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Administrative data

Endpoint:
acute toxicity: oral
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016-08-29 to 2016-10-06
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
Version / remarks:
2001-12-17
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
Version / remarks:
(EC) No 440/2008, 2008-05_30
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OPPTS 870.1100 (Acute Oral Toxicity)
Version / remarks:
2002-12
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: Japan MAFF Testing Guideline of 12 Nousan No. 8147
Version / remarks:
2000-11-24
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test type:
acute toxic class method
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Reaction mass of sodium [2,4-dihydro-4-[(2-hydroxy-5-nitrophenyl)azo]-5-methyl-2-phenyl-3H-pyrazol-3-onato(2-)][1-[(2-hydroxy-5-nitrophenyl)azo]-2-naphtholato(2-)]chromate(1-) and sodium bis[1-[(2-hydroxy-5-nitrophenyl)azo]-2-naphtholato(2-)]chromate(1-) and sodium bis[2,4-dihydro-4-[(2-hydroxy-5-nitrophenyl)azo]-5-methyl-2-phenyl-3H-pyrazol-3-onato(2-)]chromate(1-)
EC Number:
915-758-6
Molecular formula:
C32H18CrN6O8.Na / C32H22CrN10O8.Na / C32H20CrN8O8.Na
IUPAC Name:
Reaction mass of sodium [2,4-dihydro-4-[(2-hydroxy-5-nitrophenyl)azo]-5-methyl-2-phenyl-3H-pyrazol-3-onato(2-)][1-[(2-hydroxy-5-nitrophenyl)azo]-2-naphtholato(2-)]chromate(1-) and sodium bis[1-[(2-hydroxy-5-nitrophenyl)azo]-2-naphtholato(2-)]chromate(1-) and sodium bis[2,4-dihydro-4-[(2-hydroxy-5-nitrophenyl)azo]-5-methyl-2-phenyl-3H-pyrazol-3-onato(2-)]chromate(1-)
Test material form:
solid
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Batch No. of test material: N01-131001
- Expiration date of the lot/batch: 2023-03-15

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: Room temperature

OTHER SPECIFICS: Solid / dark brown

Test animals

Species:
rat
Strain:
Wistar
Remarks:
Crl:WI (Han) SPF
Sex:
female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Wiga GmbH, Germany
- Females nulliparous and non-pregnant: yes
- Age at study initiation: ~ 10 weeks
- Weight at study initiation: 173 - 190 g
- Fasting period before study: at least 16 hours before administration, but water was available ad libitum.
- Housing: Single housing, in fully air-conditioned rooms, Makrolon cage, type III
- Diet: VRF1(P); SDS Special Diets Services, 67122 Altrip, Germany, ad libitum
- Water: Tap water, ad libitum
- Acclimation period: least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 30 - 70
- Air changes (per hr): ~ 10
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Remarks:
Ph.Eur.
Details on oral exposure:
VEHICLE:
- Concentration in vehicle: 20 g/100 mL (2000 mg/kg bw), 3 g/100 mL (300 mg/kg bw)
- Amount of vehicle: 10 mL/kg bw
- Justification for choice of vehicle: Good homogeneity of the test substance in corn oil Ph.Eur.

MAXIMUM DOSE VOLUME APPLIED:
10 mL/kg bw

DOSAGE PREPARATION:
The test item preparation for each test group was produced shortly before administration by stirring with a high speed homogenizer (Ultra-Turrax) and a magnetic stirrer.

CLASS METHOD:
Rationale for the selection of the starting dose: request of the sponsor
Doses:
1000 and 300 mg/kg bw
No. of animals per sex per dose:
3 females/dose
Control animals:
no
Details on study design:
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Clinical signs for each animal were recorded several times on the day of administration and at least once during each workday thereafter. Individual body weights shortly before administration (day 0), weekly thereafter and on the last day of observation. In addition on the day of death starting with study day 1. A check for any dead or moribund animals was made at least once each workday.
- Necropsy of survivors performed: yes, necropsy with gross pathological examination was performed on the last day of the observation period after sacrifice by CO2-inhalation in a chamber with gradually increasing concentrations. Necropsy of all animals that died was performed as early as possible after death.
Statistics:
Calculations were performed using Microsoft Excel 2010 and checked with a calculator.

Results and discussion

Effect levels
Key result
Sex:
female
Dose descriptor:
LD50
Effect level:
> 300 - < 2 000 mg/kg bw
Based on:
test mat.
Mortality:
In the single 2000 mg/kg bw test group all animals died at hour 5, day 1 or day 2 after administration, respectively.
No mortality occurred in both 300 mg/kg test groups.
Clinical signs:
Clinical signs in the single 2000 mg/kg test group revealed in all animals an impaired general state and piloerection at hour 4 or from hour 4 until hour 5 after administration. Additionally, one animal showed dyspnea at hour 4, while another animal showed a dark discolored skin on day 2.
Clinical signs in the first 300 mg/kg bw test group revealed in all animals an impaired general state and piloerection from hour 3 until hour 4 after administration. In addition, a dark discolored skin was seen in these animals from day 1 until day 3.
No clinical signs were observed in the second 300 mg/kg bw test group during clinical examination.
Body weight:
The body weights of the surviving animals increased within the normal range throughout the study period with one exception in the first 300 mg/kg bw test group. This animal gained weight in a normal range during the first week, but showed a slight weight gain during the second week. This effect is observed at times in the rat strain used, because in the required age range the female animals have already reached the phase of slow growth.
Gross pathology:
The following pathological findings were observed in the animals that died (2000 mg/kg bw test group, 3 females):
- Blackish discoloration of the stomach contents in all animals
- Gassed stomach in one animal
- Dark/blackish discolored liver in all animals
- Blackish discoloration of the small intestine's contents in one animal
There were no pathological findings in the animals sacrificed at the end of the observation period (6 females; both 300 mg/kg bw test groups).

Applicant's summary and conclusion

Interpretation of results:
Category 4 based on GHS criteria