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EC number: 201-663-0 | CAS number: 86-30-6
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Endpoint summary
Administrative data
Description of key information
Acute toxicity studies were performed on n-nitrosodiphenylamine by oral and dermal route.
Based on the results, the LD50 for both routes are higher than 2000 mg/kg/day.
Key value for chemical safety assessment
Acute toxicity: via oral route
Link to relevant study records
- Endpoint:
- acute toxicity: oral
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 08 August 2017 to 25 August 2017
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)
- Version / remarks:
- OECD Guidelines for Testing of Chemicals No. 423. Acute Oral Toxicity – Acute Toxic Class Method. Adopted: 17 December 2001
- Deviations:
- yes
- Remarks:
- Due to technical reason temperature values (maximum of 25.6°C) out of the target range 19-25°C were observed, however these minor differences in the environmental parameter were considered not to adversely affect the study.
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)
- Version / remarks:
- Commission Regulation (EC) No 440/2008 of 30 May 2008, B.1.Tris
- Deviations:
- yes
- Remarks:
- Due to technical reason temperature values (maximum of 25.6°C) out of the target range 19-25°C were observed, however these minor differences in the environmental parameter were considered not to adversely affect the study.
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 870.1100 (Acute Oral Toxicity)
- Version / remarks:
- EPA Health Effects Test Guidelines (OPPTS 870.1100), United States, EPA 712-C-02-190 (2002)
- Deviations:
- yes
- Remarks:
- Due to technical reason temperature values (maximum of 25.6°C) out of the target range 19-25°C were observed, however these minor differences in the environmental parameter were considered not to adversely affect the study.
- GLP compliance:
- yes (incl. QA statement)
- Test type:
- acute toxic class method
- Limit test:
- yes
- Species:
- rat
- Strain:
- Wistar
- Sex:
- female
- Details on test animals or test system and environmental conditions:
- Species and strain: Crl:WI Wistar rats
Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, D- 97633 Sulzfeld
Hygienic level at arrival: SPF
Hygienic level during the study: Standard housing conditions
Number of animals: 6 animals, 3 animals / group
Sex: Female, nulliparous and non-pregnant.
Age of animals at dosing: Young healthy adult rats, 8 weeks old
Body weight at treatment: 184 – 198 g
Acclimation period: at least 5 days
Husbandry
Animal health: Only healthy animals were used for the test. The staff Veterinarian certified their health status.
Number of animal room: 522/2
Housing: 3 animals / cage
Cage type: Type II polypropylene/polycarbonate
Bedding: LIGNOCEL ¾ S certified wooden chips (J. Rettenmaier & Söhne GmbH + Co. KG 73494, Holzmühle, Rosenberg, Germany) was available to animals during the study.
Nesting: ARBOCEL crinklets natural nest building material (J. Rettenmaier & Söhne GmbH + Co. KG, 73494, Holzmühle, Rosenberg, Germany) was available to animals during the study.
Lighting period: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 21.6 – 25.6°C
Relative humidity: 35 – 68%
Ventilation: 15 – 20 air exchanges/hour
Enrichment: Animals were housed by group to allow social interaction and with deep wood sawdust bedding to allow digging and other normal rodent activities.
Food and Water Supply
Animals received ssniff® SM R/M "Autoclavable complete diet for rats and mice – breeding and maintenance" produced by ssniff Spezialdiäten GmbH, D-59494 Soest,
Germany (Batch no.: 262 21592, expiry date: 31 January 2018), ad libitum, except for the night before treatment. The food is considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study. Animals received tap water from the municipal supply from 500 mL bottles, ad libitum. The water was fit for human consumption and was considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study.
Animal Identification
Animals were individually identified using numbers written on the tail with an indelible marker pen. The numbers were given on the basis of Citoxlab Hungary Ltd.'s Master File, for each animal allocated to the treatment groups. The cages were identified by cards, with information about study code, sex, dose group, cage number and individual animal numbers. - Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on oral exposure:
- Formulation
The test item was freshly formulated at a concentration of 200 mg/mL in the vehicle, in the Pharmacy of Citoxlab Hungary Ltd. on the day of administration. The test item was dusted in order to enhance the formulation. Formulation container was magnetic stirred continuously up to the end of dose administration procedures.
Vehicle: Corn oil
Lot number: A0384372
Expiry date: 31 May 2019
Dose volume: 10 mL/kg bw
Produced by: Acros Organics, Belgium
The initial dose level was selected by the Study Director to be that which is most likely to produce mortality in some of the dosed animals. Based on the preliminary toxicological information in the SDS, limit dose of 2000 mg/kg bw was selected as a starting dose. - Doses:
- 2000 mg/kg bw
- No. of animals per sex per dose:
- 6 animals (3 per group)
- Control animals:
- no
- Details on study design:
- ADMINISTRATION OF THE TEST ITEM
Justification of the dose:
The initial dose level was selected by the Study Director to be that which is most likely to produce mortality in some of the dosed animals. Based on the preliminary toxicological information in the SDS, limit dose of 2000 mg/kg bw was selected as a starting dose.
Initially, 3 female animals were treated with N-NITROSODIPHENYLAMINE at dose level of 2000 mg/kg bw. No mortality was observed, therefore further 3 animals were treated at the dose level of 2000 mg/kg bw. As only 1 animal died in this second dose group, further testing was not required according to the test guidelines (OECD 423, Commission Regulation (EC) No 440/2008 of 30 May 2008, B.1.Tris).
Procedure
A single oral gavage administration was followed by a fourteen-day observation period in the surviving animals or until the day of death as applicable. On the night before treatment, the animals were fasted. The food but not water was withheld during an overnight period. Animals were weighed just before treatment. The test item was administered by oral gavage in the morning. The food was returned 3 hours after the treatment.
OBSERVATIONS
Clinical Observations
Clinical observations were performed on all animals at 30 minutes, 1, 2, 3, 4 and 6 hours after dosing and daily for 14 days thereafter in all surviving animals or until the day of death as applicable. Individual observations were performed on the skin, fur, eyes, mucous membranes, respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.
Body Weight Measurement
The body weight was recorded on the day before treatment (Day -1), on the day of the treatment (Day 0) and weekly thereafter or on the day of death.
NECROPSY
Macroscopic examination was performed on all animals. The animals were sacrificed by exsanguination under pentobarbital anaesthesia (Release 30% inj.; Lot No.: 106075, Expiry Date: 31 July 2018, Produced by: Wirtschaftsgenossenschaft deutscher Tierärzte eG; Siemensstr. 14, 30827 Garbsen, Germany). After examination of the external appearance, the cranial, thoracic and the abdominal cavities were opened and the organs and the tissues were observed. Macroscopic abnormalities were recorded. - Statistics:
- The method used was not intended to allow the calculation of a precise LD50 value.
- Key result
- Sex:
- female
- Dose descriptor:
- LD50
- Effect level:
- > 2 000 mg/kg bw
- Based on:
- test mat.
- Remarks on result:
- other: 1 animal died
- Mortality:
- N-NITROSODIPHENYLAMINE caused the death of 1/6 animals (Day 1, ID: 377) at a dose level of 2000 mg/kg bw.
- Clinical signs:
- At dose level of 2000 mg/kg bw, decreased activity (score 1 or 2, up to Day 2), hunched back (up to Day 2), incoordination (score 1 or 2, up to Day 1), piloerection (up to Day 2) in all animals and diuresis (between Day 2 and Day 4) in all surviving animals were observed.
From Day 5 all surviving animals were symptom-free. - Body weight:
- There were no effects on body weights or body weight gains that could be attributed to treatment with of N-NITROSODIPHENYLAMINE.
- Gross pathology:
- In the rat, dosed at 2000 mg/kg bw, that was found dead on Day 1, the lungs (all lobes) were collapsed and dark red (diffuse).
There was no evidence of the macroscopic observations in the surviving animals dosed at 2000 mg/kg bw and terminated on Day 14. - Other findings:
- No further findings reference in the study report.
- Interpretation of results:
- Category 5 based on GHS criteria
- Conclusions:
- Under the conditions of the study, the acute oral LD50 value of the test item N-NITROSODIPHENYLAMINE was found to be above 2000 mg/kg bw in female Crl:WI rats.
According to the GHS criteria, N-NITROSODIPHENYLAMINE can be ranked as "Category 5" for acute oral exposure. - Executive summary:
The single-dose oral toxicity of N-NITROSODIPHENYLAMINE was performed according to the acute toxic class method (OECD 423 and Commission Regulation (EC) No 440/2008 of 30 May 2008, B.1.Tris) in Crl:WI Wistar rats.
Two groups of 3 female Crl:WI rats were treated with the test item at a dose level of 2000 mg/kg bw (Group 1 and Group 2).
A single oral treatment was carried out by gavage for each animal after an overnight food withdrawal. Food was made available again 3 hours after the treatment. The test item was formulated in corn oil at a concentration of 200 mg/mL at a dose volume of 10 mL/kg bw.
Initially, three females (Group 1) were treated at a dose level of 2000 mg/kg bw. As no mortality was observed, a confirmatory group (Group 2) was treated at the same dose level and 1 animal died on Day 1 from this group. As no other mortality was observed in the confirmatory group, therefore no further testing was required according to OECD 423 and Commission Regulation (EC) NO 440/2008 of 30 May 2008, B.1.Tris.
Clinical observations were performed at 30 minutes, 1, 2, 3, 4 and 6 hours after dosing and daily for 14 days thereafter or until the day of death. Body weight was measured on Days -1, 0, 7 and 14 (before necropsy), or on the day of death. All animals were subjected to a necropsy and a macroscopic examination.
The results of the study were summarized as follows:
Mortality
N-NITROSODIPHENYLAMINE caused the death of 1/6 animals (Day 1, ID: 377) at a dose level of 2000 mg/kg bw.
Clinical Observations
At dose level of 2000 mg/kg bw, decreased activity (score 1 or 2, up to Day 2), hunched back (up to Day 2), incoordination (score 1 or 2, up to Day 1), piloerection (up to Day 2) in all animals and diuresis (between Day 2 and Day 4) in all surviving animals were observed.
From Day 5 all surviving animals were symptom-free.
Body Weight and Body Weight Gain
There were no effects on body weights or body weight gains that could be attributed to treatment with N-NITROSODIPHENYLAMINE.
Necropsy
In the rat, dosed at 2000 mg/kg bw, that was found dead on Day 1, the lungs (all lobes) were collapsed and dark red (diffuse).
There was no evidence of the macroscopic observations in the surviving animals dosed at 2000 mg/kg bw and terminated on Day 14.
Conclusion:
Under the conditions of the study, the acute oral LD50 value of the test item N-NITROSODIPHENYLAMINE was found to be above 2000 mg/kg bw in female Crl:WI rats.
Reference
CLINICAL OBSERVATIONS
DOSE LEVEL: 2000 mg/kg bw, treatment on Day 0 SEX: FEMALE
Cage No. |
Animal Number |
Observations |
Observation days |
Frequency |
||||||||||||
0 |
1 |
2 |
3 |
4 |
5 |
6 |
7-14 |
|||||||||
30’ |
1h |
2h |
3h |
4h |
6h |
|||||||||||
1 |
374 |
Symptom Free |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
10/20 |
Activity decreased |
1 |
1 |
1 |
1 |
1 |
1 |
1 |
1 |
- |
- |
- |
- |
- |
8/20 |
||
Hunched back |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
- |
7/20 |
||
Incoordination |
- |
- |
- |
1 |
1 |
2 |
1 |
- |
- |
- |
- |
- |
- |
4/20 |
||
Piloerection |
- |
- |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
- |
5/20 |
||
Diuresis |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
- |
- |
- |
3/20 |
||
375 |
Symptom free |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
10/20 |
|
Activity decreased |
1 |
1 |
1 |
1 |
1 |
2 |
1 |
- |
- |
- |
- |
- |
- |
7/20 |
||
Hunched back |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
8/20 |
||
Incoordination |
- |
- |
- |
1 |
2 |
2 |
1 |
- |
- |
- |
- |
- |
- |
4/20 |
||
Piloerection |
- |
- |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
- |
5/20 |
||
Diuresis |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
- |
- |
- |
3/20 |
||
376 |
Symptom free |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
10/20 |
|
Activity decreased |
1 |
1 |
1 |
1 |
1 |
2 |
1 |
- |
- |
- |
- |
- |
- |
7/20 |
||
Hunched back |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
8/20 |
||
Incoordination |
- |
- |
- |
1 |
1 |
2 |
1 |
- |
- |
- |
- |
- |
- |
4/20 |
||
Piloerection |
- |
- |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
- |
5/20 |
||
Diuresis |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
- |
- |
- |
3/20 |
||
2 |
377# |
Activity decreased |
- |
1 |
1 |
1 |
1 |
2 |
|
5/6 |
||||||
Hunched back |
+ |
+ |
+ |
+ |
+ |
+ |
6/6 |
|||||||||
Incoordination |
- |
- |
1 |
1 |
1 |
2 |
4/6 |
|||||||||
Piloerection |
- |
- |
+ |
+ |
+ |
+ |
4/6 |
|||||||||
Found dead |
- |
- |
- |
- |
- |
- |
+ |
|
- |
|||||||
378 |
Symptom free |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
10/20 |
|
Activity decreased |
- |
1 |
1 |
1 |
1 |
2 |
2 |
1 |
- |
- |
- |
- |
- |
7/20 |
||
Hunched back |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
8/20 |
||
Incoordination |
- |
- |
1 |
1 |
1 |
2 |
1 |
- |
- |
- |
- |
- |
- |
5/20 |
||
Piloerection |
- |
- |
- |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
5/20 |
||
Diuresis |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
- |
- |
- |
3/20 |
||
379 |
Symptom free |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
10/20 |
|
Activity decreased |
- |
1 |
1 |
1 |
1 |
1 |
1 |
1 |
- |
- |
- |
- |
- |
7/20 |
||
Hunched back |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
8/20 |
||
Incoordination |
- |
- |
1 |
1 |
1 |
1 |
1 |
- |
- |
- |
- |
- |
- |
5/20 |
||
Piloerection |
- |
- |
- |
+ |
+ |
+ |
+ |
+ |
- |
- |
- |
- |
- |
5/20 |
||
Diuresis |
- |
- |
- |
- |
- |
- |
- |
+ |
+ |
+ |
- |
- |
- |
3/20 |
Remarks: + = present - = absent
h = hour ‘ = minute
# = Found dead
Frequency of observations = number of occurrence of observations / total number of observations
Severities: 1 = Slight/Small/Few; 2 = Moderate/medium; 3 = Marked/Large/Many
BODY WEIGHT DATA
DOSE LEVEL: 2000 mg/kg bw, Treatment on Day 0 SEX: FEMALE
Cage No. |
Animal Number |
Body weight (g) Days |
Day/Body Weight (g) Death |
Body Weight Gain (g) |
||||||
-1 |
0 |
7 |
14 |
-1-0 |
0-7 |
7-14 |
-1-14 |
|||
1 |
374 |
200 |
186 |
220 |
247 |
- |
-14 |
34 |
27 |
47 |
375 |
214 |
194 |
227 |
248 |
- |
-20 |
33 |
21 |
34 |
|
376 |
201 |
184 |
216 |
243 |
- |
-17 |
32 |
27 |
42 |
|
2 |
377# |
212 |
198 |
- |
- |
1/187 |
-14 |
- |
- |
- |
378 |
211 |
198 |
229 |
240 |
- |
-13 |
31 |
11 |
29 |
|
379 |
202 |
192 |
223 |
235 |
- |
-10 |
31 |
12 |
33 |
|
Mean: |
206.7 |
192.0 |
223.0 |
242.6 |
- |
-14.7 |
32.2 |
19.6 |
37.0 |
|
Standard deviation: |
6.3 |
5.9 |
5.2 |
5.3 |
- |
3.4 |
1.3 |
7.8 |
7.3 |
- = No data
# = Found dead
NECROPSY FINDINGS
DOSE LEVEL: 2000 mg/kg bw, Treatment on Day 0 SEX: FEMALE
Cage No. |
Animal Number |
Necropsy Date/ Necropsy Day |
External Observations |
Internal Observations |
Organ/Tissue |
1 |
374 |
22 August 2017 Day 14 |
No external observations recorded |
No internal observations recorded |
Not applicable |
375 |
22 August 2017 Day 14 |
No external observations recorded |
No internal observations recorded |
Not applicable |
|
376 |
22 August 2017 Day 14 |
No external observations recorded |
No internal observations recorded |
Not applicable |
|
2 |
377# |
12 August 2017 Day 1 |
No external observations recorded |
Collapsed |
Lungs |
Discoloration, red, diffuse, all lobes |
|||||
378 |
25 August 2017 Day 14 |
No external observations recorded |
No internal observations recorded |
Not applicable |
|
379 |
25 August 2017 Day 14 |
No external observations recorded |
No internal observations recorded |
Not applicable |
# = Found dead
Endpoint conclusion
- Endpoint conclusion:
- adverse effect observed
- Dose descriptor:
- LD50
- Value:
- 2 000 mg/kg bw
- Quality of whole database:
- The study is considered as reliable (klimish score of 1).
Acute toxicity: via inhalation route
Endpoint conclusion
- Endpoint conclusion:
- no study available
Acute toxicity: via dermal route
Link to relevant study records
- Endpoint:
- acute toxicity: dermal
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 20 February 2018 to 08 March 2018
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 402 (Acute Dermal Toxicity)
- Version / remarks:
- OECD GUIDELINES FOR TESTING OF CHEMICALS (No.: 402, 09th Oct. 2017)
- Deviations:
- yes
- Remarks:
- Due to technical reason, relative humidity values (minimum of 28%) out of the target range 30-70% were observed, however these minor differences in the environmental parameter were considered not to adversely affect the study.
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.3 (Acute Toxicity (Dermal))
- Version / remarks:
- Commission Regulation (EC) No 440/2008, B.3 (L 142, 30 May 2008)
- Deviations:
- yes
- Remarks:
- Due to technical reason, relative humidity values (minimum of 28%) out of the target range 30-70% were observed, however these minor differences in the environmental parameter were considered not to adversely affect the study.
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 870.1200 (Acute Dermal Toxicity)
- Version / remarks:
- OPPTS 870.1200 (EPA 712-C-98-192, August 1998)
- Deviations:
- yes
- Remarks:
- Due to technical reason, relative humidity values (minimum of 28%) out of the target range 30-70% were observed, however these minor differences in the environmental parameter were considered not to adversely affect the study.
- GLP compliance:
- yes (incl. QA statement)
- Test type:
- standard acute method
- Limit test:
- yes
- Species:
- rat
- Strain:
- Wistar
- Sex:
- female
- Details on test animals or test system and environmental conditions:
- Species and strain: Crl:WI Wistar rats
Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, D-97633 Sulzfeld, Germany
Hygienic level at arrival: SPF
Hygienic level during the study: Standard housing conditions
Justification of strain: The Wistar rat is one of the standard rodent species used in acute toxicity studies.
Number of animals: 3 animals
Sex: Female, nulliparous and non-pregnant.
Age of animals at study start: ~9 weeks old
Body weight range at dosing: Between 239 g and 260 g
Acclimation time: 5 or 7 days
Husbandry
Animal health: Only healthy animals were used for the study. The staff Veterinarian certified the health status.
Room-Box: 242/3
Housing: Individual and group caging
Cage type: Type II. polypropylene/polycarbonate
Bedding: Lignocel 3/4-S Hygienic Animal Bedding (produced by J. Rettenmaier & Söhne GmbH+Co.KG, Germany) was available to animals during the study. The quality of the bedding was guaranteed by the supplier. Details of bedding quality are archived with the raw data.
Nesting: Nest building material Arbocel Crinklets natural (produced by J. Rettenmaier & Söhne GmbH+Co.KG, Germany) was available to animals during the study. The quality of the nest building material was guaranteed by the supplier. Details of nest building material quality are archived with the raw data.
Light: 12 hours daily, from 6.00 a.m. to 6.00 p.m.
Temperature: 19.0–24.2°C
Relative humidity: 28–45%
Ventilation: 15-20 air exchanges/hour
Enrichment: Rodents were housed with deep wood sawdust bedding to allow digging and other normal rodent activities.
Temperature and relative humidity were recorded twice daily during the study.
Food and Water Supply
Animals received ssniff® SM R/M "Autoclavable complete diet for rats and mice – breeding and maintenance" produced by ssniff Spezialdiäten GmbH, D-59494 Soest, Germany, ad libitum, and tap water from the municipal supply, as for human consumption from a 500 mL bottle, ad libitum. The food is considered not to contain any contaminants that could reasonably be expected to affect the purpose or integrity of the study and the water was considered fit for human consumption.
The supplier provided an analytical certificate for the batch used. A copy of the certificate will be archived with the raw data.
Water quality control analysis is performed once every three months and microbiological assessment is performed monthly by Veszprém County Institute of State Public Health and Medical Officer Service (ÁNTSZ, H-8201 Veszprém, József A.u.36., Hungary).
Animal Identification
Animals were individually identified using numbers written on the tail with an indelible pen. The numbers were given on the basis of Citoxlab Hungary Ltd.' s Master File for each animal allocated to the treatment groups. The cages were identified by cards containing information about study code, sex, dose group, cage number and individual animal number. - Type of coverage:
- semiocclusive
- Vehicle:
- water
- Remarks:
- to moisten area only.
- Details on dermal exposure:
- The back of each animal was shaved (approximately 10% area of the total body surface) approximately 24 hours prior to treatment. The test item was applied to the shaved skin as a single dose and remained in contact with the skin for the 24-hour exposure period. Sufficient amount water was used to moisten the test item to ensure good contact with the skin. Sterile gauze pads were placed on the skin of rats to cover the test item. These gauze pads were kept in contact with the skin using a patch with adhesive hypoallergenic plaster. The entire trunk of the animal was then wrapped with semi occlusive plastic wrap for 24 hours.
At the end of the exposure period, the treated area of skin with the test item was washed with water at body temperature. - Duration of exposure:
- 24-hour exposure period
- Doses:
- A single dermal application of 2000 mg/kg bw
- No. of animals per sex per dose:
- One female rat was dosed initially (sentinel animal) than 2 female animals were dosed 48 hours later.
- Control animals:
- no
- Details on study design:
- OBSERVATIONS
Clinical Observations
Clinical observations were performed on the day of treatment at 30 minutes, 1, 2 and 5 hours after application of the test item and once each day for 14 days thereafter. Observations included the skin and fur, eyes and mucous membranes, the respiratory, circulatory, autonomic and central nervous system, somatomotor activity and behaviour pattern. Particular attention was directed to observation of tremors, convulsions, salivation, diarrhoea, lethargy, sleep and coma.
Skin Irritation
Adverse skin reactions at the site of application were recorded daily following the removal of the dressing.
Measurement of Body Weight
The body weights were recorded on Day 0 (before the test item administration) and on Days 7 and 14 (before necropsy).
NECROPSY
Macroscopic examination was performed on all animals. All animals were anaesthetised with pentobarbital sodium (details in 3.3) and exsanguinated. Following confirmation of death, after examination of the external appearance, the cranial, thoracic and abdominal cavities were opened and the appearance of the tissues and organs was observed. All macroscopic changes were recorded. - Statistics:
- The method used was not intended to allow the calculation of a precise LD50 value.
Body weight and body weight gain are summarised in tabular form. Clinical signs and necropsy findings are described and summarised in tabular form. - Key result
- Sex:
- female
- Dose descriptor:
- LD0
- Effect level:
- > 2 000 mg/kg bw
- Based on:
- test mat.
- Remarks on result:
- other: no mortality occurred
- Mortality:
- The test item did not cause mortality at the dose level of 2000 mg/kg bw.
- Clinical signs:
- There were no systemic clinical signs noted in any animal throughout the study.
- Body weight:
- There was no treatment related effects on body weight or body weight gain during the observation period. Body weights were within the range commonly recorded for this strain and age.
- Gross pathology:
- There was no evidence of any gross macroscopic changes at a dose level of 2000 mg/kg bw.
- Other findings:
- Local Dermal Signs
No adverse local dermal signs were observed after treatment with the test item or during the 14 days observation period.
The test item coloured the treatment area to yellowish on Day 1 (3/3 animals) and Day 2 (2/3) animals. - Interpretation of results:
- GHS criteria not met
- Conclusions:
- The acute dermal median lethal dose (LD50) of the test item N-NITROSODIPHENYLAMINE was found to be greater than 2000 mg/kg body weight in female Crl:WI rats.
According to the GHS criteria, N-NITROSODIPHENYLAMINE can be ranked as "Unclassified" for acute dermal exposure. - Executive summary:
An acute dermal toxicity study was performed with the test item N-NITROSODIPHENYLAMINE in female Crl:WI Wistar rats, in compliance with OECD Guideline No. 402 (2017), Commission Regulation (EC) No 440/2008, B.3 and OPPTS 870.1200.
This study was being performed with vertebrate animals as no in vitro alternative is available. The Sponsor confirmed previously that the specific regulatory purpose of this study did not allow a waiving of this dermal acute study, taking account of the OECD guidance document 237.
The study has been designed such that the minimum number of animals were used. Single animal at dose level of 2000 mg/kg body weight (bw) was used in a range-finding phase in female rats, followed by two animals to confirm the expected non-lethal dose level. The test item was applied as a single dermal 24-hour exposure followed by a 14-day observation period.
Clinical observations were performed on all animals at 30 minutes, 1, 2, 5 hours after dosing and daily for 14 days thereafter. Body weight was measured on Day 0 (prior to dosing) and on Days 7 and 14 (before necropsy). Gross macroscopic examination was performed on all animals at necropsy at the end of the 2-week observation period (Day 14).
The results of the study were summarised as follows:
Mortality
Test item did not cause mortality at the dose level of 2000 mg/kg bw.
Systemic clinical signs
There were no systemic clinical signs noted in any animal throughout the study.
Local dermal signs
No adverse local dermal signs were observed after treatment with the test item or during the 14 days observation period.
The test item coloured the treatment area to yellowish on Day 1 (3/3 animals) and Day 2 (2/3) animals.
Body weight and body weight gain
There was no treatment related effects on body weight or body weight gain during the observation period. Body weights were within the range commonly recorded for this strain and age.
Necropsy
There was no evidence of any macroscopic changes at a dose level of 2000 mg/kg bw.
Conclusions
The acute dermal median lethal dose (LD50) of the test item N-NITROSODIPHENYLAMINE was found to be greater than 2000 mg/kg body weight in female Crl:WI rats.
According to the GHS criteria, N-NITROSODIPHENYLAMINE can be ranked as "Unclassified" for acute dermal exposure.
Reference
Clinical Observations
DOSE LEVEL: 2000 mg/kg bw SEX: FEMALE
Cage No. |
Animal No. |
Observations |
Observation days |
Frequency |
|||||||||||||||||
0 |
1 |
2 |
3 |
4 |
5 |
6 |
7 |
8 |
9 |
10 |
11 |
12 |
13 |
14 |
|||||||
30’ |
1h |
2h |
5h |
||||||||||||||||||
1 |
3126 |
Symptom Free |
+ |
+ |
+ |
+ |
- |
- |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
16/18 |
Skin colour (yellowish-treated area) |
- |
- |
- |
- |
+ |
+ |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
2/18 |
||
2 |
3127 |
Symptom Free |
+ |
+ |
+ |
+ |
- |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
17/18 |
Skin colour (yellowish-treated area) |
- |
- |
- |
- |
+ |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
1/18 |
||
3 |
3128 |
Symptom Free |
+ |
+ |
+ |
+ |
- |
- |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
+ |
16/18 |
Skin colour (yellowish-treated area) |
- |
- |
- |
- |
+ |
+ |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
- |
2/18 |
Remarks: + = present - = absent
h = hour Treatment day = Day 0
‘ = minute
Frequency of observation = number of occurrence of observation / total number of observations
Body Weight Data
DOSE LEVEL: 2000 mg/kg bw SEX: FEMALE
Cage No. |
Animal No. |
Body weight (g) Days |
Body Weight Gain (g) |
||||
0 |
7 |
14 |
0-7 |
7-14 |
0-14 |
||
1 |
3126 |
260 |
271 |
282 |
11 |
11 |
22 |
2 |
3127 |
247 |
279 |
294 |
32 |
15 |
47 |
3 |
3128 |
239 |
250 |
257 |
11 |
7 |
18 |
Mean: |
248.7 |
266.7 |
277.7 |
18.0 |
11.0 |
29.0 |
|
Standard deviation: |
10.6 |
15.0 |
18.9 |
1.1 |
4.0 |
15.7 |
Macroscopic Findings
DOSE LEVEL: 2000 mg/kg bw SEX: FEMALE
Cage No. |
Animal No. |
Necropsy Date / Necropsy Day |
External Observations |
Internal Observations |
Organ/Tissue |
1 |
3126 |
06 March 2018 Day 14 |
No external observations |
No internal observations |
Not applicable |
2 |
3127 |
08 March 2018 Day 14 |
No external observations |
No internal observations |
Not applicable |
3 |
3128 |
08 March 2018 Day 14 |
No external observations |
No internal observations |
Not applicable |
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed
- Dose descriptor:
- discriminating dose
- Value:
- 2 000 mg/kg bw
- Quality of whole database:
- The study is considered as reliable (klimish score of 1).
Additional information
Acute toxicity study by oral route (Tarcai 2018) :
The single-dose oral toxicity of n-nitrosodihenylamine was performed according to the acute toxic class method (OECD 423 and Commission Regulation (EC) No 440/2008 of 30 May 2008, B.1.Tris) in Crl:WI Wistar rats.
Two groups of 3 female Crl:WI rats were treated with the test item at a dose level of 2000 mg/kg bw (Group 1 and Group 2).
A single oral treatment was carried out by gavage for each animal after an overnight food withdrawal. Food was made available again 3 hours after the treatment. The test item was formulated in corn oil at a concentration of 200 mg/mL at a dose volume of 10 mL/kg bw.
Initially, three females (Group 1) were treated at a dose level of 2000 mg/kg bw. As no mortality was observed, a confirmatory group (Group 2) was treated at the same dose level and 1 animal died on Day 1 from this group. As no other mortality was observed in the confirmatory group, therefore no further testing was required according to OECD 423 and Commission Regulation (EC) NO 440/2008 of 30 May 2008, B.1.Tris.
Clinical observations were performed at 30 minutes, 1, 2, 3, 4 and 6 hours after dosing and daily for 14 days thereafter or until the day of death. Body weight was measured on Days -1, 0, 7 and 14 (before necropsy), or on the day of death. All animals were subjected to a necropsy and a macroscopic examination.
The substance caused the death of 1/6 animals at a dose level of 2000 mg/kg bw. In the rat, dosed at 2000 mg/kg bw, that was found dead on Day 1, the lungs (all lobes) were collapsed and dark red (diffuse).There was no evidence of the macroscopic observations in the surviving animals dosed at 2000 mg/kg bw and terminated on Day 14.
At dose level of 2000 mg/kg bw, decreased activity (score 1 or 2, up to Day 2), hunched back (up to Day 2), incoordination (score 1 or 2, up to Day 1), piloerection (up to Day 2) in all animals and diuresis (between Day 2 and Day 4) in all surviving animals were observed. From Day 5 all surviving animals were symptom-free.
There were no effects on body weights or body weight gains that could be attributed to treatment with the test substance.
Under the conditions of the study, the acute oral LD50 value of the test item was found to be above 2000 mg/kg bw in female Crl:WI rats.
Acute toxicity study by dermal route (Tarcai 2018) :
An acute dermal toxicity study was performed with the test item n-nitrosodihenylamine in female Crl:WI Wistar rats, in compliance with OECD Guideline No. 402 (2017).
Test item did not cause mortality at the dose level of 2000 mg/kg bw. There were no systemic clinical signs noted in any animal throughout the study. No adverse local dermal signs were observed after treatment with the test item or during the 14 days observation period. The test item coloured the treatment area to yellowish on Day 1 (3/3 animals) and Day 2 (2/3) animals. There was no treatment related effects on body weight or body weight gain during the observation period. Body weights were within the range commonly recorded for this strain and age. There was no evidence of any macroscopic changes at a dose level of 2000 mg/kg bw. The acute dermal median lethal dose (LD50) of the test item n-nitrosodiphenylamine was found to be greater than 2000 mg/kg body weight in female Crl:WI rats.
Justification for classification or non-classification
Based on the available data, no classification for the acute toxicity is required for Nitrosodiphenylamine according to the Regulation EC n°1272/2008.
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