Calcium 4,5-dichloro-2-[[4,5-dihydro-3-methyl-5-oxo-1-(3-sulphonatophenyl)-1H-pyrazol-4-yl]azo]benzenesulphonate

Endpoint summary

Description of key information

Three studies were performed to evaluate acute oral, dermal and inhalative toxicity of the test substance to the

rat (according OECD 401, 402, 403). Neither the test substance nor structural analogues induce mortalities, abnormalities or clinical signs when applied oral or dermal. Also single administration via respiratory system did not cause health effects or mortalities. The LD50 for oral and dermal toxicity is considered to be > 2000 mg/kg bw, LC50 is > 5.5 mg/l air.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

(mostly due to reduced reporting in times before GLP), limited data to test substance

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 401 (Acute Oral Toxicity)

Deviations:

yes

Remarks:

The application volume was not calculated by the individual weight, but by the mean weight per sex

Principles of method if other than guideline:

Standardized test method (BASF-Test)

GLP compliance:

no

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

Acclimatization in the animal care unit for at least 1 week.
Age of the animals at the beginning of the study: about 12 weeks
Type of cage: Stainless steel wire mesh cages, type DK-III (Becker & Co., Castrop-Rauxel, FRG)
No. of animals per cage: 5
Animal identification: Identification of groups (5 animals) using cage cards
Room temperature: 20 - 26°C
Humidity: 45 - 75%
Day/night rhythm: 12 h/12 h (6.00 - 18.00 hours/18.00 - 6.00 hours)
Drinking water: Demineralized water each workday, tap water on holidays; ad libitum
Diet: Ssniff R; SSNIFF, Versuchstierdiaeten, Soest, Germany
Fasting period: The animals are given no feed 16 hours before administration, but water is available ad libitum.

Route of administration:

oral: gavage

Vehicle:

other: 0.5% aqueous carboxymethyl cellulose and 1 - 2 drops of Cremophor EL

Details on oral exposure:

Aqueous formulation corresponds to the physiological medium.
Form of administration: suspension

Doses:

5000 mg/kg

No. of animals per sex per dose:

5

Control animals:

no

Details on study design:

Duration of observation period following administration: 14 days
Weight check: Animals of comparable weight (+/- 10 g) in one cage; weighing of groups before administration, 2nd weighing on the 3rd day, 3rd weighing on the 7th day, 4th weighing on the 13th day after administration.
Signs and symptoms: Recording of signs and symptoms < 15 min, 15 min, 30 min, 1 h, 2 h, 4 h and 5 h after administration of the test substance and then once each workday. Check for moribund and dead animals twice each workday and once on holidays.
Pathology: Withdrawal of food 16 hours before sacrifice with C02; then necropsy with gross-pathological examination. Necropsy of all animals that die as early as possible.

Statistics:

-

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 5 000 mg/kg bw

Based on:

test mat.

Remarks on result:

other: No mortality, no toxicity observed.

Mortality:

none

Clinical signs:

other: 1 day after treatment: yellow feces

Gross pathology:

Sacrificed animals: organs: no abnormalities detected.

Interpretation of results:

GHS criteria not met

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

5 000 mg/kg bw

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

limited data to test substance

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 403 (Acute Inhalation Toxicity)

Deviations:

yes

Remarks:

No analytical purity; gravimetric analyses of the concentration without analytical confirmation of the composition and reanalysis of the airborne material, respectively; single instead of twice particle size distribution determination.

GLP compliance:

no

Test type:

standard acute method

Limit test:

yes

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

Mean body weight at the beginning of the test: male animals: 307 +/- 16 g, female animals 228 +/- 17 g.
Age at the beginning of the test: about 8 -10 weeks.
Identification: The animals were identified by toe amputation.
The animals were offered SSNIFF R complete diet for rats and mice, manufacturer: SSNIFF-Versuchstierdiaeten GmbH, Soest, Germany, and tap water ad libitum during the post-exposure observation period.
The animals were accommodated in fully air-conditioned rooms (required temperature 22 + 20°C, required humidity 55 + 5%) with a light/dark rhythm of 12 hours. They were housed in groups of five in cages of Becker, type D III, without bedding.

Route of administration:

inhalation: mixture of vapour and aerosol / mist

Type of inhalation exposure:

nose/head only

Vehicle:

air

Details on inhalation exposure:

Head-nose inhalation system INA 20 (glass/steel construction, BASF Aktiengesellschaft, V ca. 55 l); the animals are restrained in tubes and their snouts project into the inhalation chamber.
A mixture of dust and air was generated by means of a vibration aerosol generator (fluidized bed).
By means of a dust generator the substance to be tested was generated into a dust aerosol, which was passed into the inhalation system.
A vibration aerosol generator was used for generating dust. The concentration was adjusted by varying the amplitude and frequency of the vibrator.
The rate of flow was adjusted as follows: 1500 l/h of compressed air through the vibrator.
The inhalation mixture was offered to the animals for inhalation for 4 hours.
By means of an exhaust air system the pressure ratios in the inhalation chamber were adjusted in such a way that the amount of fresh air was about 10% lower. This ensured that the mixture of test substance and air was not diluted by laboratory air in the breathing zones of the animals.

Analytical verification of test atmosphere concentrations:

yes

Duration of exposure:

4 h

Concentrations:

5.5 mg/l

No. of animals per sex per dose:

10

Control animals:

yes

Details on study design:

- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: before starting the study, after 7 days and after the end of the observation period
- Necropsy of survivors performed: yes

Statistics:

The statistical evaluation of the concentration-response relationship was carried out in accordance with the binominal test (Wittig, H.: Mathematische Statistik 1974, pp. 32 - 35) according to tables of the BASF Computer Center.
The particle size was determined in the Department of Toxicology of BASF Aktiengesellschaft in accordance with mathematical and graphical methods of evaluating particle measurements (Silverman, L.: Particle Size Analysis in Industrial Hygiene, 1971, pp. 235-259).

Sex:

male/female

Dose descriptor:

LC50

Effect level:

> 5.5 mg/L air

Based on:

test mat.

Exp. duration:

4 h

Remarks on result:

other: No mortality.

Mortality:

none

Clinical signs:

other: During exposure: substance-colored fur in the region of the head. After exposure: substance-colored fur, partly agitated behavior, after 1 day nothing abnormal detected.

Body weight:

Mean body weight:
Test animals; before start of the study: 307 g (males), 228 g (females); after 7 days: 341 g (males), 241 g (females); after 14 days: 370 g (males), 252 g (females)
Control animals: before start of the study 307 g (males), 230 g (females); after 7 days 340 g (males), 240 g (females); after 14 days 370 g (males), 249 g (females)
The body weight of the animals showed no adverse effects in comparison with that of the control.

Gross pathology:

Sacrificed animals: no abnormalities detected.

Interpretation of results:

GHS criteria not met

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LC50

Value:

5.5 mg/m³ air

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Type of information:

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

Adequacy of study:

key study

Reason / purpose for cross-reference:

read-across source

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 2 000 mg/kg bw

Based on:

other: Read-across

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

LD50

Value:

2 000 mg/kg bw

Additional information

Procedure and observations

 

To evaluate the acute oral toxicity, a single dose (5000 mg/kg bw) of the test article was administrated to a group of 5 male and 5 female rats by oral gavage (BASF AG 1981a). Following dosing, the animals were observed for 14d. Examination of clinical signs and viability were performed daily, weighing on day 1, 3, 7 and 13 after treatment. There were no deaths as a result of treatment with the test article. Clinical signs of toxicity or changes in body weight gain were not observed during the observation period. Gross necropsy was without any findings.

 

To evaluate the acute inhalative toxicity, male and female rats (10/sex/dose) were exposed for 4h to a dust aerosol (5.5 mg/l air, 1.500 l/h, nose only) of the test item (BASF AG 1982a). Following dosing, the animals were observed for 14d. Examination of clinical signs and viability were performed daily, weighing on day 1, 3, 7 and 13 after treatment. There were no deaths as a result of treatment with the test article. Clinical signs of toxicity or changes in body weight gain were not observed during the observation period. Gross necropsy was without any findings.

read across EC 416-730-1

The test item was not tested for acute dermal toxicity. It is acceptable to derive the dermal toxicity from experimental data of a structural analogue (di-ammonium salt) since both are salts with comparable structures. In addition, lower solubility and the higher molecular weight of the test item give a safety margin (a detailed read across justification is given in CSR, Annex I)

Dermal toxicity of an analogue the di-ammonium salt was evaluated on a group of five male and 5 female rats which were treated with the test article at 2000 mg/kg by dermal application (Ciba 1994a). The substance was suspended in water at a concentration of 0.5 g/ml and administered at a volume of 4 ml/kg. Four times during day 1 and once daily during days 2-15 the animals were examined for clinical signs and viability. Body weights were recorded on day 1 before administration and on days 8 and 15. All animals were necropsied and examined macroscopically. No deaths occurred during the study period. Neither clinical signs of systemic toxicity nor local effects of the test article on the skin at the application site were observed during the observation period. After treatment, a residual yellow staining on skin was seen. A slightly lower bodyweight gain was recorded for one male and one female on Day 8 and in three males on Day 15. All other rats achieved the anticipated bodyweight gains throughout the study. Macroscopic organ findings were not observed at necropsy.

In addition to the regular acute toxicity testing, toxicity after intraperitoneal application according an internal guideline was performed (BASF AG 1981b). 5 male and female rats received 2000 mg/kg bw (10 ml/kg bw) of the test item dissolved in 0.5% CMC plus castor oil by intrapritoneal injection. The animals were observed for 14d post treatment and were weighted on days 2, 7 and 13. Apathy and dyspnea were observed after treatment and animals were of poor general state. Mortalities did not occur. Gross necropsy revealed intraabdominal incorporations of the test substance

 

Discussion

Application of the test substance via oral or inhalative route did not induce any signs of toxicity. None of the animals died, viability and bodyweight gain were unaffected by the test article. Also dermal application of the di-ammonium salt (structural analogue) did not induce any symptoms or mortalities. Intraperitoneal injection of the test item resulted in a poor general state and symptoms commonly seen after intraperitoneal administration. The effects are, therefore, regarded as not-treatment related. In addition, from todays perspective the study does not meet basic scientific criteria and toxicological relevance.

Justification for classification or non-classification

 Classification, Labelling, and Packaging Regulation (EC) No. 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation 1272/2008. As a result the substance is not considered to be classified for acute toxicity under Regulation (EC) No. 1272/2008, as amended for the thirteenth time in Commission Regulation (EU) 2018/1480 of Oct 4 2018.