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EC number: 201-132-3 | CAS number: 78-67-1
- 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
Developmental toxicity / teratogenicity
Administrative data
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 19 OCTOBER 2012 to 24 JANUARY 2014
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: Conducted according to current test guidelines and GLP compliant
Cross-reference
- Reason / purpose for cross-reference:
- reference to other study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 014
- Report date:
- 2014
Materials and methods
Test guideline
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- Deviations:
- no
- GLP compliance:
- yes
- Limit test:
- no
Test material
- Reference substance name:
- AZDN
- IUPAC Name:
- AZDN
- Reference substance name:
- 2,2'-dimethyl-2,2'-azodipropiononitrile
- EC Number:
- 201-132-3
- EC Name:
- 2,2'-dimethyl-2,2'-azodipropiononitrile
- Cas Number:
- 78-67-1
- Molecular formula:
- C8H12N4
- IUPAC Name:
- 2,2'-dimethyl-2,2'-azodipropiononitrile
- Reference substance name:
- 2,2'-dimethyl-2,2'azopropionitrile
- IUPAC Name:
- 2,2'-dimethyl-2,2'azopropionitrile
- Test material form:
- solid: particulate/powder
- Remarks:
- migrated information: powder
- Details on test material:
- - Name of test material (as cited in study report): AZDN
- Physical state: White powder
- Analytical purity: 99%
- Purity test date: 08 August 2012
- Lot/batch No.: 7582
- Expiration date of the lot/batch: 31 March 2013
- Stability under test conditions: Suspensions of 0.1 and 15 mg/mL were found to be stable for up to 11 days at 2 to 8C (study no. 8266542).
- Storage condition of test material: 2 to 8C in the dark
Constituent 1
Constituent 2
Constituent 3
Test animals
- Species:
- rat
- Strain:
- other: Crl:CD(SD)
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Laboratories, Margate, UK
- Age at study initiation: 9-10 weeks old at time of mating
- Weight at study initiation: 200.1 to 271.8 g at time of mating
- Fasting period before study: No
- Housing: Animals housed singly in cages that conform with the 'Code of practice for the housing and care of animals used in scientific procedures' (Home Office, London, 1989). Suitable wood bedding provided weekly to each cage. Wooden Aspen chew blocks and sizzle nest provided as environmental enrichment.
- Diet (e.g. ad libitum): SQC Rat and Mouse Breeder Diet No 3, Expanded (Special Diets Services Ltd, Witham, UK) ad libitum
- Water (e.g. ad libitum): Mains water ad libitum
- Acclimation period: No. Female animals delivered to the testing laboratory on Day 3 of gestation and examined on receipt. All animals found to be in good health.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 24C
- Humidity (%): 45 to 65%
- Air changes (per hr): 15 to 20
- Photoperiod (hrs dark / hrs light): 12 hours light, 12 hours dark
IN-LIFE DATES: From: 19 October 2012 to 08 November 2012
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS: Formulations were prepared on three occasions. The test substance was formulated as a suspension in corn oil following dispensary SOPs and the formulation method 8266543_O_01D.
VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil selected due to stability of test substance in the vehicle, and use in previous study R-95-007.
- Concentration in vehicle: Not specified
- Amount of vehicle (if gavage): 5 mL/kg
- Lot/batch no. (if required): Not specified
- Purity: Not available - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Stability and homogeneity previously assessed at 0.1 and 15 mg/mL in study 8266542.
CONCENTRATION VERIFICATION: Samples (three random aliquots from test substance formulations; two random aliquots from control formulations) prepared for use on the first and last day of dosing were taken for analysis of achieved concentration. - Details on mating procedure:
- - Impregnation procedure: purchased timed pregnant
Mating was performed overnight at the supplier's laboratory and confirmed by the presence of a vaginal plug in situ. - Duration of treatment / exposure:
- The test and control substances were administered orally, by gavage, to mated female rats daily from Day 6 to Day 19 of gestation.
- Frequency of treatment:
- Daily
- Duration of test:
- The females were maintained to Day 20 of gestation when they were killed and their uterine contents examined.
- No. of animals per sex per dose:
- 22 females per dose concentration
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale: The high dose level of 20 mg/kg/day was selected as this was expected to elicit mild maternal toxicity and was based on the range-finding study 8266558, where a dose level of 25 mg/kg/day elicited maternal toxicity in the form of slight body weight loss and reduced food consumption. The intermediate dose level of 5 mg/kg/day was selected as this is the geometric mean of the high and low dose levels. The low dose level of 1 mg/kg/day was selected as this was expected to be a no observed effect level (NOEL).
- Rationale for animal assignment (if not random): On Day 3 of gestation the animals were assigned to treatment groups using a randomisation procedure based on body weight and day of mating.
Examinations
- Maternal examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Beginning and end of the working day for signs of ill health and overt toxicity.
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: Each animal was given a detailed physical examination on the days of body weight recording.
BODY WEIGHT: Yes
- Time schedule for examinations: The body weight of each female was recorded on Days 3, 6, 7, 8, 9, 12, 15, 17, 19 and 20 of gestation.
FOOD CONSUMPTION):
Individual food consumption was recorded for Days 3 to 5, 6, 7, 8, 9 to 11, 12 to 14, 15 to 16 and 17 to 18 and 19 of gestation.
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation Day 20
- Organs examined: Ovaries and uteri - Ovaries and uterine content:
- The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
Early intrauterine deaths were classified as those which showed decidual or placental tissue only. Late intrauterine deaths showed embryonic or foetal tissue in addition to placental tissue. Dead foetuses were classified as those which appeared to have died shortly before necropsy. - Fetal examinations:
- Live foetuses were killed by a subcutaneous injection of sodium pentobarbitone.
Individual foetal and placental weights were recorded and foetuses were examined externally and sexed. Approximately one half of the foetuses in each litter, selected by systematic sampling, were examined for visceral abnormalities by microdissection. They were then eviscerated and the carcasses processed to stain the ossified skeleton by the Alizarin technique and cartilage processed to stain using Alcian Blue. The skeletons were examined, preserved and stored in glycerol/propylene glycol.
The remaining foetuses were placed in Bouin's solution for at least two weeks to allow fixation and partial decalcification. At examination, the head was removed by a cut through the mouth, pharynx and back of the head and coronal sections of the head were examined. The remaining portion of the foetus was examined by dissection and was preserved, with the head sections, in 10% neutral buffered formalin and stored in plastic vials.
The dissection of the foetuses and the examination of the stained skeletons were performed using low power binocular magnification.
Foetal abnormalities were classified as malformations (rare and/or potentially lethal defects) and variations (commonly occurring non lethal abnormalities). - Statistics:
- The control group (Group 1) was taken as the baseline group with which the treated groups (Groups 2, 3 and 4) were compared.
Statistical methods included one-way analysis of variance (ANOVA), Levene's test, Dunnett's test, Kruskal-Wallis ANOVA, the Terpstra-Jonckheere test, the Wilcoxon rank sum test, analysis of covariance (ANCOVA), Cochran-Armitage test and Fisher's exact test. - Indices:
- Not applicable
- Historical control data:
- Not applicable
Results and discussion
Results: maternal animals
General toxicity (maternal animals)
- Clinical signs:
- effects observed, non-treatment-related
- Dermal irritation (if dermal study):
- no effects observed
- Mortality:
- mortality observed, non-treatment-related
- Description (incidence):
- One animal receiving 1 mg/kg/day (number 23) died on Day 7 of gestation and one animal receiving 5 mg/kg/day (number 62) died on Day 20 of gestation. Findings for these animals were consistent with dosing accidents including thoracic cavity containing abnormal liquid contents.
- Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- Slight body weight loss was seen in animals receiving 5 and 20 mg/kg/day from Days 6 to 8 of gestation inclusive. Statistically significant reduced body weight gain from Days 7 to 8 of gestation at 5 and 20 mg/kg/day (79% and 173% less, or P<0.05 and P<0.001 respectively). Percentage body weight change (corrected) on Day 20 of gestation was lower than control at 5 and 20 mg/kg/day (12% and 29% lower respectively). These were considered to be effects of treatment.
- Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- In animals receiving 5 and 20 mg/kg/day there was a statistically significant treatment-related reduction in food intake from Days 6 to 8 of gestation inclusive (between 5% and 47% less, or P<0.05 for both), with recovery seen afterwards. This was an effect of treatment.
- Food efficiency:
- not examined
- Water consumption and compound intake (if drinking water study):
- not examined
- Ophthalmological findings:
- not examined
- Haematological findings:
- not examined
- Clinical biochemistry findings:
- not examined
- Urinalysis findings:
- not examined
- Behaviour (functional findings):
- not examined
- Immunological findings:
- not examined
- Organ weight findings including organ / body weight ratios:
- effects observed, non-treatment-related
- Gross pathological findings:
- effects observed, non-treatment-related
- Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- not examined
- Histopathological findings: neoplastic:
- not examined
Maternal developmental toxicity
- Number of abortions:
- no effects observed
- Description (incidence and severity):
- There were 22, 22, 22 and 22 pregnancies leading to 22, 21, 21 and 22 litters in animals receiving 0, 1, 5 and 20 mg/kg/day respectively. The mean number of corpora lutea, the mean incidence of pre- and post-implantation loss and mean litter size were all unaffected by treatment.
- Pre- and post-implantation loss:
- no effects observed
- Total litter losses by resorption:
- no effects observed
- Description (incidence and severity):
- Sex ratio, mean litter weight, mean placental weight and mean foetal weight all showed no effect of treatment.
- Early or late resorptions:
- no effects observed
- Dead fetuses:
- no effects observed
- Changes in number of pregnant:
- no effects observed
- Details on maternal toxic effects:
- Maternal toxic effects:yes
Details on maternal toxic effects:
Morbidity and mortality:
One animal receiving 1 mg/kg/day (number 23) died on Day 7 of gestation and one animal receiving 5 mg/kg/day (number 62) died on Day 20 of gestation. Findings for these animals were consistent with dosing accidents including thoracic cavity containing abnormal liquid contents.
There were no unscheduled treatment-related deaths.
Clinical signs:
There were no significant, treatment-related clinical signs recorded.
Post-dosing observations:
On the first dosing the post-dosing observation mouth rubbing was seen in two animals receiving 1 mg/kg/day and three animals receiving 5 mg/kg/day. During the rest of the dose period mouth rubbing was seen on most days in up to six animals receiving 20 mg/kg/day. This commonly recorded observation is considered to be a result of taste-aversion rather than systemic toxicity.
Body weight:
Slight body weight loss was seen in animals receiving 5 and 20 mg/kg/day from Days 6 to 8 of gestation inclusive. Statistically significant reduced body weight gain from Days 7 to 8 of gestation at 5 and 20 mg/kg/day (79% and 173% less, or P<0.05 and P<0.001 respectively). Percentage body weight change (corrected) on Day 20 of gestation was lower than control at 5 and 20 mg/kg/day (12% and 29% lower respectively). These were considered to be effects of treatment.
Food consumption:
In animals receiving 5 and 20 mg/kg/day there was a statistically significant treatment-related reduction in food intake from Days 6 to 8 of gestation inclusive (between 5% and 47% less, or P<0.05 for both), with recovery seen afterwards. This was an effect of treatment.
Effect levels (maternal animals)
- Dose descriptor:
- NOAEL
- Effect level:
- 1 mg/kg bw/day (nominal)
- Based on:
- test mat.
- Basis for effect level:
- other: maternal toxicity
Results (fetuses)
- Fetal body weight changes:
- no effects observed
- Reduction in number of live offspring:
- no effects observed
- Changes in sex ratio:
- no effects observed
- Changes in litter size and weights:
- no effects observed
- Changes in postnatal survival:
- no effects observed
- External malformations:
- effects observed, non-treatment-related
- Skeletal malformations:
- effects observed, non-treatment-related
- Visceral malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Malformations were noted in three foetuses from three litters in the control group (one external/visceral, two skeletal), four foetuses from three litters at 1 mg/kg/day (skeletal), five foetuses from two litters at 5 mg/kg/day (skeletal), and nine foetuses from eight litters at 20 mg/kg/day (three external/visceral, six skeletal). The incidence and intergroup distribution of these foetal malformations and variations do not indicate an adverse effect of treatment. Findings which on occasion achieved statistical significance generally fell within the background range (abnormal shaped head, thymus cervical remnant, metacarpal 5 unossified), or were commonly seen findings which did not show a clear treatment-related increase across the groups and/or were not considered to be biologically significant (kidney cavitation increased, ureter distended – fluid contents).
- Other effects:
- no effects observed
- Details on embryotoxic / teratogenic effects:
- Embryotoxic / teratogenic effects:no effects
Details on embryotoxic / teratogenic effects:
Uterine/implantation data:
There were 22, 22, 22 and 22 pregnant females leading to 22, 21, 21 and 22 litters in animals receiving 0, 1, 5 and 20 mg/kg/day respectively. The mean number of corpora lutea, the mean incidence of pre- and post-implantation loss and mean litter size were all unaffected by treatment.
Foetal data:
Sex ratio, mean litter weight, mean placental weight and mean foetal weight all showed no effect of treatment.
Foetal defect data:
Malformations were noted in three foetuses from three litters in the control group (one external/visceral, two skeletal), four foetuses from three litters at 1 mg/kg/day (skeletal), five foetuses from two litters at 5 mg/kg/day (skeletal), and nine foetuses from eight litters at 20 mg/kg/day (three external/visceral, six skeletal). The incidence and intergroup distribution of these foetal malformations and variations do not indicate an adverse effect of treatment. Findings which on occasion achieved statistical significance generally fell within the background range (abnormal shaped head, thymus cervical remnant, metacarpal 5 unossified), or were commonly seen findings which did not show a clear treatment-related increase across the groups and/or were not considered to be biologically significant (kidney cavitation increased, ureter distended – fluid contents) (see attached tables).
Effect levels (fetuses)
- Dose descriptor:
- NOAEL
- Effect level:
- > 20 mg/kg bw/day (nominal)
- Sex:
- male/female
- Basis for effect level:
- other: no effect
Fetal abnormalities
- Abnormalities:
- effects observed, non-treatment-related
Overall developmental toxicity
- Developmental effects observed:
- not specified
Any other information on results incl. tables
Summary of Foetal Defects |
|||||
Defect |
Mean incidence – number of foetuses (mean percentage of foetuses) |
Background Incidence minimum %, maximum % |
|||
|
Group |
||||
|
1 |
2 |
3 |
4 |
|
External/visceral findings |
|||||
Malformations |
|||||
Aortic arch, retro-oesophageal |
- |
- |
- |
1 (0.3) |
|
Eye, anterior chamber, internal haemorrhage |
- |
- |
- |
1 (0.3) |
|
Eyes, one absent |
- |
- |
- |
1 (0.5) |
|
Eyes, severely reduced in size |
1 (0.4) |
- |
- |
1 (0.5) |
|
Variations |
|
|
|
|
|
Head, abnormal shape, domed |
2 (0.9) |
- |
2 (0.7) |
9 (3.4)DR* |
0.7, 4.4 |
Kidney, cavitation increased |
6 (3.0) |
5 (1.7) |
4 (1.7) |
18 (10.7)* |
0.8, 4.9 |
Thymus, cervical remnant |
1 (0.4) |
6 (2.1) |
7 (2.6) |
11 (4.0)* |
0.0, 11.2 |
Ureter, distended – fluid contents |
10 (4.6) |
6 (2.1) |
8 (3.5) |
18 (10.8) |
1.9, 6.6 |
Skeletal findings |
|||||
Malformations |
|||||
Forelimbs, severely shortened and bent with bent scapulae |
- |
- |
- |
1 (0.9) |
|
Ribs, ossification interrupted |
- |
1 (0.8) |
- |
- |
|
Vertebral thoracic centrum, cleft cartilaginous centres |
2 (1.3) |
1 (0.7) |
2 (1.4) |
2 (1.6) |
|
Vertebral cervical arches, additional cartilaginous ventral plate |
- |
1 (0.6) |
- |
- |
|
Variations |
|
|
|
|
|
Metacarpal 5 , unossified |
26 (21.7) |
14 (11.6) |
38 (30.2) |
38 (26.7)* |
17.1, 45.2 |
|
|
|
|
|
|
* P<0.05
DR = significant dose response test
Applicant's summary and conclusion
- Conclusions:
- In conclusion, administration of AZDN to pregnant rats from Days 6 to 19 of gestation at dose levels of 1 ,5 and 20 mg/kg/day resulted in maternal toxicity from 5 mg/kg/day upwards based on transient effects of reduced body weight gain and reduced mean food intake while no effects on foetal development were observed.
At 1 mg/kg/day, no adverse effects were observed in both parental and foetal development.
There was no evidence of embryotoxicity at any dose level tested.
The maternal no observed adverse-effect level (NOAEL) was 1 mg/kg/day based on a significant decrease in maternal body weight gain and food consumption.
The embryo-foetal NOAEL was 20 mg/kg/day. - Executive summary:
The objective of the study was to determine the effects of the test substance, AZDN(2,2’-dimethyl-2,2’azodipropionitrile), on the embryonic and foetal development of the rat.
The test item and control substance (corn oil) were administered orally, by oral gavage, to mated female rats daily from Day 6 to Day 19 of gestation, inclusive. The animals were dosed in ascending group order. The females were maintained to Day 20 of gestation when they were killed and their uterine contents examined.
Groups of rats of the Crl:CD(SD) strain were dosed as follows:
Group Number
Description
Dose level (mg/kg/day)
Number of animals in group
1
Control
0
22
2
Low
1
22
3
Intermediate
5
22
4
High
20
22
Two females one from each of the low and intermediate dose groups died on Days 7 and 20 of gestation respectively due to dosing accidents. Findings for these animals were consistent with dosing accidents including thoracic cavity containing abnormal liquid contents. There were no treatment-related deaths, clinical signs, or macroscopic necropsy findings.
The minor post-dosing observation mouth rubbing was seen in all treated groups with frequency of observation increasing with increasing dose.
Treatment-related reduced food intake (between 5% and 47% less, or P<0.05) for both 5 and 20 mg/kg/day and a consequent slight body weight loss (79% and 173% less, or P<0.05 and P<0.001 respectively) were seen from Days 6 to 8 of gestation inclusive.
Mean uterine/implantation and mean foetal data showed no adverse effect of treatment. There was no overall increase in the mean incidence or inter-group distribution of external, visceral or skeletal foetal variations or malformations.
In conclusion, administration of AZDN to pregnant rats from Days 6 to 19 of gestation at dose levels of 1 ,5 and 20 mg/kg/day resulted in maternal toxicity from 5 mg/kg/day upwards based on transient effects of reduced body weight gain and reduced mean food intake while no effects on foetal development were observed.
At 1 mg/kg/day, no adverse effects were observed in both parental and foetal development.
There was no evidence of embryotoxicity at any dose level tested.
The maternal no-observed-adverse-effect-level (NOAEL) was 1 mg/kg/day based on a significant decrease in maternal body weight gain and food consumption from 5 mg/kg/day upwards.
The embryo-foetal NOAEL was 20 mg/kg/day.
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