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EC number: 911-254-5 | CAS number: -
- 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
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Remarks:
- Meets the requirements of GLP. There are no deviations from the recommended guideline.
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 013
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.31 (Prenatal Developmental Toxicity Study)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Limit test:
- no
Test material
- Reference substance name:
- Reaction mass of 2-tert-butyl-4,6-dimethylphenol and 4-tert-butyl-2,5-dimethylphenol
- EC Number:
- 911-254-5
- Molecular formula:
- Not applicable - Multiconstituent substance
- IUPAC Name:
- Reaction mass of 2-tert-butyl-4,6-dimethylphenol and 4-tert-butyl-2,5-dimethylphenol
- Test material form:
- liquid
- Remarks:
- (yellow-reddish liquid)
Constituent 1
Test animals
- Species:
- rat
- Strain:
- Wistar
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Experimental Medicine Centre at the Medical University in Białystok
- Age at study initiation: 13 weeks old
- Weight at study initiation: Average body weight of females of control group and treated groups: 232.5 g.
- Housing: Animals were kept in plastic cages with metal wire lid and dimensions: 37 x 22 x 15 cm (lenght x width x height) on UV sterilized bedding. At mating one female was placed in one cage with one male. At pregnancy the females were housed individually.
- Diet (e.g. ad libitum): “Murigran” standard laboratory fodder, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: Minimum 3 weeks.
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 19-23 ºC
- Humidity (%): 30-80%
- Air changes (per hr): about 16 times/hour
- Photoperiod (hrs dark / hrs light): 12 hours light/12 hours dark
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS:
The test item was diluted in corn oil directly before administration, in concentration correct for each dose.
A constant volume of solution (0.4mL of solution / 100 g bw) was used at every level of dosage. - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- In order to confirm correct preparation of the test item, each of three doses was analyzed during the whole administration period using a liquid chromatographic method. The linearity of response of the analytical method, its specificity, precision, recovery, limit of quantification and detection were assessed in the process of the analytical method validation.
- Details on mating procedure:
- - If cohoused: The females were mated with not related males. The males came from the same husbandry and stock and were at a similar age.
- M/F ratio per cage: One male was used to mate with maximally three females.
- Length of cohabitation: from 1 to 4 days. In case of unsuccessful pairing the first male was replaced by a second one (4 cases), a third one (2 cases) or a fourth male (1 case).
- Proof of pregnancy: For each mating one female was placed in a cage with one male. Everyday in the morning, vaginal smears were taken from females. The sperm-positive females were accepted as mated and the day of observation of sperm in the smear was accepted as 0 day of gestation. - Duration of treatment / exposure:
- The test item in treated groups and the corn oil in control group were given to pregnant females by gavage daily from 0 to 19th day of gestation.
- Frequency of treatment:
- Daily
- Duration of test:
- On day 20 of gestation (one day prior to the expected day of delivery), females were euthanised by intraperitoneal administration of morbital and subjected to caesarean section.
Doses / concentrationsopen allclose all
- Dose / conc.:
- 8.3 mg/kg bw/day (actual dose received)
- Remarks:
- Group 1
- Dose / conc.:
- 50 mg/kg bw/day (actual dose received)
- Remarks:
- Group 2
- Dose / conc.:
- 300 mg/kg bw/day (actual dose received)
- Remarks:
- Group 3
- No. of animals per sex per dose:
- 25 females per group
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- - Dose selection rationale:
Sighting range-finding study:
Three different doses of the test item (10, 50 and 250 mg/kg bw) were administered orally to three groups of pregnant females. A concurrent control group of pregnant females was not given the test item. Seven pregnant females were used per group.
The doses were selected on the grounds of the LD50 value which is about 500 mg/kg bw (300 mg/kg bw: no mortality occurred; 2000 mg/kg bw: three animals died). The highest dose should be between 1/2-1/5 of the LD50, the medium dose should be between 1/10-1/20 of the LD50 and the lowest dose should be between 1/50-1/100 of the LD50.
The test item in the used doses administered from 5th to 19th gestation day, did not indicate any toxic influence on pregnant females, except for the decrease of food consumption from 5th to 8th in group 3. A statistically significant reduction on the weight of placenta was observed in the treated groups. No embryotoxicity, no fetotoxicity and no teratogenic effects were stated in the examined fetuses.
Examinations
- Maternal examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: The evaluation of general condition of the animals, i.e. the observation of all animals for morbidity and mortality was performed twice a day during labour week and once a day on days off.
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: The detailed clinical observations were performed for all females on day of beginning of experiment (day 0) and then on 2nd, 5th, 8th, 11th, 14th, 17th and 20th day of gestation. The observations comprised: skin changes, coat changes, changes in eyes and mucous membranes, respiratory system, circulatory system, autonomous and central nervous system, somatic activity and behavior.
BODY WEIGHT: Yes
- Time schedule for examinations: Body weights of the females were determined on days 0, 2, 5, 8, 11, 14, 17, and 20 of gestation.
FOOD CONSUMPTION: Yes
- Time schedule for examinations: Days 0, 2, 5, 8, 11, 14, 17, and 20 of gestation.
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: At necropsy, females were examined macroscopically for any abnormalities in body structure or pathological changes which could have influenced gestation. - Ovaries and uterine content:
- The ovaries and uterine content was examined after termination: Yes.
After caesarean section, uterus from each female with fetuses was dissected. After removing of fetuses left and right uteri horn were examined. Number of viable and dead fetuses as well as number of resorptions was determined in each horn. Each pregnant uterus after removing of fetuses was weighed. Uteri of non pregnant females and non pregnant horns of uteri were fixed between two glass plates and overexposed by electric lamp to confirm the non-pregnant status. Then these uteri were stained using ammonium sulphide and then again overexpose with light by electric lamp in order to confirm the non-pregnant status. At the necropsy ovaries from each pregnant female were collected and fixed in 10% solution of formalin in order to determine number of corpora lutea.
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 - Fetal examinations:
- - External examinations: Yes. Sex of each fetus, body weight, length with tail and without tail was determined. Weight of fetuses with placenta and fetal membranes was determined first and then weight of fetuses without placenta and fetal membranes as well as weight of placenta were measured. After removal of fetal membranes each fetus was measured with tail and without tail, and detailed gross evaluation was performed.
During the gross evaluation attention was paid to:
- reactions to tactile impulses
- formation of body coverings
- formation of limbs
- number of fingers and toes
- shape of head
- formation of auricular conchas
- presence of nasal apertures
- formation of oral cavity
- presence of anus and tail
- Skeletal examinations: Yes. In every group, half of fetuses of each litter was subjected to evaluation of skeleton. The fetuses were fixed in 95% ethanol then overexposed in 1% KOH and stained with alizarin S, decolorized in Moll solution and moved to glycerine. The stained fetuses were examined and formation of skull bones, back-bone, ribs, acromial and pelvic girdles with limbs was evaluated. Points of ossification in sternum, metacarpus of fore limbs and metatarsus of hind limbs were counted during evaluation.
- Soft tissue examinations: Yes. The remaining fetuses from each litter were fixed in 10% solution of formalin, then cut in sagittal plane. Formation of particular cavities and presence of internal organs was evaluated.
- Head examinations: Yes. - Statistics:
- Evaluation of the study was performed for pregnant females from which fetuses were obtained at necropsy. The obtained results (body weight of pregnant females, food consumption of pregnant females, number of corpora lutea, number of fetuses in litter, number of males and females in litter, weight of uteri, weight of fetuses, length of fetuses, number of ossification points in sternum and limbs) were elaborated statistically with the use of t-Student test with p ≤ 0.05.
Results and discussion
Results: maternal animals
General toxicity (maternal animals)
- Clinical signs:
- effects observed, treatment-related
- Description (incidence and severity):
- Coat thinning in one female of group 1 and in one female of group 3 should not be connected with the test item. This sign occurred also in three females in control group and should be connected rather with hormonal changes during gestation. Clinical signs on the last day of experiment observed in one female of group 3 were connected with death of fetuses at the last period of gestation. No other clinical signs were stated in the females.
- Dermal irritation (if dermal study):
- not examined
- Mortality:
- no mortality observed
- Description (incidence):
- All females survived during the period of experiment.
- Body weight and weight changes:
- no effects observed
- Description (incidence and severity):
- No statistically significant changes were observed in the mean body weights of the treated groups when compared to the control group at the end of the study period.
- Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- Statistically significantly lower food consumption in group 3 from 0 to 5th day of experiment was reflected in statistically significant lower body weight of females on 2nd and 5th day of gestation. Statistically significant greater food consumption from 8th to 11th day and from 14th to 20th day of experiment caused compensation of the low body weight of females at the initial period of experiment.
- 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 specified
- Urinalysis findings:
- not specified
- Behaviour (functional findings):
- not specified
- Immunological findings:
- not examined
- Organ weight findings including organ / body weight ratios:
- not specified
- Gross pathological findings:
- no effects observed
- Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- not specified
- Histopathological findings: neoplastic:
- not specified
- Other effects:
- not specified
Maternal developmental toxicity
- Number of abortions:
- not specified
- Pre- and post-implantation loss:
- effects observed, treatment-related
- Description (incidence and severity):
- In group 3, in one female fifteen implantation sites and lack of fetuses were stated in uterus.
- Total litter losses by resorption:
- effects observed, treatment-related
- Description (incidence and severity):
- In group 3, two resorptions in two females in group 3 were observed.
- Early or late resorptions:
- not specified
- Dead fetuses:
- effects observed, treatment-related
- Description (incidence and severity):
- One dead fetus in group 1 should not be connected with test item and should be considered as incidental.
In group 3 in one female, in which on day of dissection some clinical signs were observed, only dead fetuses were stated. - Changes in pregnancy duration:
- not specified
- Changes in number of pregnant:
- not specified
- Other effects:
- not specified
Effect levels (maternal animals)
open allclose all
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 50 mg/kg bw/day (actual dose received)
- Basis for effect level:
- other: maternal toxicity
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 50 mg/kg bw/day (actual dose received)
- Basis for effect level:
- other: developmental toxicity
Maternal abnormalities
- Key result
- Abnormalities:
- no effects observed
Results (fetuses)
- Fetal body weight changes:
- effects observed, treatment-related
- Description (incidence and severity):
- Differences on average body weight of fetuses in treated groups compared to control group were observed but there is no relationship between the dose and the effect. Decreased body weight of fetuses without placenta was observed in group 1 and group 3. However, increased body weight of fetuses without placenta was observed in group 2. The lowest weight of fetuses without placenta was observed in group 3. Average weight of fetuses in control group, group 1 and group 2 are within the range from literature data, while average weight of fetuses without placenta in group 3 – 3.338 g is below the lower range (3.35 g to 3.85 g).
The length of fetuses with tail and without tail was lower in group 3. It may confirm delay of the growth of fetuses in group 3 considering the changes in body weight and length of fetuses. - Reduction in number of live offspring:
- effects observed, treatment-related
- Description (incidence and severity):
- One dead fetus in group 1 should not be connected with test item and should be considered as incidental.
In group 3 in one female, in which on day of dissection some clinical signs were observed, only dead fetuses were stated. - Changes in sex ratio:
- not specified
- Changes in litter size and weights:
- not specified
- Changes in postnatal survival:
- not examined
- External malformations:
- no effects observed
- Skeletal malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- Dislocation of sternum ossification points and ribs attachment in one fetuses in group 1 should not be connected with test item. This case was noticed only in one fetus and similar changes in ribs and sternum were observed in fetuses of control groups in other prenatal developmental toxicity studies performed at the testing facility. Therefore, this change should be considered as a spontaneous disorder.
Decrease in number of ossification points is considered as a change associated with delayed development due to toxicity which does not influence postnatal development and is later compensated. A statistically significant lower number of ossification points of metacarpus in group 1 results from greater percentage of fetuses with 3 ossification points of metacarpus in group 1, whereas lower number of ossification points of metatarsus in group 3 results from the fact that in one female of this group 4 fetuses had 3 ossification points of metatarsus and 1 fetus had no ossification points of metatarsus. All other fetuses in group 3 had 4 ossification points of metatarsus. Additionally, in control group, which treated groups are compared to, average number of ossification points of metatarsus was above 4, because 7 fetuses from one female in control group had 5 ossification points of metacarpus (likewise in group 2). - Visceral malformations:
- no effects observed
- Other effects:
- no effects observed
- Description (incidence and severity):
- In each of the treated groups one case of conjoined placenta was stated. This finding is not considered to be treatment related. Subcutaneous blood hemorrhages observed in fetuses in different parts of the body occurred in control group and in treated groups and should not be connected with the test item. Subcutaneous blood hemorrhages were observed often in fetuses in other prenatal developmental toxicity studies performed in the testing facility and they should be considered as typical disturbances in fetuses.
Effect levels (fetuses)
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 50 mg/kg bw/day (actual dose received)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- reduction in number of live offspring
- fetal/pup body weight changes
Fetal abnormalities
- Key result
- Abnormalities:
- no effects observed
Overall developmental toxicity
- Developmental effects observed:
- not specified
Any other information on results incl. tables
Table 1 - Results of mating - main study
Parameter |
Number of females |
|||
Group 0 |
Group 1 |
Group 2 |
Group 3 |
|
Number of females per group |
25 |
25 |
25 |
25 |
Number of pregnant females |
21 |
23 |
22 |
20 |
Number of not pregnant females |
4 |
2 |
3 |
5 |
Females with all dead fetuses |
- |
- |
- |
1 |
Females with implantations and without fetuses |
- |
- |
- |
1 |
Females with resorptions |
6 [1,10,17,18, 19(2),20] |
3 [2,11,24] |
4 [3,7,24(4),25] |
2 [8,21] |
Number of females for evaluation |
21 |
23 |
22 |
18 |
[ ] computer numbers of females
( ) number of resorptions
Table 2 - Body weights of females (g) - main study
Day of gestation |
Group |
|||
0 |
1 |
2 |
3 |
|
0 |
233.71 ± 20.83 |
235.13 ± 20.47 |
230.23 ± 15.38 |
230.61 ± 15.44 |
2 |
237.48 ± 20.52 |
237.30 ± 23.32 |
232.05 ± 16.27 |
219.83 ± 14.26* |
5 |
244.67 ± 20.29 |
248.13 ± 21.41 |
245.77 ± 15.78 |
228.67 ± 21.77* |
8 |
253.14 ± 21.77 |
254.00 ± 21.68 |
252.18 ± 18.22 |
243.06 ± 20.82 |
11 |
264.90 ± 19.68 |
267.13 ± 23.76 |
265.95 ± 17.66 |
258.44 ± 25.22 |
14 |
275.29 ± 20.12 |
277.04 ± 22.44 |
278.45 ± 17.22 |
274.94 ± 20.76 |
17 |
300.57 ± 22.66 |
301.57 ± 23.53 |
304.14 ± 20.99 |
300.67 ± 23.53 |
20 |
335.81 ± 24.24 |
340.70 ± 25.87 |
340.64 ± 25.65 |
339.78 ± 30.57 |
* statistically significant difference with p ≤ 0.05
Table 3 - Food consumption (g/100 g bw/day) - main study
Day of gestation |
Group |
|||
0 |
1 |
2 |
3 |
|
0 – 2 |
5.7 ± 1.84 |
5.1 ± 1.38 |
4.8 ± 1.08 |
2.2 ± 1.13 * |
2 – 5 |
7.2 ± 1.29 |
7.0 ± 0.86 |
7.2 ± 0.93 |
4.7 ± 1.91 * |
5 – 8 |
7.1 ± 0.98 |
7.0 ± 1.01 |
7.2 ± 0.84 |
7.3 ± 1.26 |
8 – 11 |
6.8 ± 0.85 |
6.9 ± 1.02 |
6.9 ± 0.79 |
7.8 ± 1.12 * |
11 – 14 |
6.9 ± 0.93 |
6.7 ± 0.88 |
6.7 ± 0.81 |
7.4 ± 1.52 |
14 – 17 |
6.7 ± 1.06 |
6.6 ± 0.79 |
6.8 ± 0.84 |
7.6 ± 0.67 * |
17 – 20 |
6.4 ± 0.64 |
6.6 ± 0.68 |
6.6 ± 0.59 |
6.9 ± 0.85 * |
* statistically significant difference with p ≤ 0.05
Table 4 - Average weight of uterus of pregnant females - main study
Group |
Number of pregnant females |
Weight of uterus |
|
0 |
21 |
5.19 ± 0.84 |
|
1 |
23 |
5.51 ± 0.75 |
|
2 |
22 |
5.12 ± 1.14 |
|
3 |
18 |
5.08 ± 1.31 |
|
Table 5 - Number of fetuses - main study
Group |
Number of pregnant females |
Number of fetuses |
|||||
Total
|
in litters min. – max. |
average in litter in 1 female |
females |
males |
dead |
||
0 |
21 |
247 |
2 – 16 |
11.76 ± 2.91 |
5.76 ± 2.28 |
6.00 ± 2.17 |
0 |
1 |
23 |
270 |
6 – 15 |
11.74 ± 1.94 |
6.22 ± 2.00 |
5.52 ± 1.86 |
1 |
2 |
22 |
244 |
5 – 14 |
11.09 ± 2.39 |
6.45 ± 2.09 |
4.64 ± 1.94 |
0 |
3 |
18 |
199 |
1 – 15 |
11.06 ± 3.69 |
5.39 ± 1.91 |
6.38 ± 1.41 |
13* |
*- all fetuses dead from one female (female not accounted in statistical evaluation)
Table 6 - Number of implantations and number or resorptions - main study
Group |
Number of pregnant females |
Average number of corpora lutea per one female |
Number of implantations |
Number of resorptions |
||
total |
in females min - max |
average in one female |
||||
0 |
21 |
13.00 ± 1.61 |
0 |
7 |
0 – 2 |
0.33 ± 0.58 |
1 |
23 |
12.57 ± 1.62 |
0 |
3 |
0 – 1 |
0.13 ± 0.34 |
2 |
22 |
12.55 ± 1.63 |
0 |
7 |
0 – 4 |
0.32 ± 0.89 |
3 |
18 |
13.11 ± 1.91 |
15* |
2 |
0 – 1 |
0.11 ± 0.32 |
*- all implantations from one female (female not accounted in statistical evaluation)
Table 7 - Average weight of fetuses and placenta (g) - main study
Group |
Number of examined fetuses |
Weight of fetuses |
Weight of placenta |
|
with placenta (x) |
without placenta |
|||
0 |
247 |
4.706 ± 0.607 |
3.431 ± 0.535 |
0.535 ± 0.098 |
1 |
270 |
4.627 ± 0.364 |
3.354 ± 0.307 * |
0.535 ± 0.086 |
2 |
244 |
4.884 ± 0.437 * |
3.467 ± 0.442 * |
0.555 ± 0.115 * |
3 |
199 |
4.809 ± 0.604 |
3.338 ± 0.512 * |
0.557 ± 0.180 |
*statistically significant difference with p ≤ 0.05
(x) with placenta and fetal membranes
Table 8 - Average length of fetuses and placenta (cm) - main study
Group |
Number of examined fetuses |
Length of fetuses |
|
with tail |
without tail |
||
0 |
247 |
5,32 ± 0,24 |
3,75 ± 0,20 |
1 |
270 |
5,28 ± 0,23 |
3,74 ± 0,17 |
2 |
244 |
5,30 ± 0,27 |
3,78 ± 0,21 |
3 |
199 |
5,18 ± 0,34 * |
3,69 ± 0,26 * |
* statistically significant difference with p ≤ 0.05
Table 9 - Pathological changes in fetuses - main study
Pathological changes |
Number of fetuses |
|||
Group 0 |
Group 1 |
Group 2 |
Group 3 |
|
Total number of fetuses |
247 |
271 |
244 |
212 |
Number of alive fetuses |
247 |
270 |
244 |
199 |
Number of dead fetuses |
0 |
1[7] |
0 |
13*[3] |
Number of blood hemorrhage |
6 |
6 |
5 |
3 |
Blood hemorrhage on back |
1 [5] |
3 [4, 6, 17] |
5 [3, 6(2), 10(2)] |
3 [6 (3)] |
Blood hemorrhage on metatarsus |
1 [4] |
- |
- |
- |
Blood hemorrhage on thigh |
- |
3 [2(2), 6] |
- |
- |
Blood hemorrhage on shank |
1 [8] |
- |
- |
- |
Blood hemorrhage on arm |
1 [6] |
- |
- |
- |
Blood hemorrhage on forearm |
1 [6] |
- |
- |
- |
Blood hemorrhage on tail |
1 [4] |
- |
- |
- |
Conjoined placenta |
- |
1 [7] ♀** i ♂ |
1 [10] ♂ i ♂ |
1 [22] ♀ i ♂ |
*–all fetuses dead from one female (female not accounted in statistical evaluation)
**– dead fetuses
[ ] –computer numbers of females
( )–number of fetuses in a given female, if more than 1
Table 10 - Pathological changes in skeletal system - main study
Pathological changes
|
Number of fetuses |
|||
Group 0 |
Group 1 |
Group 2 |
Group 3 |
|
No ossification points of sternum |
- |
- |
- |
1 [10] |
1 ossification point of sternum |
- |
- |
- |
2 [10(2)] |
2 ossification points of sternum |
- |
- |
- |
2 [10(2)] |
3 ossification points of sternum |
5 [10(3),14,16] |
1 [21] |
2 [16(2)] |
3 [8(2),12] |
Dislocation of sternum ossification points and rib attachment |
- |
1 [10] |
- |
- |
No ossification points of metatarsus |
- |
- |
- |
1 [10] |
3 ossification points of metatarsus |
- |
- |
- |
4 [10] |
1 ossification point of metacarpus |
- |
- |
- |
1 [10] |
5 ossification points of metatarsus |
7 [25(7)] |
- |
7 [17(7)] |
- |
Total number of all pathological changes (Table 22 and 23) |
18 |
9 |
15 |
18 |
[ ] computer numbers of females
( ) number of fetuses in a given female, if more than 1
Table 11 - Percentage of fetuses with a given number of ossification points of sternum (%) - main study
Group |
Number of examined fetuses |
Number of ossification points |
|||||||
0 |
1 |
2 |
3 |
4 |
5 |
6 |
7 |
||
0 |
129 |
0.0 |
0.0 |
0.0 |
3.88 |
14.73 |
40.31 |
41.08 |
0,0 |
1 |
141 |
0.0 |
0.0 |
0.0 |
0.71 |
14.89 |
54.61 |
29.79 |
0,0 |
2 |
129 |
0.0 |
0.0 |
0.0 |
1.55 |
13.95 |
41.86 |
42.64 |
0,0 |
3 |
105 |
0.95 |
1.90 |
1.90 |
2.86 |
12.38 |
35.24 |
44.77 |
0,0 |
Table 12 - Percentage of fetuses with a given number of ossification points of limbs (%) - main study
Group |
Number of examined fetuses |
Fore limbs Number of ossification points |
Hind limbs Number of ossification points |
||||||||||||
0 |
1 |
2 |
3 |
4 |
5 |
0 |
1 |
2 |
3 |
4 |
5 |
6 |
7 |
||
0 |
129 |
0.0 |
0.0 |
0.0 |
39.53 |
60.47 |
0.0 |
0.0 |
0.0 |
0.0 |
0.0 |
94.57 |
5.43 |
0.0 |
0.0 |
1 |
141 |
0.0 |
0.0 |
0.0 |
58.87 |
41.13 |
0.0 |
0.0 |
0.0 |
0.0 |
0.0 |
100,0 |
0.0 |
0.0 |
0.0 |
2 |
129 |
0.0 |
0.0 |
0.0 |
47.29 |
52.71 |
0.0 |
0.0 |
0.0 |
0.0 |
0.0 |
94.57 |
5.43 |
0.0 |
0.0 |
3 |
105 |
0.0 |
0.95 |
0.0 |
39.05 |
60.00 |
0.0 |
0.95 |
0.0 |
0.0 |
3.81 |
95.24 |
0.0 |
0.0 |
0.0 |
Table 13 - Average number of ossification points of sternum and limbs - main study
Group |
Number of examined fetuses |
Sternum |
Metacarpus of fore limbs |
Metatarsus of hind limbs |
0 |
129 |
5.19 ± 0.83 |
3.60 ± 0.49 |
4.05 ± 0.23 |
1 |
141 |
5.13 ± 0.68 |
3.41 ± 0.49 * |
4,00 ± 0,00 |
2 |
129 |
5.26 ± 0.75 |
3.53 ± 0.50 |
4.05 ± 0.23 |
3 |
105 |
5.09 ± 1.19 |
3.58 ± 0.55 |
3.92 ± 0.43 * |
* statistically significant difference with p ≤ 0.05
Applicant's summary and conclusion
- Conclusions:
- The NOAEL for both maternal toxicity and developmental toxicity was determined to be 50 mg/kg bw/day.
- Executive summary:
A prenatal developmental toxicity study on rats was performed according to OECD guideline 414 and EU method B.31 in order to determine test item influence and possible toxic effects to pregnant females and their fetuses due to treatment during period of gestation. The study was conducted on 100 females divided into four experimental groups: three treated groups and one control group. Every group was consisted of 25 females. From day 0 to 19th of gestation the females of treated group were given by gavage the test item dissolved in corn oil. The following doses of the test item were used: in group 1 – 8.3 mg/kg bw/day, in group 2 – 50 mg/kg bw/day and in group 3 – 300 mg/kg bw/day. Females of the control group (group 0) were given corn oil. During the study, clinical observations for mortality and signs of toxic influence of the test item were performed in females. Body weight and food consumption were recorded. On Day 20 of gestation, the females were killed and caesarean section with gross examination was performed in each female. The following endpoints were determined in each pregnant females in the sighting study and the main study: number of alive and dead fetuses, number of resorptions. After the removing of fetuses each uterus was weighed. The non-gravid uteri and non-gravid horns were overexposed by electric lamp and were stained using ammonium sulphide in order to confirm the non-pregnant status. In the main study the number of corpora lutea was determined in fixed ovaries. The following endpoints were determined in all fetuses: sex, body weight with and without placenta, weight of placenta, body length with and without tail. Each fetus was subjected to gross examination. Half of fetuses from each litter was stained with alizarin and subjected to evaluation of skeleton. The rest of them was fixed in 10% solution of formalin and evaluated for formation of body cavities and internal organs. Based on the results, the dose levels 8.3 and 50 mg/kg bw /day did not indicate any toxic influence on pregnant females. The dose level of 300 mg/kg bw/day caused increased intrauterine mortality and reduction of body weight and body length in fetuses. The dose levels 8.3, 50 and 300 mg/kg bw/day did not cause morphological changes in fetuses of treated females.
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