Registration Dossier

Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
13 January 2015 to 03 February 2015
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2015
Report Date:
2015

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.1 bis (Acute Oral Toxicity - Fixed Dose Procedure)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
fixed dose procedure
Limit test:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Physical state: Dark yellow viscous liquid
- Analytical purity: 100 %
- Expiration date of the lot/batch: 12 June 2016
- Stability under test conditions: Room temperature in the dark

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals and environmental conditions:
ANIMALS and ANIMAL HUSBANDRY

- Female Wistar rats were supplied by Harlan Laboratories UK Ltd, Oxon, UK.
- The animals were randomly allocated to cages on receipt.
- Females were nulliparous and non-pregnant.
- After an acclimatisation period of at least 5 days the animals were selected at random and given a unique number within the study by indelible ink-marking on the tail and the number was written on a cage card.
- At the start of the study the animals were 8 to 12 weeks of age.
- Body weight did not exceed ± 20 % of the body weight of the initially dosed animal.
- Animals were housed in groups of up to four in suspended solid-floor polypropylene cages furnished with woodflakes.
- With the exception of an overnight fast immediately before dosing, and for approximately 3 to 4 hours after dosing, free access to mains drinking water and food (2014C Teklad Global Rodent diet supplied by Harlan Laboratories UK Ltd, Oxon, UK) was allowed throughout the study.
- Diet, drinking water and bedding were routinely analysed and were considered not to contain any contaminants that would reasonably be expected to affect purpose or integrity of the study.
- Temperature and relative humidity were set to achieve limits of 19 to 25 °C and 30 to 70 % respectively.
- Rate of air exchange was at least 15 changes per hour.
- Lighting was controlled by a time switch to give 12 hours continuous light (06:00 to 18:00) and 12 hours darkness.
- Animals were provided with environmental enrichment items considered not to contain any contaminant at a level that might affect the purpose or integrity of the study.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
arachis oil
Details on oral exposure:
TEST ITEM FORMULATION and EXPERIMENTAL PREPARATION
- The test item was freshly prepared as a solution in arachis oil BP when required.
- Arachis oil was used because the test material did not dissolve/suspend in distilled water.
- The test item was formulated within 2 hours of being applied to the test system and it was assumed that the formulation was stable for that duration.
- No analysis was conducted to determine the homogeneity, concentration or stability of the test item formulation. This is an exception with regard to GLP and was reflected in the GLP compliance statement.
Doses:
- Dose level: 2000 mg/kg bw
- Concentration: 200 mg/mL
- Dose volume: 10 mL/kg
No. of animals per sex per dose:
Single female animal followed, in the absence of toxicity, by an additional group of four female animals.
Control animals:
no
Details on study design:
- All animals were dosed once by gavage using a metal cannula attached to a graduated syringe.
- The volume administered to each animal was calculated according to the fasted body weight at the time of dosing.
- Treatment of animals was sequential.
- Sufficient time was allowed between each dose level to confirm the survival of the previously dosed animals.
- Clinical observations were made 0.5, 1, 2 and 4 hours after dosing and then daily for 14 days.
- Morbidity and motality checks were made twice daily.
- Individual body weights were recorded on Day 0 (the day of dosing) and on Days 7 and 14.
- Animals were killed by cervical dislocation at the end of the observation period.
- All animals were subjected to gross necropsy consisting of external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded but no tissues were retained.
Statistics:
EVALUATION OF DATA

The test item was evaluated according to Annex 3 of the OECD Guidelines for Testing of Chemicals No 420 "Acute Oral Toxicity - Fixed Dose Method (adopted 17 December 2001) as shown in the flow chart in Appendix 2 (attached):

- Number of animals that died during the study (or that were killed for humane reasons).
- Determination of the nature, severity, onset and duration of toxic effects (evident toxicity refers to the toxic effects of sufficient severity that administration of the next higher dose level could result in development of severe signs of toxicity and probable mortality).
- Effects on body weights and abnormalities noted at necropsy were also identified.
- Using the mortality data obtained, an estimate of the acute oral median lethal dose (LD50) of the test item was made.

Results and discussion

Effect levels
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
test mat.
Mortality:
- No animal deaths took place during the study.
Clinical signs:
- Individual clinical observations and mortality data are given in Table 1 (attached).
Body weight:
- Individual body weights and body weight changes are given in Table 2 (attached).
- All animals showed expected gains in body weight over the observation period.
Gross pathology:
- Individual necropsy findings are shown in Table 3 (attached).
- No abnormalities were noted at necropsy.

Applicant's summary and conclusion

Conclusions:
The acute oral median lethal dose (LD50) of the test item in the femal Wistar strain rat was considered to be greater than 2000 mg/kg body weight.
Executive summary:

GUIDELINE

The study was designed to be compatible with OECD Guideline for Testing of Chemicals No 420 "Acute Oral Toxicity - Fixed Dose Method" (2001) and Method B1 bis Acute Toxicity (Oral) of Commission Regulation (EC) No 440/2008.

METHOD

Following a sighting test at a dose level of 2000 mg/kg, an additional four fasted female animals were given a single oral dose of the test item, as a solution in arachis oil BP, at a dose level of 2000 mg/kg body weight. Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

RESULTS

Mortality: There were no deaths.

Clinical observations: There were no signs of systemic toxicity.

Body weight: All animals showed expected gains in body weight.

Necropsy: No abnormalities were noted at necropsy.

CONCLUSION

The acute oral median lethal dose (LD50) of the test item in the female Wistar strain rat was considered to be greater than 2000 mg/kg body weight.