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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:
Experimental start date: 22 December 2016 Experimental completion date: 15 February 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 guideline
Qualifier:
according to guideline
Guideline:
OECD Guideline 420 (Acute Oral Toxicity - Fixed Dose Method)
GLP compliance:
yes (incl. QA statement)
Test type:
fixed dose procedure
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
[Reaction mass of 2,7-Naphthalenedisulfonic acid, 5-amino-3-[[4-[2-[4-[(7-amino-1-hydroxy-3-sulfo-2-naphthalenyl)azo]-2-sulfophenyl]ethenyl]-3-sulfophenyl]azo]-4-hydroxy-, compd. with 2,2',2''-nitrilotris[ethanol] (1:5) and 3,3'-[ethylenebis[(3-sulpho-p-phenylene)azo]]bis[5-amino-4-hydroxynaphthalene-2,7-disulphonic] acid, compound with 2,2',2''-nitrilotriethanol (1:6) and 3,3'-[vinylenebis[(3-sulpho-p-phenylene)azo]]bis[6-amino-4-hydroxynaphthalene-2-sulphonic] acid, compound with 2,2',2''-nitrilotriethanol (1:4)]
EC Number:
916-899-6
Molecular formula:
not available
IUPAC Name:
[Reaction mass of 2,7-Naphthalenedisulfonic acid, 5-amino-3-[[4-[2-[4-[(7-amino-1-hydroxy-3-sulfo-2-naphthalenyl)azo]-2-sulfophenyl]ethenyl]-3-sulfophenyl]azo]-4-hydroxy-, compd. with 2,2',2''-nitrilotris[ethanol] (1:5) and 3,3'-[ethylenebis[(3-sulpho-p-phenylene)azo]]bis[5-amino-4-hydroxynaphthalene-2,7-disulphonic] acid, compound with 2,2',2''-nitrilotriethanol (1:6) and 3,3'-[vinylenebis[(3-sulpho-p-phenylene)azo]]bis[6-amino-4-hydroxynaphthalene-2-sulphonic] acid, compound with 2,2',2''-nitrilotriethanol (1:4)]
impurity 1
Reference substance name:
not applicable
Molecular formula:
not applicable
IUPAC Name:
not applicable
Test material form:
solid
Specific details on test material used for the study:
Identification: Bayscript Blaukomponente TEA
EC number: 916-899-6
Empirical formula: UVCB substance
Correction factor: 1.1236
Physical state / Appearance: Sponsor description: dark blue solid
Envigo description: dark brown solid
Expiry Date: 28 April 2017
Storage Conditions: room temperature in the dark
Stability under test conditions: Homogeneity/stability analysis was performed under Envigo Study Number CY71SC prior to commencement of this study.
The dose formulations were adjusted for purity

Test animals

Species:
rat
Strain:
Wistar
Sex:
female
Details on test animals or test system and environmental conditions:
Animal Information
Female Wistar (RccHan™:WIST) strain rats were supplied by Envigo RMS (UK) Limited, Oxon, UK. On receipt the animals were randomly allocated to cages. The females were nulliparous and non pregnant. After an acclimatization period of at least 5 days the animals were selected at random and given a number unique within the study by indelible ink marking on the tail and a number written on a cage card. At the start of the study the animals were 8 to 12 weeks of age. The body weight variation did not exceed ±20% of the mean body weight at the start of treatment.
Animal Care and Husbandry
The 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 Envigo RMS (UK) Limited, Oxon, UK) was allowed throughout the study. The diet, drinking water and bedding were routinely analyzed and were considered not to contain any contaminants that would reasonably be expected to affect the purpose or integrity of the study.
The temperature and relative humidity were set to achieve limits of 19 to 25 °C and 30 to 70% respectively. The rate of air exchange was at least fifteen changes per hour and the lighting was controlled by a time switch to give 12 hours continuous light and 12 hours darkness.
The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
water
Details on oral exposure:
Test Item Preparation and Analysis
For the purpose of the study the test item was freshly prepared, as required, as a solution in distilled water.
Homogeneity/stability analysis was performed under Envigo Study Number CY71SC prior to commencement of this study.
An analytical investigation was conducted to determine the concentration of the test item formulation in the main test (2247 mg/kg dose level, equivalent to 2000 mg active ingredient/kg body weight).
MAXIMUM DOSE VOLUME APPLIED: 10 mL/kg
Doses:
338 mg/kg and 2247 mg/kg (eq. to 300 and 2000 mg/kg active ingredient, respectively)
No. of animals per sex per dose:
1 female at 338 mg/kg
5 females at 2247 mg/kg
Control animals:
no
Details on study design:
Using available information on the toxicity of the test item, 338 mg/kg (equivalent to 300 mg active ingredient/kg body weight) was chosen as the starting dose.

In the absence of mortality at a dose level of 338 mg/kg (equivalent to 300 mg active ingredient/kg body weight), an additional animal was treated at 2247 mg/kg (equivalent to 2000 mg active ingredient/kg body weight).

In the absence of mortality at a dose level of 2247 mg/kg (equivalent to 2000 mg active ingredient/kg body weight), an additional group of animals was treated at 2247 mg/kg (equivalent to 2000 mg active ingredient/kg body weight).

A total of five animals were therefore treated at a dose level of 2247 mg/kg (equivalent to 2000 mg active ingredient/kg body weight) in the study.
All animals were dosed once only 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 group to confirm the survival of the previously dosed animals.
Clinical observations were made 30 minutes, 1, 2, and 4 hours after dosing and then daily for 14 days. Morbidity and mortality checks were made twice daily, early and late during normal working days, and once daily at weekends and public holidays.
Individual body weights were recorded on Day 0 (the day of dosing) and on Days 7 and 14.
At the end of the observation period the animals were killed by cervical dislocation. All animals were subjected to gross necropsy. This consisted of an external examination and opening of the abdominal and thoracic cavities. The appearance of any macroscopic abnormalities was recorded. No tissues were retained.

Results and discussion

Effect levelsopen allclose all
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 247 mg/kg bw
Based on:
test mat.
Sex:
female
Dose descriptor:
LD50
Effect level:
> 2 000 mg/kg bw
Based on:
act. ingr.
Mortality:
There was no mortality at 338 mg/kg, or 2247 mg/kg.
Clinical signs:
At 338 mg/kg:
No signs of systemic toxicity were noted during the observation period.
Purple/blue stained urine was noted in the cage during the day of dosing. Purple/black stained feces was noted in the cage 3 to 6 days after dosing.

At 2247 mg/kg:
No signs of systemic toxicity were noted during the observation period.
Purple or blue stained urine and feces were noted in the cages 1 to 4 days after dosing. Purple stained feces persisted 5 days after dosing in the cage of the initial treated animal.
A blue colored tint of the extremities and skin was noted in the additional four treated animals 4 hours and 1 to 4 days after dosing.
Body weight:
The animal showed expected gains in body weight over the observation period at both 338 mg/kg and 2247 mg/kg.
Gross pathology:
At 338 mg/kg:
No abnormalities were noted at necropsy.

At 2247 mg/kg:
No abnormalities were noted at necropsy of the initial treated animal.
Dark kidneys were noted at necropsy of the four additional treated animals.

Applicant's summary and conclusion

Conclusions:
The acute oral median lethal dose (LD50) of Bayscript Blaukomponente TEA in the female Wistar strain rat was estimated to be greater than 2247 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight).
Executive summary:

The study according to OECD TG 420 was performed to assess the acute oral toxicity of the test item in the Wistar strain rat. Following a sighting test at dose levels of 338 mg/kg and 2247 mg/kg (equivalent to 300 and 2000 mg active ingredient/kg body weight, respectively), a further group of four fasted females was given a single oral dose of test item, as a solution in distilled water, at a dose level of 2247 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight). Clinical signs and body weight development were monitored during the study. All animals were subjected to gross necropsy.

Results

There were no deaths. There were no signs of systemic toxicity. Purple/blue stained urine and blue/purple/black stained feces were noted at both dose levels. A blue colored tint of the extremities and skin was noted in the additional four animals treated at a dose level of 2247 mg/kg (equivalent to 2000 mg active ingredient/kg body weight). All animals showed expected gains in body weight. Dark kidneys were noted at necropsy of the four additional animals treated at a dose level of 2247 mg/kg (equivalent to 2000 mg active ingredient/kg body weight). No abnormalities were noted at necropsy of the animal treated at a dose level of 338 mg/kg (equivalent to 300 mg active ingredient/kg body weight) and the initial animal treated at a dose level of 2247 mg/kg (equivalent to 2000 mg active ingredient/kg body weight).

Conclusion

The acute oral median lethal dose (LD50) of Bayscript Blaukomponente TEA in the female Wistar strain rat was estimated to be greater than 2247 mg/kg body weight (equivalent to 2000 mg active ingredient/kg body weight).