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EC number: 206-992-3 | CAS number: 420-04-2
- 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:
- 2013-05-21 to 2013-06-20
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- Rationale for repetition of study: Cyanamide is registered in the USA as a plant growth regulator and is currently reviewed by EPA. In order to support an acceptable reference dose for short-term dermal exposure AlzChem decided to prepare a study according to OECD 414 and OPPTS 870.3700. A previous prenatal development study in the Sprague-Dawley rat with Hydrogen Cyanamide (HLA2319-124) did not establish a clear no effect level for maternal toxicity, so this study was performed in order to further characterize previous findings whilst using a wider range of doses. In addition, the relevant guidelines have been updated since the previous study was conducted. Therefore, this study provides all data as required in the current guidelines.
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 014
- Report date:
- 2014
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- Version / remarks:
- 2001
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EPA OPPTS 870.3700 (Prenatal Developmental Toxicity Study)
- Version / remarks:
- 1998
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
Test material
- Reference substance name:
- Cyanamide
- EC Number:
- 206-992-3
- EC Name:
- Cyanamide
- Cas Number:
- 420-04-2
- Molecular formula:
- CH2N2
- IUPAC Name:
- cyanamide
- Test material form:
- other: aqueous solution
Constituent 1
Test animals
- Species:
- rat
- Strain:
- Sprague-Dawley
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River Laboratories France, Domaine des Oncins, 69210 Saint-Germain-Nuelles, France
- Age at study initiation: age at mating 10 to 13 weeks
- Weight at study initiation: 198 to 245 g at mating
- Housing: One air-conditioned room in a barrier protected unit
- Diet: ad libitum, pelleted commercial complete rodent diet ad libitum (Diet reference A04C-10) sterilised by irradiation and analysed for a predefined list of chemical and bacteriological contaminants.
- Water: Filtered (0.2 µm) mains drinking water was available ad libitum (via bottles).
- Acclimation period: 6 days between animal arrival and the start of treatment.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 + 3
- Humidity (%): Between 35 and 70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12 / 12
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- water
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS:
The test item was prepared as a solution in the vehicle at concentrations of 0.3, 1.5 and 4.5 mg Hydrogen Cyanamide/mL based on a content of 526.4 mg/mL which took into account the actual density (1.057 g/cm3 at +20 °C) and concentration of Hydrogen Cyanamide (49.8 % w/w). Dosing solution was prepared weekly.
VEHICLE
- Amount of vehicle: Dose volume: 10 mL/kg - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Duplicate 3 mL samples were taken from each newly prepared formulation, including the vehicle, to ensure that the nominal concentrations were in line with the analytically determined concentrations. One set of samples was analysed immediately at the Test Facility using the validated method.
The transfer and validation of the analytical method as well as the stability analysis was the subject of a separate study plan (WIL Research study number AB18598). - Details on mating procedure:
- The females were mated at the supplier with a documented day of mating. They were received at the Test Facility on day 0 of gestation.
- Duration of treatment / exposure:
- From day 6 (G 6) to day 19 (G 19) of gestation inclusive.
- Frequency of treatment:
- once daily
- Duration of test:
- Animals were killed on the day after the last treatment, which was day 20 of gestation. Treatment had started on day 6.
Doses / concentrationsopen allclose all
- Dose / conc.:
- 0 mg/kg bw/day
- Dose / conc.:
- 3 mg/kg bw/day
- Remarks:
- active substance
- Dose / conc.:
- 15 mg/kg bw/day
- Remarks:
- active substance
- Dose / conc.:
- 45 mg/kg bw/day
- Remarks:
- active substance
- No. of animals per sex per dose:
- 25 females per dose
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale: The mid and high dose levels were selected solely to replicate a previous prenatal development study in the Sprague-Dawley rat (HLA 2319-124) which yielded equivocal results. In that study a marked maternal toxicity (including body weight loss) was elicited in the high dose group. The low dose was selected to try and establish a no effect level for maternal toxicity.
Examinations
- Maternal examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily
BODY WEIGHT: Yes
- Time schedule for examinations: on days 0, 3, 6, 9, 12, 15, 18 and 20 of gestation
FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: The ovaries and uterus of each female were removed and examined.
The placentae were also examined - Ovaries and uterine content:
- The ovaries and uterine content was examined after termination: Yes
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Other: -individual foetal weights, foetal sex - Fetal examinations:
- - External examinations: Yes [all per litter]
- Soft tissue examinations: Yes [all per litter]
- Skeletal examinations: Yes [appr. 50 % per litter]
- Head examinations: Yes [appr. 50 % per litter] - Statistics:
- Statistical analysis was performed by the Provantis data acquisition system, where appropriate, as follows:
The best transformation for the data (none, log or rank) was determined depending upon:
-the normality of the data distribution tested by the Shapiro-Wilk's test
-the homogeneity of the variances across groups tested by the Bartlett's test.
Non- or log-transformed data were analysed by parametric methods.
Rank transformed data were analysed using non-parametric methods.
Data were then analysed to test for a dose-related trend to detect the lowest dose at which there was a significant effect based on the Williams test for parametric data or the Shirley's test for non-parametric data.
Homogeneity of means was assessed by analysis of variances (ANOVA) for parametric data or Kruskal-Wallis test for non parametric data.
If no trend was found and means were not homogeneous, the data were analysed by parametric or non-parametric Dunnett's test to look for significant differences from the control group.
The number of resorptions, number of dead foetuses and all litter-based percentages were analysed using non-parametric methods, i.e. Kruskal-Wallis test followed by non parametric Dunnett’s test if the Kruskal-Wallis was significant.
Selected incidence data were analysed using a chi2 test for all groups followed by Fisher’s two-tailed test with Bonferroni correction for each treated group versus the control if the chi2 was significant. - Historical control data:
- Results were compared to Historical control data; Rat - CD® IGS (Crl:CD[SD]), 1996-2009; Charles River Laboratories Preclinical Services Montreal; All routes of administration; Embryo-fetal development studies.
Results and discussion
Results: maternal animals
General toxicity (maternal animals)
- Body weight and weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- There was a dose-related and statistically significant reduction in overall mean body weight gain and food consumption during the dosing period in the 15 and 45 mg/kg bw/d groups leading to lower mean terminal body weight (-6 % and -27 % in groups 3 and 4, respectively) compared with the control. The effect in the high dose group was associated with body weight loss during the first 6 days of the dosing period (GD 6 to GD 12). The effect was confirmed by lower mean net body weight change (i.e. maternal body weight change from GD 6 to GD 20 minus gravid uterus weight) in the 15 and 45 mg/kg bw/d groups (27 g and -34 g, respectively) compared with the control (48 g).
Maternal developmental toxicity
- Details on maternal toxic effects:
- Maternal toxic effects:yes
Details on maternal toxic effects:
Treatment-related clinical signs were restricted to the 45 mg/kg bw/day group and included piloerection for all females and decreased activity, intermittent whole body tremors and pale faeces for approximately half of the animals. The health status of three females in the high dose group was closely monitored due to severe body weight loss during the first 6 days of the dosing period, in order to decide if an ethical sacrifice was necessary. There was a dose-related and statistically significant reduction in overall mean body weight gain and food consumption during the dosing period (G 6 to G 20) in the 15 and 45 mg/kg bw/day groups leading to lower mean terminal body weight (-6 % and -27 % in groups 3 and 4, respectively) compared with the control. The effect in the high dose group was associated with body weight loss during the first 6 days of the dosing period (G 6 to G 12). The effect was confirmed by lower mean net body weight change (i.e. maternal body weight change from G 6 to G 20 minus gravid uterus weight) in the 15 and 45 mg/kg bw/day groups (27 g and -34 g, respectively) compared with the control (48 g).
There were no treatment-related macroscopic findings in any group.
Effect levels (maternal animals)
open allclose all
- Key result
- Dose descriptor:
- NOEL
- Effect level:
- 3 mg/kg bw/day
- Based on:
- test mat.
- Basis for effect level:
- other: maternal toxicity
- Key result
- Dose descriptor:
- NOEL
- Effect level:
- 3 mg/kg bw/day
- Based on:
- test mat.
- Basis for effect level:
- other: developmental toxicity
Maternal abnormalities
- Key result
- Abnormalities:
- no effects observed
Results (fetuses)
- Details on embryotoxic / teratogenic effects:
- Embryotoxic / teratogenic effects:yes
Details on embryotoxic / teratogenic effects:
Consistent with the marked maternal toxicity, there was clear evidence of general developmental toxicity (delay in development) in the 45 mg/kg bw/day group with markedly reduced mean foetal weight (-38 %) (resulting in lower mean gravid uterus weight), widespread incomplete/absent ossification of axial and appendicular skeletal bones and thickened and/or wavy ribs (7 % and 31 %, respectively) compared with the control. Also consistent with the low foetal weight and the delayed development, there were also 7 foetuses from the same number of litters with a malpositioned (not completely descended) testis. It is worthy of note that these malformations were observed in only one female out of the three which had the most pronounced maternal toxicity.
Other malformations noted in the high dose group including 10 (7) foetuses (litters) with a small diaphragmatic hernia (no impact on the lung size) and 4 (4) foetuses (litters) with defects of the great blood vessels can not be clearly attributed to the observed maternal toxicity. However, due to the severity of the maternal toxicity in the high dose group (27 % lower terminal mean body weight compared with the control), foetal well-being was compromised, so the regulatory value of the data in this group are likely to be of limited value for determining developmental effects.
In the 15 mg/kg bw/day dose group, there was also a treatment-related lower mean foetal weight in comparison with the concurrent control (-8 %) (resulting in a lower mean gravid uterus weight of 13 %). The presence of a single foetus (1/312 foetuses – 0.3 % incidence) in the mid dose group with great blood vessel defects is probably incidental since great vessels defects are also seen at a low frequency (0.00 - 0.69 %) in the historical control data for the Sprague-Dawley rat.
Effect levels (fetuses)
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 15 mg/kg bw/day
- Based on:
- act. ingr.
- Sex:
- male/female
- Basis for effect level:
- fetal/pup body weight changes
- skeletal malformations
- other: malpositioned testes, diaphragmatic hernia, defects of the great bood vessels
Fetal abnormalities
- Key result
- Abnormalities:
- effects observed, treatment-related
- Localisation:
- skeletal: rib
- visceral/soft tissue: cardiovascular
- visceral/soft tissue: male reproductive system
- Description (incidence and severity):
- There was clear evidence of general developmental toxicity (delay in development) in the 45 mg/kg bw/d group with markedly reduced mean foetal weight (-38 %) (resulting in lower mean gravid uterus weight), widespread incomplete/absent ossification of axial and appendicular skeletal bones and thickened and/or wavy ribs (7 % and 31 %, respectively) compared with the control. There were also 7 foetuses from the same number of litters with a malpositioned (not completely descended) testis. Other malformations noted in the high dose group included 10 (7) foetuses (litters) with a small diaphragmatic hernia (no impact on the lung size) and 4 (4) foetuses (litters) with defects of the great blood vessels.
Overall developmental toxicity
- Key result
- Developmental effects observed:
- yes
- Lowest effective dose / conc.:
- 45 mg/kg bw/day
- Treatment related:
- yes
- Relation to maternal toxicity:
- not specified
- Dose response relationship:
- yes
Any other information on results incl. tables
Table 1: Summary of maternal and litter data
Dose level (mg/kg bw/d) | 0 | 5 | 15 | 45 |
No of females in study | 25 | 25 | 25 | 25 |
Pregnant dams | 25 | 24 | 25 | 23 |
Corpora lutea (mean) | 15.2 | 14.7 | 14.6 | 14.6 |
Implantation sites (mean) | 14.1 ± 2.0 | 14.0 ± 2.5 | 14.0 ± 2.0 | 14.1 ± 1.7 |
Early resorptions (%) | 6.96 ± 8.18 | 8.63 ± 9.98 | 10.23 ± 7.25 | 8.06 ± 8.07 |
Late resorptions (mean % ± SD) | 0.87 ± 2.42 | 3.40 ± 12.30 | 0.77 ± 2.87 | 3.14 ± 8.39 |
Pre-implantation loss (mean % ± SD) | 6.51 ± 9.30 | 5.36 ± 8.75 | 3.72 ± 7.48 | 3.2 ± 4.78 |
Post implantation loss (mean %) | 7.83 ± 9.27 | 12.03 ± 14.13 | 11.00 ± 7.88 | 11.20 ± 13.27 |
|
|
|
|
|
Body weight gain day 6-16 (g) | 50.2 | 40.9* | 31.9* | 6.2* |
Body weight gain day 6-20 (g) | 105.8 | 94.2* | 81.4* | 51.0* |
Food consumption day 6-16 (% of control) | 100 | 96 | 89* | 76* |
Food consumption day 16-20 (% of control) | 100 | 96 | 92* | 77* |
Carcass weight (adjusted) (g) | 315.7 | 315.2 | 300.7* | 236.6* |
Net weight change (adjusted) from day GD6-20 (g) | 47.75 ± 10.67 | 43.76 ± 10.59 | 27.15 ± 11.24** | -34.34 ± 20.30*** |
Gravid uterus weight (g) | 80.18 | 75.48 | 69.80* | 51.39*** |
Live litter size (mean) | 13 ± 2.4 | 12.5 ± 2.9 | 12.5 ± 2.2 | 12.6 ± 2.7 |
Mean foetal weight (G) | 4.01 ± 0.21 | 3.97 ± 0.30 | 3.67 ± 0.31** | 2.47 ± 0.35*** |
Dead foetuses (%) | 0 | 0 | 0 | 0 |
% male foetuses | 47.49 | 50.97 | 50.14 | 44.70 |
*/** statistically significant (p-values not indicated)
Table 2: Summary of relevant foetal observations
Dose level (mg/kg bw/d) | 0 | 3 | 15 | 45 |
|
|
|
|
|
Diaphragma hernia | 0 | 0 | 0 | 10 (7)** |
Testis malpositioned | 0 | 0 | 0 | 7 (7)** |
Great blood vessel defects - aortic arch narrowed - multiple abnormalities | 0 0 0 | 0 0 0 | 1 (1) 0 1 (1) | 4 (4) 1 (1) 3 (3) |
Fused sternebrae | 0 | 0 | 0 | 2 (1) |
Hemisterebrae | 0 | 0 | 0 | 3 (3) |
Wavy/bent ribs | 0 | 0 | 0 | 47 (19)** |
Incomplete ossification - skull - limbs - sternebrae - ribs - vertebrae - pervic girdle | - - - - - - - | - - - - - - - | - - - - - - - | ↑↑↑** ↑↑↑** ↑↑↑** ↑↑↑** ↑↑↑** ↑↑↑** ↑↑↑** |
Thickened ribs | 0 | 0 | 0 | 10 (16)* |
Short rib | 0 | 0 | 0 | 4 (4) |
Thoracic Vertebra: bipartite ossification of centrum | 1 (1) | 1 (1) | 2 (2) | 14 (10)** |
Hemicentrum | 0 | 0 | 0 | 2 (1)** |
*/** statistically significant (p-values not indicated)
Applicant's summary and conclusion
- Conclusions:
- Oral (gavage) administration of hydrogen cyanamide to Sprague-Dawley rats from days 6 to 19 of gestation at doses of 3, 15 and 45 mg/kg bw/day active ingredient was associated with dose-related lower mean body weight gain and food consumption in the mid and high dose groups leading to 6 % and 27 % lower mean terminal body weight (effects tending to be more pronounced between G 6 and G 12), respectively, compared with the control. In the high dose group, the effect led to a net body weight loss (-34 g compared with +48 g in the control) and it was associated with marked clinical effects (i.e. piloerection, decreased activity, intermittent whole body tremors and pale faeces). Although there was no mortality, a premature sacrifice was considered for three females in the high dose group (45 mg/kg bw/day) due to the severe maternal toxicity.
Foetal development was clearly compromised in the high dose group (45 mg/kg bw/day) with a marked reduction in foetal weight, a delayed testicular descent, a delay in ossification of axial and appendicular skeletal bones and rib anomalies. However, within the context of this study, it is not possible to establish if other developmental effects (small diaphragmatic hernia and great blood vessel changes) can be attributed to the marked maternal toxicity and the compromised foetal well-being. Therefore, the regulatory value of the data in this group is likely to be of limited value for determining developmental effects.
The only developmental effect clearly attributable to treatment in the mid dose group of 15 mg/kg bw/day group was a slight reduction in mean foetal weight.
The low dose of 3 mg/kg bw/day is a no observed effect level (NOEL) for both maternal and embryo-foetal toxicity. - Executive summary:
The test item, hydrogen cyanamide, was administered by gavage at dose levels of 3, 15 and 45 mg/kg bw/day (active ingredient) to groups of 25 mated female Sprague-Dawley rats from days 6 to 19 of gestation inclusive. A control group received the same dose volume (10 mL/kg bw) of the vehicle (water for injection). Clinical condition, body weights and food consumption were monitored throughout the study. The females were submitted to a caesarean examination on day 20 of gestation. At necropsy, the females were examined macroscopically, the gravid uterus was weighed and litter parameters were recorded. All foetuses were weighed, sexed and examined for external andfresh visceral (abdominal and thoracic cavities) abnormalities.The heads of approximately half of the foetuses/litter were removedand subsequently submitted to fixed soft tissue examinationby serial sectioning. The eviscerated carcasses of the remaining half of the foetuses were submitted to skeletal examination. There was no mortality in any group.
Treatment-related clinical signs were restricted to the 45 mg/kg bw/day group and included piloerection for all females and decreased activity, intermittent whole body tremors and pale faeces for approximately half of the animals. The health status of three females in the high dose group was closely monitored due to severe body weight loss during the first 6 days of the dosing period, in order to decide if an ethical sacrifice was necessary.
There was a dose-related and statistically significant reduction in overall mean body weight gain and food consumption during the dosing period (G 6 to G 20) in the 15 and 45 mg/kg bw/day groups leading to lower mean terminal body weight (-6 % and -27 % in groups 3 and 4, respectively) compared with the control. The effect in the high dose group was associated with body weight loss during the first 6 days of the dosing period (G 6 to G 12). The effect was confirmed by lower mean net body weight change (i.e. maternal body weight change from G 6 to G 20 minus gravid uterus weight) in the 15 and 45 mg/kg bw/day groups (27 g and -34 g, respectively) compared with the control (48 g).
There were no treatment-related macroscopic findings in any group.
There were 25, 24, 25 and 23 pregnant females in the control, low, mid and high dose groups, respectively, at the terminal caesarean examinations, all of which had viable foetuses.
There was no influence of treatment on embryo-foetal survival or on sex ratio in any group.
Consistent with the marked maternal toxicity, there was clear evidence of general developmental toxicity (delay in development) in the 45 mg/kg bw/day group with markedly reduced mean foetal weight (-38 %) (resulting in lower mean gravid uterus weight), widespread incomplete/absent ossification of axial and appendicular skeletal bones and thickened and/or wavy ribs (7 % and 31 %, respectively) compared with the control. Also consistent with the low foetal weight and the delayed development, there were also 7 foetuses from the same number of litters with a malpositioned (not completely descended) testis. It is worthy of note that these malformations were observed in only one female out of the three which had the most pronounced maternal toxicity.
Other malformations noted in the high dose group including 10 (7) foetuses (litters) with a small diaphragmatic hernia (no impact on the lung size) and 4 (4) foetuses (litters) with defects of the great blood vessels can not be clearly attributed to the observed maternal toxicity. However, due to the severity of the maternal toxicity in the high dose group (27 % lower terminal mean body weight compared with the control), foetal well-being was compromised, so the regulatory value of the data in this group are likely to be of limited value for determining developmental effects.
In the 15 mg/kg bw/day dose group, there was also a treatment-related lower mean foetal weight in comparison with the concurrent control (-8 %) (resulting in a lower mean gravid uterus weight of 13 %). The presence of a single foetus (1/312 foetuses – 0.3 % incidence) in the mid dose group with great blood vessel defects is probably incidental since great vessels defects are also seen at a low frequency (0.00 - 0.69 %) in the historical control data for the Sprague-Dawley rat.
Based on the observed findings, the low dose of 3 mg/kg bw/day is a no observed effect level (NOEL) for both maternal and embryo-foetal toxicity.
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