Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
09 - 26 Aug 2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
/ no justification for exceptional limit dose of 5000 mg/kg bw provided

Data source

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

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
17 Dec 2001
Deviations:
yes
Remarks:
/ no justification for exceptional limit dose of 5000 mg/kg bw provided
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
30 May 2008
Deviations:
yes
Remarks:
/ no justification for exceptional limit dose of 5000 mg/kg bw provided
GLP compliance:
yes (incl. certificate)
Remarks:
OGYÉI, Országos Gyógyszerészeti és Élelmezés-egészségügyi Intézet, Budapest, Hungary
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:
STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: at room temperature, protected from light and humidity

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Crl:WI rats
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: TOXI COOP ZRT., Budapest, Hungary
- Females (if applicable) nulliparous and non-pregnant: yes
- Age at study initiation: 11 weeks
- Weight at study initiation:
group of 1st step: 244 g
group 2nd step: 230 - 231 g
group of 3rd step: 247 - 257 g
- Fasting period before study: animals were fasted over-night before and 3 hours after treatment
- Housing: 3 animals of the same sex per cage in Type II polypropylene/polycarbonate cages, laboratory bedding
- Diet: ssniff® SM R/M-Z+H complete diet for rats and mice (ssniff Spezialdiäten GmbH, Soest, Germany), ad libitum
- Water: tap water, ad libitum (analysis was performed)
- Acclimation period: 27 days (group of 1st step), 28 days (group of 2nd step) and 29 days (group of 3rd step)

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): >10
- Photoperiod (hrs dark / hrs light): 12/ 12

IN-LIFE DATES:
group of 1st step: From: 09 To: 9 Aug 2016
group 2nd step: From: 10 To: 25 Aug 2016
group of 3rd step: From: 11 To: 26 Aug 2016

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: Helianthi Annui Oleum Raffinatum (sunflower oil)
Details on oral exposure:
VEHICLE
- Concentration in vehicle: 500 and 200 mg/mL
- Amount of vehicle (if gavage): 10 mL/kg bw
- Justification for choice of vehicle: The applied vehicle is one of the standard vehicles used in acute toxicity studies.
- Lot/batch no. (if required): 8001528001

MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg bw

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: The starting dose was selected on basis of the available information about the test item. The acute toxic class method was carried out involving a stepwise procedure (according to OECD 423) with the use of 5000 mg/kg bw as the starting dose in one female rat (group of 1st step). Due to the observed death of this female, dosing was proceeded at 2000 mg/kg bw in three females (group of 2nd step) in accordance with the flow charts provided in Annex 2 of OECD 423. No animal died in the second step at 2000 mg/kg bw dose level, so treatment with 2000 mg/kg bw was repeated on further three female rats (group of 3rd step). No animal died in the third step, and hence the test was finished.
Doses:
5000 and 2000 mg/kg bw
No. of animals per sex per dose:
1 animal for 5000 mg/kg bw
3 animals for 2000 mg/kg bw per step (6 in total)
Control animals:
no
Remarks:
not required
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:
mortality: at least once during the first 30 minutes after substance application, then 1, 2, 3 and 4 h after gavage (all groups) and thereafter twice per day for 14 days (groups of 2nd + 3rd step)
general observations: at least once during first 30 minutes after substance application, then 1, 2, 3 and 4 h after gavage (all groups) and thereafter daily for 14 days (groups of 2nd + 3rd step)
weighing: on Day 0 (all groups), 7 and on Day 15 (groups of 2nd + 3rd step)
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, general state, external appearance, behavior, gross histopathology
Statistics:
No statistical anylysis were performed.

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 - <= 5 000 mg/kg bw
Based on:
test mat. (dissolved fraction)
Mortality:
After administration of 5000 mg/kg bw, the test female died within 4 h after treatment. No mortality was observed after administration of 2000 mg/kg bw.
Clinical signs:
5000 mg/kg bw, group of 1st step: On Day 1 (0.5 - 4 h after treatment) several clinical symptoms were observed: decreased activity (4 / 5 observations), vocalisation (3/5), tremor (1/5), abnormal gait (3/5), bedding digging (1/5), closed eyes (3/5), salivation (1/5), piloerection (2/5) and dyspnea (3/5).
2000 mg/kg bw, group of 2nd step: On Day 1 (1 - 2 h after treatment) irritability was observed in all animals.
2000 mg/kg bw, group of 3rd step: No clinical signs of toxicity were observed during the observation period.
Body weight:
All animals showed normal body weight gain during the observation period. Due to the mortality of the female dosed with 5000 mg/kg bw, no body weight was determined for this female.
Gross pathology:
No pathological changes related to the test item were observed.

2000 mg/kg bw, group of 2nd step: In 2/3 animals moderate hydrometra was observed. Hydrometra is a physiological finding and connected to the cycle of the animal. Thus, the effect is not considered as treatment-related.
2000 mg/kg bw, group of 3rd step: In 1/3 animals moderate hydrometra was observed. 1/3 animals showed a colon full of gas, which is regarded as an individual variation. Most likely the observation is a consequence of the anaesthesia.

Any other information on results incl. tables

Table 1: Results of acute oral toxicity study

Dose
[mg/kg bw]

Toxicological results*

Duration of clinical signs

Time of death

Mortality (%)

2000

0/3/3

1-2 h - day 14

---

0

2000

0/0/3

---

---

0

5000

1/1/1

0.5 h - death

Day 1

100

LD50 = >2000 - ≤ 5000 mg/kg bw

*first number = number of dead animals

 second number = number of animals with clinical signs

 third number = number of animals used

Applicant's summary and conclusion

Interpretation of results:
other: CLP/ EU GHS criteria not met, no classification required according to Regulation (EC) No 1272/2008
Conclusions:
CLP: not classified