Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From the 13nd of October, 2016 to the 12nd of April, 2017
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 (Acute Toxicity (Oral))
Qualifier:
according to
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
GLP compliance:
yes (incl. certificate)
Test type:
acute toxic class method

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
During the study,the test item was suspended as follows:
Vehicle: Corn oil Concentration: 200mg/mL.
The concentration was calculated and expressed interms of test item corrected for purity (iron complexes, sodium salt).

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
female
Details on test animals and environmental conditions:
ANIMAL SUPPLY AND ACCLIMATISATION
Species and strain: Rat, Hsd: Sprague Dawley SD
Sex: Females (nulliparous and non-pregnant)
Age and weight at order: 7 weeks old; 168-170 grams
Supplier: Envigo RMS srl, San Pietro al Natisone (UD), Italy
Date of arrival: 15 March 2017
Weight range at arrival: 153.2-162.3 grams
Acclimatisation period: At least 5 days
Veterinary health check: During acclimatisation period

ANIMAL HUSBANDRY
Animals per cage: 3 during the study; up to 5 during acclimatisation
Housing: Polisulphone solid bottomed cages measuring 59.5×38×20 cm with nesting material provided into suitable bedding bags
Cage control: Daily inspected and changed as necessary (at least 3 times/week)
Water: Drinking water supplied to each cage via a water bottle
Water: supply ad libitum
Diet: 4 RF 18 (Mucedola S.r.l., Via G. Galilei, 4, 20019, Settimo Milanese (MI) Italy)
Diet supply: ad libitum throughout the study except for the dosing procedure
Room: lighting Artificial (fluorescent tubes), daily light/dark cycle of 12/12 hours
Air: changes Approximately 15 to 20 air changes per hour
Temperature range: 22 °C ± 2 °C
Relative humidity range: 55 % ± 15 %

SELECTION/ALLOCATION
Random at arrival. The body weight of each individual was within 20% of the group’s mean. Animals were unequivocally numbered within the study.
The animal number together with the study number ensured a unique animal numbering for any study employing computerised data collection. The computer system used in this study was Pristima version 6.3.2.

ANIMAL IDENTIFICATION
Animals were permanently identified, following arrival, by a combination of ear notch (units) and tattoo on the hind feet. Animals were identified by odd numbers.

FASTING PROCEDURE
Food was removed from the cages overnight prior to dosing (Day -1) and was made available approximately 4 hours after dosing.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on oral exposure:
A first group of 3 female animals was dosed at a level of 2000 mg/kg (Step 1).
A second sub-group, similarly composed, was then dosed at the same dose level (Step 2).
No further doses were investigated since the objective of the study had been achieved (no mortality).

FREQUENCY OF TREATMENT
Animals were dosed once only on Day 1.

DOSE CALCULATION
On the day of dosing (Day 1), the amount of the formulated test item to be administered was
calculated for each fasted animal according to body weight.

DOSING METHOD
The formulated test item was administered, by gavage, at a dose volume of 10 mL/kg using a
plastic feeding tube attached to a graded syringe.
Doses:
2000 mg/kg
No. of animals per sex per dose:
3 female per group
Details on study design:
TREATMENT (mg/kg)
2000 in terms of test item corrected for purity (60.42% iron complex, sodium salt)
Rat Numbers Females (odd)
29, 31, 33 (Group 1 - Step 1)
41, 43, 45 (Group 3 - Step 2)

OBSERVATION
Mortality and morbidity
Throughout the study all animals were checked twice daily.
Clinical signs
Animals were observed for clinical signs as indicated below:
– Day of dosing
· Session 1: on dosing
· Session 2: approximately 0.5 hour after dosing
· Session 3: approximately 2 hours after dosing
· Session 4: approximately 4 hours after dosing
– Daily thereafter for a total of 14 days (Session 1).

Body weight
All animals were weighed at allocation to the study (Day -1), on the day of dosing (Day 1) and on Days 2, 8 and 15.
Body weight change calculated for Days 2, 8 and 15 of the dosing phase was relevant to Day 1 of the phase.

TERMINAL STUDY
Termination
All animals were sacrificed on Day 15.
Euthanasia method
Animals were sacrificed by carbon dioxide narcosis.
Necropsy procedure
Necropsy was carried out on all animals (gross necropsy examination for both external and internal abnormalities, with particular attention to the gastro-intestinal tract).

Results and discussion

Effect levelsopen allclose all
Sex:
female
Dose descriptor:
LD0
Effect level:
> 2 000 mg/kg bw
Based on:
act. ingr.
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
act. ingr.
Mortality:
No mortality occurred in the first group of animals initially dosed at 2000 mg/kg (Group 3, Step 1) and in the further group of 3 females dosed at
the same dose level (Group 5, Step 2).
Clinical signs:
No clinical signs were observed in the first group of animals initially dosed at 2000mg/kg (Group 1, Step 1) and in the further group of 3 females dosed at the same dose level (Group 3, Step 2).
Body weight:
Changes in body weight observed during the study were within the expected range for this strain and age of animals.
Gross pathology:
No abnormalities were observed at necropsy examination performed on all animals dosed at 2000 mg/kg (Groups 3 and 5) at the end of the observation period.

Applicant's summary and conclusion

Interpretation of results:
other: Not classified
Remarks:
Classification criteria according to the CLP Regulation 1272/2008 and its amendments
Conclusions:
LD50 2000 mg/kg
no mortality
Executive summary:

Method

The acute toxicity of the substance was investigated following a single oral administration to the Sprague Dawley rat followed by a 14-day observation period, according to OECD 423.

A first group of 3 female animals was initially dosed at 2000 mg/kg (Step 1).

Observations

No mortality occurred and no clinical signs were observed.

A second group of 3 female animals was then dosed at the same dose level (Step 2). No death occurred and no clinical signs were noted.

Body weight changes recorded during the study were within the expected range for this strain and age of animals.

No abnormalities were observed at necropsy examination performed at the end of the observation period on animals of both groups.

Results

These results indicate that the test item did not induce toxic effects in the rat following oral administration of a single dose at a level of 2000 mg/kg. The lack of mortality demonstrates the acute toxicity expected (ATE)to be greater than 2000 mg/kg body weight.