Reaction products of diazotised 5,5’-methylenedianthranilic acid, coupled with resorcinol, subsequently coupled with diazotised 4-aminobenzene-1,3-disulfonic acid, chelated with copper sulphate, sodium salt

Endpoint summary

Description of key information

LD50 > 2000 mg/kg bw

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

other: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

From September 29 to October 11, 2016

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Reliability of the source study is 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2001

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2008

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Species:

rat

Strain:

Wistar

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Species: laboratory albino rat (it is the preferred rodent species according to the guideline)
- Supplier: SPF breeding, VELAZ s.r.o.
- Age: 8 weeks at the time application
- Acclimatisation: 12 days
- Housing: animal room with monitoring conditions – 3 animals of one sex in one plastic breeding cage
- Diet: pelleted standard diet for experimental animals ad libitum
- Water: drinking tap water ad libitum (quality corresponding to Regulation No. 252/2004 Czech Coll. of Law)

ENVIRONMENTAL CONDITIONS
- Temperature: room temperature 22 ± 3 °C, permanently monitored
- Relative humidity: 30 – 70 %, permanently monitored
- Photoperiod: 12-hour light/12 hour dark
- Bedding: shavings of soft wood

- Randomisation: according to internal SOP No.42
- Identification of animals: colour marks 1 - 3 on tail of animals, each cage was marked with the number of study, sex and dose of the test substance
- Health condition: certificate of good health condition – from breeding farm; no signs of diseases were observed at clinical check-in, during the acclimatisation period and before the start of study.

Route of administration:

oral: gavage

Vehicle:

water

Details on oral exposure:

VEHICLE
- Lot/batch no.: 1605100264

MAXIMUM DOSE VOLUME APPLIED:
1 mL/100 g of animal body weight

Doses:

2000 mg/kg of body weight

No. of animals per sex per dose:

Animal per sex per dose: 3
Total number of animals: 6

Control animals:

not specified

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing:
- Body weight: before application, 8th day and before euthanasia of animals
- Mortality: dailyClinical examination: daily
- Pathological examination: 15th day
- Necropsy of survivors performed: yes
- Other examinations performed: after application the animals were observed individually:
* the first day: twice (30 minutes and 3 hours after application)
* the second day: twice (in the morning and in the afternoon) and daily thereafter for 14 days.
Observations included changes in skin and fur, eyes, visible mucous membranes, behaviour of animals, somatomotor activity, reactions to stimuli, and presence of lacrimation, salivation and discharge from nostrils, function of respiratory, digestive and urogenital system. The results of the observations were recorded on special data sheets. All test animals survived to the end of study were sacrificed on the 15th day and gross necropsy was carried out. Nutritious status, body surface, body foramina, thoracic, abdominal and cranial cavity were evaluated. All gross macroscopic changes of organs and tissues were recorded on special data sheets.

Sex:

female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

test mat.

Mortality:

No mortality observed

Clinical signs:

other: Changes of colour of faeces associated with the colour of the test substance were observed first and second day during the clinical observation after the application. Soft faeces were observed on the day of the application. No other clinical signs of into

Gross pathology:

No pathologic macroscopic changes were diagnosed during pathological examination

Preparation of experimental animals

About twenty hours before oral administration the animals were not fed, drinking water was given ad libitum. Immediately before application the animals were weighed and distributed to groups with 3 animals. The feed was given to animals 3 hours after application of the test substance.

Course of study

The dose level of 2000 mg/kg was used as the starting dose, because no toxicity of the test substance was estimated on the basis of information about testing of similar substances. Because this dose caused no death of females, the same dose of 2000 mg/kg level was sequentially applied for confirmation (application with time distance 24 hours) to group of 3 females. No death of animals was observed in this group of 3 females.

Testing schedule (according to OECD Test Guideline No. 423: Acute Oral Toxicity – Acute Toxic Class Method, Adopted 17th December 2001)

START: 2000 mg/kg – 3 females (Step No. 1) – no deaths ►2000 mg/kg – 3 females (Step No. 2): no deaths ► END of study

Body weight recording

Animals were weighed before application, at the 8th day of study and at the 15th day, before euthanasia of animals. Average body weight in a group was calculated from individual body weights. Body weight increments were calculated from body weight at the start of the study, the first week and at the end of the study.

Interpretation of results:

other: Not classified according to the CLP Regulation

Conclusions:

LD50 > 2000 mg/kg bw

Executive summary:

Method

The test was performed according EU Method B.1 tris: Acute Oral Toxicity - Acute Toxic Class Method. 

The test substance was administered in a single dose as a suspension in vehicle (water), given orally via gavage to female Wistar rats. The volume of administered suspension was 1 ml/100 g body weight of animals. 

The dose level of 2000 mg/kg bw was used as the starting dose, because no toxicity of the test substance was estimated on the basis of the information about testing of similar substances.

The dosing was performed sequentially in two groups of three females: group No. 1 - using the starting dose of 2000 mg/kg of body weight. No death of females caused this dose, dose of 2000 mg/kg of body weight was sequentially applied for confirmation to group No. 2.

 

Observation

The test substance administered at the dose of 2000 mg/kg bw caused no death of animals. Only changes of colour of faeces associated with the colour of the test substance were observed first and second day during the clinical observation after the application. Soft faeces were observed only on the day of the application. No other clinical signs of intoxication were detected during the whole study in all sixanimals administered by the dose 2000 mg/kg/body weight. No pathologic macroscopic changes were diagnosed during pathological examination.

 

Conclusion

LD50 > 2000 mg/kg bw

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Acute toxicity: via inhalation route

Endpoint conclusion

Endpoint conclusion:

no study available

Acute toxicity: via dermal route

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

According to the CLP Regulation (EC 1272/2008), substances can be allocated to one of four toxicity categories based on acute toxicity by the oral, dermal or inhalation route according to the numeric criteria. Acute toxicity values are expressed as (approximate) LD50 (oral, dermal) or LC50 (inhalation) values or as acute toxicity estimates (ATE).

In the case of oral exposure route, the acute toxicity hazard categories and acute toxicity estimates (ATE) defining the respective categories are:

- category 1: ATE ≤ 5 mg/kg bw

- category 2: 5 < ATE ≤ 50 mg/kg bw

- category 3: 50 < ATE ≤ 300 mg/kg bw

- category 4: 300 < ATE ≤ 2000 mg/kg bw

The substance shall not be classified for Oral toxicity if the LD50 resulted in animal test is > 2000 mg/kg (mg/kg bodyweight). According to the CLP criteria n.1272/2008, the test substance is not classified as toxic for the oral exposure as its LD50 is > 2000 mg/kg bw.