Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
05 March 2018 - 27 March 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
December 2001
Deviations:
no
Qualifier:
according to
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Version / remarks:
December 2002
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
May 2008
Deviations:
no
Qualifier:
according to
Guideline:
other: Appendix to Director General Notification, No. 12-Nousan-8147. Agricultural Production Bureau, Ministry of Agriculture, Forestry and Fisheries of Japan (JMAFF)
Version / remarks:
November 2000, including the most recent revisions
Deviations:
no
GLP compliance:
yes
Test type:
acute toxic class method
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid: viscous
Details on test material:
- Physical appearance: dark brown to black viscous liquid
- Storage conditions: in refrigerator (2-8°C) protected from light
Specific details on test material used for the study:
pH: 7.3
Specific gravity/density: 1.10 g/mL

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Crl: WI(Han)
Sex:
female
Details on test animals and environmental conditions:
- Source: Charles River Deutschland, Sulzfeld, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: young adult animals of approx. 8-9 weeks old
- Weight at study initiation: 148 to 165 g
- Fasting period before study: Animals were deprived of food overnight (for a maximum of 20 hours) prior to dosing and until 3-4 hours after administration of the test item.
- Housing: Group housing of 3 animals per cage in polycarbonate cages (MIV type; height 18 cm.), containing sterilized sawdust as bedding material and paper as cage-enrichtment.
- Diet: Free access to pelleted rodent diet (SM R/M-Z from SSNIFF® Spezialdiäten GmbH, Soest, Germany).
- Water: Free access to tap water.
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS (set to maintain)
- Temperature (°C): 18-24 (actual: 20-21)
- Humidity (%): 40-70 (actual: 28-51) - this deviation is considered not to have affected the integrity of the study because it did not noticeably affect the clinical condition of the animals or the outcome of the study.
- Air changes (per hr): at least 10
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 06 March 2018 - 27 March 2018

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
unchanged (no vehicle)
Details on oral exposure:
MAXIMUM DOSE VOLUME APPLIED: The dose volume was calculated as dose level (g/kg) / specific gravity/density of the test item (g/mL). The maximum dose level was 2000 mg/kg bw.

DOSAGE PREPARATION:
- Adjustment was made for specific gravity of the vehicle. No correction was made for the purity/composition of the test item.

CLASS METHOD:
- Rationale for the selection of the starting dose: The dose levels were based on the OECD test guidelines and were selected from the series 5 (lowest dose level), 50, 300 and 2000 (highest dose level) mg/kg body weight. The starting dose level should be the one that is likely to produce mortality in at least some of the animals and was selected based on available toxicity data of the test item.
Doses:
2000 mg/kg body weight
The study was conducted in a stepwise manner with two groups of 3 females. The first group was treated at a dose level of 2000 mg/kg bw. Based on the results, one additional group was dosed at 2000 mg/kg bw.
No. of animals per sex per dose:
6 females/dose (2 groups of three females in a stepwise manner)
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:
*Mortality/Viability: twice daily;
*Body weights: on the day of dosing (fasted weight) and on days 1 (pre-administration), 8 and 15;
*Clinical signs: at periodic intervals on the day of dosing (day 1) and once daily thereafter
- Necropsy of survivors performed: yes
- Other examinations performed: no
Statistics:
The LD50 cut-off value was established based on OECD guideline 423. No statistical analysis was performed (the method used is not intended to allow the calculation of a precise LD50 value).

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
No mortality occurred.
Clinical signs:
Hunched posture and piloerection were noted for the animals between days 1 and 5.
Body weight:
The body weight gain shown by the 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:
No abnormalities were found at macroscopic post mortem examination of the animals.

Applicant's summary and conclusion

Interpretation of results:
GHS criteria not met
Remarks:
Not classified according to Regulation (EC) No. 1272/2008
Conclusions:
In an acute oral toxicity study with rats, performed according to OECD/EC test guidelines, the LD50 of Cocoa Shell Extract (Pot) was established to exceed 2000 mg/kg bw. Based on the result obtained in this study, Cocoa Shell Extract (Pot) is not classified for acute toxicity under GHS and under Regulation (EC) 1272/2008.