4,5-dihydro-5-oxo-4-[[4-[[2-(sulphooxy)ethyl]sulphonyl]phenyl]azo]-1-(4-sulphophenyl)-1H-pyrazole-3-carboxylic acid, sodium salt

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2016-2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Version / remarks:

adopted 17 December 2001

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Version / remarks:

May 2008

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.1100 (Acute Oral Toxicity)

Version / remarks:

EPA 712-C-02-190, December 2002

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: Procedures and facilities comply with the requirements of Directive 2010/63/EU and the national legislation defined in the animal protection law concerning the protection of animals used for experimental and other scientific procedures

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

4,5-dihydro-5-oxo-4-[[4-[[2-(sulphooxy)ethyl]sulphonyl]phenyl]azo]-1-(4-sulphophenyl)-1H-pyrazole-3-carboxylic acid, sodium salt

EC Number:

287-722-1

EC Name:

4,5-dihydro-5-oxo-4-[[4-[[2-(sulphooxy)ethyl]sulphonyl]phenyl]azo]-1-(4-sulphophenyl)-1H-pyrazole-3-carboxylic acid, sodium salt

Cas Number:

85567-10-8

Molecular formula:

C18H13N4Na3O12S3

IUPAC Name:

trisodium 5-oxo-4-(2-{4-[2-(sulfonatooxy)ethanesulfonyl]phenyl}diazen-1-yl)-1-(4-sulfonatophenyl)-4,5-dihydro-1H-pyrazole-3-carboxylate

Test material form:

solid

Test animals

Species:

rat

Strain:

other: WISTAR Crl: WI(Han)

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: 8-12 weeks old
- Weight at day of admnistration: step 1: 165 – 198 g; step 2: 156 – 173 g
- Fasting period before study: Prior to the administration food was withheld from the test animals for 16 to 19 hours (access to water was permitted). Following the period of fasting the animals were weighed and the test item was administered. Food was provided again approximately 4 hours post dosing.
- Housing: The animals were kept in groups in IVC cages, type III H, polysulphone cages on Altromin saw fibre bedding
- Diet (e.g. ad libitum): Free access to Altromin 1324 maintenance diet for rats and mice
- Water (e.g. ad libitum): Free access to tap water, sulphur acidified to a pH value of approximately 2.8 (drinking water, municipal residue control, microbiological controls at regular intervals)
- Acclimation period: Adequate acclimatisation period (at least five days) under laboratory conditions

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3 °C
- Humidity (%): 55 ± 10%
- Air changes (per hr): 10 x / hour
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES:
From: Acclimatisation Period:
Step 1, animals no. 1 - 3: 17 November 2016 to 29 November 2016
Step 2, animals no. 4 - 6: 29 November 2016 to 05 December 2016

To: 20 December 2016

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: aqua ad injectionem

Details on oral exposure:

VEHICLE
- Amount of vehicle (if gavage): 2 g of the test item was dissolved in the vehicle to gain a final volume of 10 mL and to achieve a dose of 2000 mg/kg body weight at a dose volume of 10 mL/kg body weight.
- Justification for choice of vehicle: In order to get the test item in a solution or suspension, which is applicable to the animals, aqua ad injectionem was evaluated as vehicle and was considered to be adequate. This vehicle was chosen due to its non-toxic characteristics.
- Lot/batch no. (if required): AlleMan Pharma, lot no. 605070, expiry date: 04/2019

MAXIMUM DOSE VOLUME APPLIED: The test item was administered at a dose volume of 10 mL/kg body weight.

DOSAGE PREPARATION (if unusual): The test item was weighed out into a tared plastic vial on a precision balance.
Homogeneity of the test item in the vehicle was maintained by vortexing the prepared suspension thoroughly before each dose administration.
For all animals of the first and second step, 2 g of the test item was dissolved in the vehicle to gain a final volume of 10 mL and to achieve a dose of 2000 mg/kg body weight at a dose volume of 10 mL/kg body weight.
The dose formulations were made shortly before each dosing occasion.

CLASS METHOD (if applicable)
- Rationale for the selection of the starting dose: The starting dose was selected to be 2000 mg/kg body weight. According to OECD 423 guideline, when available information suggests that mortality is unlikely at highest starting dose level (2000 mg/kg), then a limit test should be conducted. The provided safety data sheet of the sponsor indicated no toxicity at the highest starting dose level.

Doses:

2000 mg/kg bw

No. of animals per sex per dose:

3 per step / 2 steps performed

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:
All animals were observed for 14 days after dosing for general clinical signs, morbidity and mortality. A careful clinical examination was made several times on the day of dosing (at least once during the first 30 minutes and with special attention given during the first 4 hours post-dose). Thereafter, the animals were observed for clinical signs once daily until the end of the observation period.
Cageside observations included changes in the skin and fur, eyes and mucous membranes. Also respiratory, circulatory, autonomic and central nervous systems and somatomotor activity and behaviour pattern were examined. Particular attention was directed to observations of tremor, convulsions, salivation, diarrhoea, lethargy, sleep and coma.
The animals were weighed on day 1 (prior to the administration) and on days 8 and 15.
- Necropsy of survivors performed: yes
- Other examinations performed: clinical signs, body weighgross pathological changes. In the absence of gross pathological changes no tissues were preserved for a possible histopathological evaluation.

Results and discussion

Effect levels

Key result

Sex:

female

Dose descriptor:

LD50 cut-off

Based on:

test mat.

Remarks on result:

other: no mortality at limit dose of 2000 mg/kg bw

Mortality:

The test item showed no mortality after a single dose administration.

Clinical signs:

other: The test item showed no mortality and no other acute oral toxicity characteristics after a single dose administration.

Gross pathology:

With the exception of acute injection of blood vessels in the abdominal region, which is due to the euthanasia injection, no specific gross pathological changes were recorded for any animal.

Applicant's summary and conclusion

Interpretation of results:

other: No classification required according to regulaation (EC) No. 1272/2008

Conclusions:

Under the conditions of the present study, a single oral application of the test item to rats at a dose of 2000 mg/kg body weight was associated with no signs of toxicity or mortality.
The median lethal dose of the test item after a single oral administration to female rats, observed over a period of 14 days is: LD50 cut-off (rat): ∞
According to Annex I of Regulation (EC) 1272/2008 the test item has no obligatory labelling requirement for toxicity and is not classified.

Executive summary:

Two groups, each of three female WISTAR Crl: WI(Han) rats, were treated with the test item by oral gavage administration at a dosage of 2000 mg/kg body weight (limit test). The test item was dissolved in aqua ad injectionem (sterile water) at a concentration of 0.2 g/mL and administered at a dose volume of 10 mL/kg.

All animals used in the study after their entrance at BSL were allowed to acclimatise to the laboratory conditions for at least 5 days. The animals were observed on delivery, on inclusion in the study and before administration for mortality/morbidity and other clinical signs. All animals were examined for clinical signs several times on the day of dosing and once daily until the end of the observation period. Their body weights were recorded on day 1 (prior to the administration) and on days 8 and 15. All animals were necropsied and examined macroscopically.

All animals survived until the end of the study without showing any test-item related signs of toxicity.

Throughout the 14-day observation period, the body weight gain of the test animals was within the normal range of variation for this strain.

At necropsy, no treatment-related macroscopic findings were observed in any animal of any step.

On the basis of the test results given below and in conformity with the criteria given in Annex I of Regulation (EC) 1272/2008, the substance should not be classified.