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Toxicological information

Acute Toxicity: oral

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Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report Date:
2016

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
2001
Deviations:
yes
Remarks:
No observation for clinical signs was performed on Day 7 of Group 1 (animals Nos.1-3) of the study. Evaluation: Sufficient data was available for evaluation of the study.
GLP compliance:
yes
Test type:
acute toxic class method
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Batch No. 605803
- Expiration date of the lot/batch: 23 October 2016

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: At room temperature

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Deutschland, Sulzfeld, Germany
- Females (if applicable) nulliparous and non-pregnant: Yes
- Age at study initiation: approximately 9-11 weeks old
- Weight at study initiation: 174 g
- Fasting period before study: None
- Housing: Group housing of 3 animals per cage in labeled Makrolon cages (MIV type; height 18 cm.) containing sterilized sawdust as bedding material (Lignocel S 8-15, JRS - J.Rettenmaier & Söhne GmbH + CO. KG, Rosenberg, Germany) and paper as cage-enrichment (Enviro-dri, Wm. Lillico & Son (Wonham Mill Ltd), Surrey, United Kingdom).
- Diet (e.g. ad libitum): Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water (e.g. ad libitum): Free access to tap water.
- Acclimation period:Acclimatization period was at least 5 days before start of treatment under laboratory conditions.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 to 24°C
- Humidity (%): 40 to 70%
- Air changes (per hr): at least 10 air changes/hour
- Photoperiod (hrs dark / hrs light): 12-hour light/12-hour dark cycle

IN-LIFE DATES: From: 28 April 2016 To: 07 June 2016

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
DMSO
Details on oral exposure:
VEHICLE
- Concentration in vehicle: Dosed at 300 mg/kg and 2000 mg/kg at dose volumes of 4 mL/kg bw
- Amount of vehicle (if gavage): 4 mL/kg bw
- Justification for choice of vehicle: The vehicle was selected based on trial preparations performed at Charles River Den Bosch and on test item data supplied by the Sponsor.

MAXIMUM DOSE VOLUME APPLIED: 4 mL/kg bw

DOSAGE PREPARATION (if unusual): The preparations (w/w) were kept at room temperature and were dosed within 4 hours after adding the vehicle to the test item. Homogeneity was assessed by visual inspection of the solutions and the formulations were stirred during dosing, which ensures homogeneity sufficient for these kinds of studies. Adjustment was made for specific gravity of the vehicle. No correction was made for purity of the test item.
The concentration of the test item in vehicle was varied to allow constant dosage volume in terms of mL/kg body weight.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: Based on a previous study.
Doses:
300 mg/kg bw
2000 mg/kg bw
No. of animals per sex per dose:
3/dose
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Observations for mortality twice daily. Body weights were recorded on days 1, 8, and 15 or at death.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weights

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 300 - < 2 000 mg/kg bw
Based on:
test mat.
Mortality:
At 2000 mg/kg, on Day 2 one animal was sacrificed for humane reasons after showing severe clinical signs (e.g. lethargy, tremors and pale- and lean appearance). On Day 4, one animal was found dead. At 300 mg/kg, no mortality occurred.
Clinical signs:
At 2000 mg/kg, lethargy, tremor, muscle twitching, abnormal posture, ventro-lateral recumbency, hunched posture, uncoordinated movements, loss of righting reflex, increased activity, abnormal gait, hypotonia, deep respiration, piloerection, shaking head, dehydrated, ptosis, pale appearance, red staining of the eyes, salivation and/or lean appearance were noted for the animals between Days 1 and 6. At 300 mg/kg, lethargy, hunched posture, piloerection, ptosis, rales and/or slow breathing were noted for the animals between Day 1 and 8.
Body weight:
The low body weight of animal no. 5 on Day 15 was considered not reliable since no corroborative findings (e.g. lean appearance) were noted. This body weight was excluded from interpretation of the study. Sufficient data was available for evaluation. The body weight gain shown by the remaining surviving animals over the study period was considered to be similar to that expected for normal untreated animals of the same age and strain.
Gross pathology:
At 2000 mg/kg, abnormalities of the spleen (reduced in size) and/or gastro-intestinal tract (GI-tract; distended with gas) were noted for the two animals that died during the study at macroscopic post mortem examination. Additionally, emaciation was noted for the animal sacrificed for humane reasons. At 300 mg/kg, an abnormality of the gastro-intestinal tract (GI-tract; distended with gas) was noted for one animal.

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria
Conclusions:
Based on the results of the study, the rat oral LD50 of the test article is between 300 -2000 mg/kg bw.
Executive summary:

The acute oral lethality of the test article was determined in rats. Female Wistar rats received 300 mg/kg (6 females) or 2000 mg/kg (3 females) test article dissolved in DMSO via oral gavage. Observations for mortality (recorded twice daily), body weights (day 1, 8, 15 or at death), clinical signs (once daily), and macroscopic examination (at necropsy). At 2000 mg/kg all animals exhibited clinical signs of toxicity including lethargy (3/3), spasms (3/3), abnormal and/or hunched posture (3/3), abnormal gait (3/3), uncoordinated movements (2/3), ventro-lateral recumbency (1/3), loss of righting reflex (1/3), increased activity (1/3), hypotonia (1/3), deep breathing (1/3), piloerection (3/3), shaking head (1/3), paleness (1/3), salivation (1/3), dehydration (2/3), and ptosis (3/3). On Day 2 one animal in the 2000 mg/kg group was euthanized for humane reasons after exhibiting symptoms including lethargy, tremors, and pale and lean appearance. Another animal was found dead on Day 4 in this dose group. These two animals had reduced spleen sizes, and gastrointestinal distension upon post mortem examination. At the 300 mg/kg animals exhibited piloerection and hunched posture (6/6), lethargy (3/6), ptosis (3/6), rales (1/6), and slow breathing (1/6) between Days 1 and 8. No mortality was seen at this dose level, however, and only one animal had gastrointestinal distention due to gas noted on necropsy. Based on the results of the study, the rat oral LD50 of the test article is between 300 -2000 mg/kg bw.