yes
Control and treated at the highest dose level of 1000 mg/kg were subjected to sacrifice.

Organs examined: Adrenals, Aorta, Brain (cerebrum, cerebellum and pons), Caecum, Cervix, Colon, Duodenum, Epididymides, Eyes, Heart, Ileum, Jejunum, Kidneys, Liver, Lungs, Mesenteric Lymphnodes, Oesophagus, Ovaries, Pancreas, Pituitary gland, Pharyngeal Lymphnodes, Prostate, Rectum, Skeletal Muscles, Skin with Mammary Gland, Spleen, Sternum with bone marrow, Sciatic Nerve, Spinal Cord (Cervical, mid thoracic and lumbar), Stomach, Seminal Vesicles with Coagulation Gland, Testes, Thymus, Thyroid, Trachea, Vagina, Urinary Bladder and Uterus of 1000 mg/kg/bw/day group.
Statistics:
Data were analyzed for reporting group means and standard deviations with significance between the controls and treated groups, using in-house developed and validated MS-Excel 2003 based statistical software. All the parameters characterized by continuous data such as body weight, per cent body weight change, feed consumption, organ weight, relative organ weight, haematological and clinical chemistry data were subjected to Bartlett’s test to meet the homogeneity of variance before conducting Analyses of Variance (ANOVA) and Dunnett’s t-test. Where the data did not meet the homogeneity of variance, Student’s t-test was performed to calculate significance.

Significance was calculated at 1% as well as 5% level and indicated in the summary tables as follows:

* = Significant than control at 95% level of confidence (p≤ 0.05).
** = Significant than control at 99% level of confidence (p≤ 0.01).

Results and discussion

Results of examinations

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:
not specified
Haematological findings:
no effects observed
Clinical biochemistry findings:
no effects observed
Urinalysis findings:
not specified
Behaviour (functional findings):
not specified
Organ weight findings including organ / body weight ratios:
no effects observed
Gross pathological findings:
no effects observed
Histopathological findings: non-neoplastic:
no effects observed
Histopathological findings: neoplastic:
no effects observed
Details on results:
Clinical signs and mortality
Clinical signs :
Daily clinical observations did not reveal any signs of toxicity in male and female animals from of 0 mg/kg, 250 mg/kg, 500 mg/kg and 1000 mg/kg bw/day dose groups

Mortality:
No mortality were observed in any of the traeted groups of 0 mg/kg, 250 mg/kg, 500 mg/kg and 1000 mg/kg bw/day.

Body weight and weight gain All the rat of 0 mg/kg, 250 mg/kg, 500 mg/kg and 1000 mg/kg/bw/day dose groups exhibited normal body weight gain at the end of the study period of 28 days.
Food consumption and compound intake Food intake of animals for control and 250 mg/kg, 500 mg/kg and 1000 mg/kg /bw/day dose groups were found to be normale throughout the study period of 28 days.
Food efficiency: No data available

Water consumption and compound intake No data available

Opthalmoscopic examination No data available

Haematology Increase in the values of Hb of male rats dosed at 500 mg/kg and 1000 mg/kg, MCHC of male rats dosed at 500 mg/kg, Total WBC of male rats dosed at 1000 mg/kg were observed.
Plateles of female rats dosed at 1000 mg/kg were also increase. The increase in the values of different parameters is within the normal laboratory limits.
Clinical chemistry Increase level of Sodium in male rats dosed at 250 mg/kg and 1000 mg/kg of Benzyl Propionate.

Chloride levels significantly increase in male rats dosed at 250 mg/kg of Benzyl Propionate.

Statistically significant increase of Calcium levels in female rats dosed at 250 mg/kg and increase Bilirubin levels in female rats dosed at 500 mg/kg of Benzyl Propionate were found well increase level of Sodium in female rats dosed at 250 mg/kg, 500 mg/kg and 1000 mg/kg were observed.

Statistically significant decrease of Alkaline Phosphatase levels in female rats dosed at 250 mg/kg, decrease level of Potassium in female rats dosed at 250 mg/kg and 1000 mg/kg and decrease Chloride levels in female rats dosed at 1000 mg/kg Benzyl Propionate were observed.

Although there was an increase/decrease in the values the deviations were within the range of normal laboratory limits.

Urinanalysis: No data available
Neurobehaviour: No data available

Organ weights: Organ weight of 100 mg/kg/bw/day dose group male animals sacrificed on day 29, was found to be comparable with that of controls.

Organ weight of female animals sacrificed on day 29, revealed increased relative weights of liver, ovaries and lungs of 1000 mg/kg dose group.The significant changes in organ weights were observed in female animals from high dose group, the effects are not related to Benzyl Propionate.
Gross pathology All the treatment groups control and 250 mg/kg/bw/day, 500 mg/kg/bw/day and 1000 mg/kg/bw/day of Benzyl Propionate dus not showe any chengs.

Histopathology: Minimal focal to multifocal periportal mononuclear cell infiltration in the liver; minimal interstitial haemorrhages in the kidneys; minimal alveolar haemorrhages and/or alveolar histiocytosis in the lungs; minimal multifocal haemosiderosis and/or diffused congestion in spleen; minimal eosinophilic infiltration and/or luminal dilatation in uterus; minimal luminal seminal coagulum in the urinary bladder; minimal dilatation of zona reticularis and/or presence of accessory adrenocortical tissue in adrenals; minimal multifocal haemorrhages in thymus; presence of ultimobranchial cysts in thyroid; in male or female animals of control and 1000 mg/kg/bw/day dose group. All the changes observed in the control and 1000 mg/kg/bw/day dose treatment group animals were similar and the effect observed are not due to Benzyl Propionate.

Effect levels

Dose descriptor:
NOAEL
Effect level:
1 000 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Based on Clinical biochemistry, Organ weight, Gross pathological, Histopathology and survival.

Target system / organ toxicity

Critical effects observed:
not specified

Applicant's summary and conclusion

Conclusions:
NOAEL was considered to be 1000 mg/kg bw/day when Sprague-Dawley rats exposed to Benzyl Propionate orally foe 28 days.
Executive summary:

A subacute study was conducted to evaluate the toxic effects of repeated administration of Benzyl Propionate in male and female Sprague-Dawley rats by gavage. Benzyl Propionate was administered to 6 animals/sex/species in Corn oil at doses of 0,250,500 and 1000 mg/kg/bw/day for 28 days. All rats of 250,500 and 1000 mg/kg/bw/day dose group Survive though-out the study, Benzyl Propionate have no effect on mortality. Blood samples for Clinical Biochemistry and Haematology were collected. No abnormalities occurred that could be directly attributed to Benzyl Propionate treatment. Although significant change in relative weights of liver, ovaries and lungs of female were observed in 1000 mg/kg/bw/day dose groups. No Benzyl Propionate related gross pathological or histological changes were seen and findings were not Benzyl Propionate dependent and hence considered to be of no toxicological importance. Therefore NOEAL for repeated dose toxicity study was considered to be 1000 mg/kg/bw/day in male and female Sprague-Dawley rats when exposed to Benzyl Propionate by oral route for 28 days.

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