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EC number: 207-866-0 | CAS number: 498-66-8
- 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
Key value for chemical safety assessment
Effects on fertility
Link to relevant study records
- Endpoint:
- screening for reproductive / developmental toxicity
- Remarks:
- based on test type (migrated information)
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- January 2010 - october 2010
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: GLP guideline study
- Reason / purpose for cross-reference:
- reference to same study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 422 (Combined Repeated Dose Toxicity Study with the Reproduction / Developmental Toxicity Screening Test)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Limit test:
- no
- Species:
- rat
- Strain:
- Wistar
- Sex:
- male/female
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River, 97633 Sulzfeld, Germany
- Age at study initiation: 9-10 weeks
- Weight at study initiation: males: 289 g; females: 186 g(means)
- Housing:
- Diet: ad libitum
- Water: tap water ad libitum
- Acclimation period: 7 to 10 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22°C ± 3°C
- Humidity (%): 30-70%
- Air changes (per hr): approx. 10
- Photoperiod (hrs dark / hrs light): 12/12
IN-LIFE DATES: From: pre-mating phase To: termination - Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS:
A stock solution containing 125 mg/L vehicle was prepared in a water bath (60°C).Two serial dilutions (1+1, v/v) were prepared from the high dose with equal amounts of vehicle. The solutions were stored refrigerated (5°C± 3°C) under agron until application.
VEHICLE
- Justification for use and choice of vehicle (if other than water): very low water solubility of the test substance
- Concentration in vehicle: 125, 52.5, and 31.25 g test substance/L
- Amount of vehicle (if gavage): dose volume was 4 mL/kg bw
- Supplier: Sigma
- Purity: 100% - Details on mating procedure:
- During the mating phase (up to 14 days), the female were placed with the same male until indications for mating were detected. Each morning, the females were examined for the presence of sperm or a vaginal plug. Day 0 of pregnancy was defined as the day a vaginal plug or sperm was
found. Females showing no evidence of copulation were regrouped with proven sires for a second or third mating phase. - Analytical verification of doses or concentrations:
- no
- Duration of treatment / exposure:
- 61 ± 10.0 days (because a second and third mating was necessary; report, page 24/36)
- Frequency of treatment:
- 7 per week
- Details on study schedule:
- Males were dosed daily for a minimum of four weeks, including the day before the scheduled termination of the in-life phase. This included a minimum of two weeks of dosing prior to mating and continued throughout the mating period until the study was terminated. Females were dosed daily, including two weeks prior to mating, covering at least two complete oestrous cycles, the variable time to conception, the duration of pregnancy and at least four days after delivery, up to and including the day before scheduled termination of the in-life phase. Therefore the duration of the study following acclimatisation depended on the female performance: 14 days pre-mating, up to 14 days until mating, an average of 21 days
of gestation, and a minimum of 4 days of lactation. - Remarks:
- Doses / Concentrations:
125, 250, 500 mg/kg bw and day
Basis:
actual ingested - No. of animals per sex per dose:
- 12
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- Males were dosed daily for a minimum of four weeks, including the day before the scheduled termination of the in-life phase. This included a minimum of two weeks of dosing prior to mating and continued throughout the mating period until the study was terminated.
Females were dosed two weeks prior to mating, the variable time to conception, the duration of pregnancy and at least four days after delivery, up to and including the day before scheduled termination of the in-life phase. Females showing no evidence of copulation were re-grouped with proven sires for further mating phases. Dosing was continued in both sexes during the mating period. - Positive control:
- not required
- Parental animals: Observations and examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations checked in table [No.?] were included.
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: once before beginning of treatment and weekly thereafter
BODY WEIGHT: Yes
- Time schedule for examinations: once before beginning of treatment and at least weekly thereafter. Females: dyas 0, 7, 14, 20, within 24 h post parturition and 4 days post partum.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: No; not required.
Food consumption was determined per cage at least one weakly.
FOOD EFFICIENCY:
- Body weight gain in kg/food consumption in kg per unit time X 100 calculated as time-weighted averages from the consumption and body weight gain data: No
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No; not required.
Food consumption was determined per cage twice weakly.
OPHTHALMOSCOPIC EXAMINATION: No
HAEMATOLOGY: Yes
- Time schedule for collection of blood: day 14
- Anaesthetic used for blood collection: No
- Animals fasted: No data
- How many animals: all
- Parameters checked in table No. 3 were examined.
CLINICAL CHEMISTRY: Yes
- Time schedule for collection of blood: day 14
- Anaesthetic used for blood collection: No
- Animals fasted: No data
- How many animals: all
- Parameters checked in table No. 3 were examined.
URINALYSIS: No
NEUROBEHAVIOURAL EXAMINATION: No
- Time schedule for examinations: once before beginning of application and once during the last week of exposure.
- Dose groups that were examined: no data
- Battery of functions tested: sensory activity / grip strength (cf. report section 3.2.4; results Appendix, pages 14-15) - Oestrous cyclicity (parental animals):
- no data
- Sperm parameters (parental animals):
- Parameters examined in male parental control and high dose groups :
testis weight, epididymis weight, sperm count in epididymides - Litter observations:
- PARAMETERS EXAMINED
The following parameters were examined in offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain days 0 to 4 - Postmortem examinations (parental animals):
- SACRIFICE
- Male animals: All surviving animals [describe when, e.g. as soon as possible after the last litters in each generation were produced.]
- Maternal animals: All surviving animals [describe when, e.g. after the last litter of each generation was weaned.]
GROSS NECROPSY
- Gross necropsy consisted of
RTable: Parameters examined at necropsy
No. Tissue/organ Procedure1
1 Gross lesions P 12 Adrenal gland P, W
2 Oesophagus P 13 Urinary bladder P
3 Trachea and thyroid P 14 Testes P, W
4 Stomach P 15 Epididymides P, W
5 Thymus P, W 16 Prostate / uterus / ovary P
6 Liver P, W 17 Heart P, W
7 Spleen P, W 18 Lung P
8 Duodenum P 19 Peripheral nerve P
9 Jejunum P 20 Bone marrow P
10 Ileum (with Peyer’s
patches) P 21 Sternum P
11 Cecum P 22 Spinal cord P
12 Adrenal gland P, W
13 Urinary bladder P
14 Testes P, W
16 Prostate / uterus / ovary P
17 Heart P, W
18 Lung P
19 Peripheral nerve P
20 Bone marrow P
21 Sternum P
22 Spinal cord P
23 Colon P
24 Rectum P
25 Lymph nodes (intestinal area, axillary) P
26 Kidney P, W
27 Whole brain W
28 Cerebrum P
29 Cerebellum P
30 Pons P
1: P = preservation, W = weight determination
HISTOPATHOLOGY / ORGAN WEIGHTS
Cf. table above - Postmortem examinations (offspring):
- SACRIFICE
Offspring were sacrificed at 4 days of age. These animals were subjected to external postmortem examinations. - Statistics:
- Spreadsheet calculations were performed using Microsoft® Excel® 2004 for Mac, Version 11.5.8. In addition, the statistical software Graph Pad Prism for Mac, Version 5.01c was used to calculate detailed column statistics (minimum / maximum data, 75% percentiles, standard error, upper and lower confidence interval 95%).
The significance of differences between the vehicle only and treated groups was analysed using various methods which included Bartlett's test for variances, followed either by a One-Way ANOVA (Analysis of variance), followed by a Dunnett's t-test, or followed by a Bartlett’s test for equal variances and a Kruskall-Wallis test. See the report section 3.5.2 for a full description of the statistical methods used. A decision tree is depicted on page 33 of the Appendix. - Reproductive indices:
- Not calculated
- Offspring viability indices:
- Not calculated
- Clinical signs:
- no effects observed
- Description (incidence and severity):
- Tendency of slightly decreased body weights in male and female high dose groups.
- Body weight and weight changes:
- effects observed, treatment-related
- Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Organ weight findings including organ / body weight ratios:
- effects observed, treatment-related
- Histopathological findings: non-neoplastic:
- effects observed, treatment-related
- Description (incidence and severity):
- Hyaline droplet formation was noted in the renal proximal tubular epithelium of almost all male rats from the high dose group, and to a lesser degree in most male rats of the low and intermediate dose groups. No effects on male and female reproduction or
- Other effects:
- no effects observed
- Reproductive function: oestrous cycle:
- not examined
- Reproductive function: sperm measures:
- no effects observed
- Reproductive performance:
- no effects observed
- Dose descriptor:
- NOAEL
- Remarks:
- reproduction
- Effect level:
- 500 mg/kg bw/day (actual dose received)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: no change compared to controls
- Clinical signs:
- no effects observed
- Mortality / viability:
- no mortality observed
- Body weight and weight changes:
- no effects observed
- Sexual maturation:
- not examined
- Organ weight findings including organ / body weight ratios:
- not examined
- Gross pathological findings:
- no effects observed
- Histopathological findings:
- not examined
- Dose descriptor:
- NOAEL
- Remarks:
- reproduction, viability
- Generation:
- F1
- Effect level:
- 500 mg/kg bw/day (actual dose received)
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: no treatment-related change compared to controls
- Reproductive effects observed:
- not specified
- Conclusions:
- In a repeated dose/reproduction screening study, norbornene had no effect on parental and progeny reproduction parameters of male and female rats. The NOAEL for toxicity to reproduction was therefore 500 mg/kg bw and day, the highest tested dose.
- Executive summary:
In a combined repeated dose/reproduction toxicity study (performed according to OECD TG 422 and under GLP conditions), male and female Wistar rats (12 per sex and dose) received norbornene at dose levels of 125, 250, and 500 mg/kg bw and day by oral gavage. Vehicle controls received corn oil only.
There were no mortalities or clinical signs of toxicity that were attributable to the test substance. The NOAEL for local and systemic toxicity was 500 mg/kg bw and day for both sexes in this study (cf. section 7.5.1).
Regarding reproduction parameters, norbornene treatment had no effect on the following parameters:
Duration of pregnancy; the number of dams with live pups; the mean number of alive pups at day zero and at day four post partum; the mean numbers per dam of corpora lutea, the mean numbers per dam of implantations, as well as live pups. The mean litter mass at day o and day 4 were normal for Wistar rats. A statistically significant increase of corpora lutea was detected in dams of the medium dose group but this was within the normal biological range. Of the five stillborns found in total, two were delivered by dams of the low dose group and three by dams of the medium dose group (Tab. 6, attached document); this was considered to be incidental. The sex-ratio of pups was not affected. The mean pup body weight was within the normal range for Wistar rats, and no statistically significant differences were detected between the dose groups. Data of the numbers of abnormal pups born, or the loss of offspring (pre-implantation, pre-natal and post-natal loss) were normal for rats of this strain and age.
Thus, norbornene had no effect on reproduction and development of male and female Wistar rats (12 rats per sex and dose) exposed to the test substance at 125, 250, and 500 mg/kg bw and day under the conditions of this combined repeated dose/reproduction toxicity screening test which was performed according to OECD TG 422 and under GLP conditions (Vivo, 2010).
This study is considered to be valid and suitable for assessment. The NOAEL for local and systemic toxicity was therefore 500 mg/kg bw and day for both sexes. The NOAEL for toxicity to reproduction and development was also 500 mg/kg bw and day under the conditions of this study.
Reference
All relevant raw data and calculated data were documented within the appendix.
During first mating (Tab. 5, attached document), evidence of copulation in total numbers was similar between all groups and was found within the first five days for all but one female of the low dose group. During second mating, no apparent differences occurred between the dose groups regarding the time point for evidence of copulation, which was within the first five days in all cases.
Pregnancy lasted 22 days (n=22) or 23 days (n=20) and no apparent differences occurred between the dose groups. No differences between the dose groups were detectable regarding the number of dams with live young born at day zero (d0, day of birth), the mean number of alive pups at day four (d4) of lactation, the mean numbers per dam of corpora lutea, the mean numbers per dam of implantations, as well as live pups. The mean litter mass at d0 and d4 were normal for Wistar rats. A statistically significant increase of corpora lutea was detected in dams of the medium dose group. This effect was estimated to be not test item related but within the normal biological range. Of the five stillborns found in total, two were delivered by dams of the low dose group and three by dams of the medium dose group (Tab. 6). No bias was observed in the sex-ratio data. In general, mean pup body mass of rats born in the course of the present study was within normal range for Wistar rats and no statistically significant differences were detected between the dose groups. Data of the numbers of abnormal pups born, or the loss of offspring (pre-implantation, pre-natal and post-natal loss) were normal for rats of this strain and age.
Effect on fertility: via oral route
- Endpoint conclusion:
- no adverse effect observed
- Dose descriptor:
- NOAEL
- 500 mg/kg bw/day
- Study duration:
- subchronic
- Species:
- rat
- Quality of whole database:
- High quality screening study (OECD 422 under GLP conditions)
Additional information
In a combined repeated dose/reproduction toxicity study (performed according to OECD TG 422 and under GLP conditions), male and female Wistar rats (12 per sex and dose) received norbornene at dose levels of 125, 250, and 500 mg/kg bw and day by oral gavage. Vehicle controls received corn oil only.
There were no mortalities or clinical signs of toxicity that were attributable to the test substance. The NOAEL for local and systemic toxicity was 500 mg/kg bw and day for both sexes in this study (cf. section 7.5.1).
Regarding reproduction parameters, norbornene treatment had no effect on the following parameters:
Duration of pregnancy; the number of dams with live pups; the mean number of alive pups at day zero and at day four post partum; the mean numbers per dam of corpora lutea, the mean numbers per dam of implantations, as well as live pups. The mean litter mass at day o and day 4 were normal for Wistar rats. A statistically significant increase of corpora lutea was detected in dams of the medium dose group but this was within the normal biological range. Of the five stillborns found in total, two were delivered by dams of the low dose group and three by dams of the medium dose group (Tab. 6, attached document); this was considered to be incidental. The sex-ratio of pups was not affected. The mean pup body weight was within the normal range for Wistar rats, and no statistically significant differences were detected between the dose groups. Data of the numbers of abnormal pups born, or the loss of offspring (pre-implantation, pre-natal and post-natal loss) were normal for rats of this strain and age.
Thus, norbornene had no effect on reproduction and development of male and female Wistar rats (12 rats per sex and dose) exposed to the test substance at 125, 250, and 500 mg/kg bw and day under the conditions of this combined repeated dose/reproduction toxicity screening test which was performed according to OECD TG 422 and under GLP conditions (Vivo, 2010).
This study is considered to be valid and suitable for assessment. The NOAEL for local and systemic toxicity was therefore 500 mg/kg bw and day for both sexes. The NOAEL for toxicity to reproduction and development was also 500 mg/kg bw and day under the conditions of this study.
Short description of key information:
The NOAEL for reproduction toxicity was 500 mg/kg bw and day, the highest tested dose, in male and female parental rats in a screening study (OECD TG 422, GLP, oral gavage).
Justification for selection of Effect on fertility via oral route:
High quality screening study (OECD 422 under GLP conditions)
Effects on developmental toxicity
Description of key information
The NOAEL of Norbornene in an OECD 414 study using rats was established at 300 mg/kg bw (gavage), based on highly significant foetal weight reduction at all tested dose levels (150, 300, and 600 mg/kg bw) that reached >10% in the high dose group. Maternal toxicity and teratogenicity were not seen in any dose group.
Link to relevant study records
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 21 Nov 2014 - 07 April2015
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: GLP guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- Version / remarks:
- (Adopted 22 January 2001)
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Limit test:
- no
- Species:
- rat
- Strain:
- Wistar
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Source: Charles River, Sulzfeld, Germany
- Age at study initiation: 8-12 weeks
- Weight at study initiation: approx. 200 g
- Fasting period before study: no data
- Housing: in groups of four animals in macrolon cages
- Diet (e.g. ad libitum): Altromin, No 1324 TPF, maintenance diet for rats and mice, breeding diet No. 1314 TPF, ad libitum
- Water: sterilised community tab water, ad libitum
- Acclimation period:
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22±3
- Rel. humidity (%): 30-70
- Air changes (per hr): 10
- Photoperiod (hrs dark / hrs light): 12/12 - Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS:
Heat corn oil (measured amount) and test item to 60°C for 15 minutes. Pour test item into corn oil to produce the test solution, determine net weight and volume of the test item, calculate concentration (mg/mL) of the stock solution. Add required corn oil to produce the highest application dose for the 600 mg/kg dose level. Heat for one minute and vortex for 15 seconds. Then, produce two serial dilutions (1:1) for the mid and the low dose level, respectively. Store test materials at 5°C under argon until application.
DIET PREPARATION
n. a.
VEHICLE
- Justification for use and choice of vehicle: corn oil was used because of the low solubility of Norbornene in water
- Concentration in vehicle: 150/75/36.5 mg/mL
- Amount of vehicle: 4 mL/kg bw
- Source: Sigma, catalogue No. C8267
- Purity: 100% (: - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- A validated GC/FID method was used for the determination of Norbornene in corn oil. Samples were diluted with i-hexane, and toluene was used as internal standard. 1 µL was injected split less. Instruments: column: ZB-624, ID 0.32 mm, 1.8µm; Temperature 40°C, 13 min isothermal,. Gas: hydrogen. Inlet: 250°C split less; Detector FID, 260°C; Flow 2 mL/min. The ratio of the areas test item/toluene was used for quantification. Retention times were 5.72 min for Norbornene and 9.21 min for toluene. Recoveries were in the range 103 - 112 % for solutions containing 37,5, 75, and 150 mg/mL. Linear calibration curve; y=0.00886x + 0.00731; r²= 0.99998. Further details: Appendix B
- Details on mating procedure:
- Animals were mated at the breeding facility; no details reported (section 3.1 of the study report)
- Duration of treatment / exposure:
- gestational days 6 through 20
- Frequency of treatment:
- daily
- Duration of test:
- 24 days, including acclimatisation
- Remarks:
- Doses / Concentrations:
0, 150, 300, and 600 mg/kg bw
Basis:
analytical conc. - No. of animals per sex per dose:
- Planned: 24/group
Successfully mated: 18/23/21/21 in controls/low/mid/high dose group, respectively - Control animals:
- yes, concurrent vehicle
- Details on study design:
- Dose selection rationale: dose levels were selected based on the results as follows:
-OECD 422 combined repeated dose and reproduction toxicity study, oral gavage: minor effects but no clear toxicity pattern at the highest tested dose of 500 mg/kg bw.
-OECD 413 90-day inhalation study: treatment related changes in the ovaries of females exposed to 4.98 mg/m³. No effects at 2.02 gm³, i.e. the NOAEL was 582 mg/kg bw in this study. - Maternal examinations:
- CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
- Cage side observations were not included.
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily
BODY WEIGHT: Yes
- Time schedule for examinations: days 3, 6, 9, 12, 15, 18, and 21
FOOD CONSUMPTION: Yes
- Food consumption for each animal group determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes
- Time schedule for examinations: days 3, 6, 9, 12, 15, 18, and 21
WATER CONSUMPTION: No
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: macroscopical examination following sacrifice. Uterine weight (not obtained from animals found dead during the study). Uterine content was examined for numbers of embryonic or foetal deaths, viable fetuses, and scars. In the ovaries, the number of corpora lutea was determined for pregnant animals. - Ovaries and uterine content:
- The ovaries and uterine content were 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 - Fetal examinations:
- - External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter
- Skeletal examinations: Yes: half per litter
- Head examinations: No - Statistics:
- Descriptive statistics: the arithmetic mean and the standard deviation was calculated for all grouped numerical data originating from monitoring the body weight, food consumption and gross pathology (fetal and placenta weights). Where appropriate, detailed column statistics were applied (minimum / maximum data, 25% quantiles, standard error, upper and lower confidence interval 95%).
Inductive statistics: for basic analysis the respective test item groups were compared to the vehicle group by assessing of statistical significance using a two-tailed unpaired Student´s t-test. For all calculations, the significance level was set to 0.05. In case the basic analysis returned a statistical significance additional inductive statistical analysis was applied as outlined in the main decision tree (see appendix). Most statistical hypotheses in this study were best characterised as “many to one”– comparing a vehicle control vs. three treatment groups, respectively. Therefore the adequate analysis method was a one-way ANOVA (Analysis of variance), followed by a post hoc Dunnett´s t-test. Analysis of body weight data includes the influence of two different categorical independent variables – time and dose groups. Therefore, analysis by two-way ANOVA, followed by Bonferroni post-hoc test, was assumed as the most adequate analysis method for this set of data. For all calculations, the significance level was set to 0.05. These further inductive statistics were performed using Graph Pad Prism for Mac, Version 5. All statistical data are documented in the appendix of the study report. - Indices:
- mean pre- and post-implantation loss
- Historical control data:
- no data
- Details on maternal toxic effects:
- Maternal toxic effects:no effects
Details on maternal toxic effects:
Clinical signs: salivation was seen in all treated groups. The incidence and severity increased with the dose. 2two rats were occasionally affected in the low dose group; in the mid dose group, single animals showed this sign, starting from day 9, and all animals were affected on day 20; at the high dose, 3 animals showed this signs on day 9, and almost all animals from day 11 onwards.
Mortalities: two mid dose rats dies immediately after dosing due to an application error. One high dose rat was found dead more than 8 hours after death, the cause of death could not be established.
Body weights: treatment had no effect on the gravid rat body weight of either test group
Food consumption: food consumption was comparable across all dose groups, i.e. no treatment-related effect was noted
Necropsy: a pale pancreas was noted in 19/20 high dose and in 2/20 mid dose dams. This observation was not noted in the vehicle and in the low dose groups. The macroscopic examination revealed no other pathological findings. - Dose descriptor:
- NOAEL
- Effect level:
- 600 mg/kg bw/day (actual dose received)
- Based on:
- act. ingr.
- Basis for effect level:
- other: maternal toxicity
- Dose descriptor:
- NOAEL
- Effect level:
- 300 mg/kg bw/day
- Based on:
- act. ingr.
- Basis for effect level:
- other: developmental toxicity
- Details on embryotoxic / teratogenic effects:
- Embryotoxic / teratogenic effects:yes. Remark: developmental effects
Details on embryotoxic / teratogenic effects:
Uterus contents: the mean gravid uterus weight (with ovaries) was comparable across all groups.
Ovaries: the number of corpora lutea was comparable across all groups.
Implantations: the number was comparable across all groups.
Number of live fetuses: comparable across all groups.
Dead fetuses: no dead foetus seen in any group.
Early resorptions: mean number increased at the high dose level (0.8 versus 0.2 in the control), but not statistically significantly increased.
Pre- and post-implantation losses: minor differences lacking statistical significance. Two of the control animals showed an unusual high incidence of pre-implantation losses (10 and 12). Re-calculation without these two animals did also not reveal significant differences between the animal groups.
Foetal weight: weight at birth was significantly (p<0.001) reduced in male and female fetuses in all dose groups. The effect was dose-dependent and most prominent at the high dose [-11.4% (males and -11.8% (females)]. Birth weight reductions >10% are considered to be adverse. - Dose descriptor:
- NOAEL
- Effect level:
- 600 mg/kg bw/day (actual dose received)
- Based on:
- act. ingr.
- Basis for effect level:
- other: teratogenicity
- Abnormalities:
- not specified
- Developmental effects observed:
- not specified
- Conclusions:
- The NOAEL of Norbornene in an OECD 414 study using rats was established at 300 mg/kg bw (gavage), based on highly significant foetal weight reduction at all tested dose levels (150, 300, and 600 mg/kg bw) that reached >10% in the high dose group. Maternal toxicity and teratogenicity were not seen in any dose group.
- Executive summary:
The developmental toxicity and teratogenicity of Norbornene was studied in time-mated Wistar rats according to the OECD 414 test guideline and under GLP conditions. Norbornene, dissolved in corn oil, was administered by oral gave at dose levels of 0, 150, 300 and 600 mg/kg bw, based on the results of preceding studies [OECD 422 (oral) and 413 (inhalation)] which gave NOAEL values of 500 and 582 mg/kg bw, respectively.
The NOAEL of Norbornene in the OECD 414 study using rats was established at 300 mg/kg bw (gavage), based on highly significant foetal weight reduction at all tested dose levels (150, 300, and 600 mg/kg bw) that reached >10% in the high dose group. Maternal toxicity and teratogenicity were not seen in any dose group. A significant increase of supernummary ribs was seen in fetuses of the high dose group (40% versus 8% in the control) and on litter basis (90% vs. 39% in the vehicle control. Therefore, this finding is considered to be test-item related, but not teratogen.
Overall the NOAEL of Norbornene in the Wistar rat was 600 mg/kg bw for maternal toxicity and teratogenicity, and 300 mg/kg bw for developmental toxicity.
The study is considered valid and suitable for assessment.
Reference
Table 1: Mean number of corpora lutea
|
Corpora lutea (n) |
|||
|
Left ovary |
Right ovary |
Total |
|
Dose group |
Mean |
Mean |
Mean |
SD |
600 mg/kg |
6.9 |
6.6 |
13.5 |
1.7 |
300 mg/kg |
6.8 |
6.6 |
13.3 |
1.7 |
150 mg/kg |
6.3 |
6.9 |
11.1 |
2.4 |
Vehicle |
6.5 |
6.7 |
13.2 |
1.9 |
Table 2: Mean number of implantations
|
Corpora lutea (n) |
|||
|
Left uterus horn |
Right uterus horn |
Total |
|
Dose group |
Mean |
Mean |
Mean |
SD |
600 mg/kg |
6.1 |
6.2 |
12.3 |
1.6 |
300 mg/kg |
5.7 |
6.2 |
11.9 |
1.6 |
150 mg/kg |
5.7 |
6.1 |
11.8 |
1.2 |
Vehicle |
5.5 |
5.3 |
10.8 |
3.1 |
Table 3: Mean number of foetuses and resorptions
|
Corpora lutea |
Implantation sites |
Live fetuses |
Early resorptions |
Scar |
Dead fetuses |
Dose group |
Mean |
Mean |
Mean |
Mean |
Mean |
Mean |
600 mg/kg |
13.5 |
12.3 |
11.5 |
0.8 |
0.0 |
0.0 |
300 mg/kg |
13.3 |
11.9 |
11.5 |
0.2 |
0.0 |
0.0 |
150 mg/kg |
11.1 |
11.8 |
11.5 |
0.2 |
0.0 |
0.0 |
Vehicle |
13.2 |
10.8 |
10.2 |
0.2 |
0.0 |
0.0 |
Vehicle * |
13.1 |
11.8 |
11.1 |
0.2 |
0.0 |
0.0 |
Vehicle*: calculation that excludes two dams with unusually high number of pre-implantation losses (10 and 12, respectively)
Table 4: Indices of implantation losses
|
Pre-implantation loss (CL-IS)
|
Post-implantation loss (IS-LF) |
Pre-implantation loss per group |
Post-implantation loss per group |
Dose group |
Mean |
Mean |
Mean |
Mean |
600 mg/kg |
1.2 |
0.8 |
9.76 |
7.05 |
300 mg/kg |
1.4 |
0.4 |
11.76 |
3.73 |
150 mg/kg |
1.3 |
0.3 |
11.02 |
2.64 |
Vehicle |
2.4 |
0.7 |
22.22 |
6.55 |
Vehicle * |
1.3 |
0.7 |
11.1 |
6.55 |
Legend:
Vehicle*: calculation that excludes two dams with unusually high number of pre-implantation losses (10 and 12, respectively)
CL
= Corpora lutea
IS = Implantation site
LF = Life fetuses
Indices were calculated as follows:
Pre implantation loss: mean no. of corpora lutea – mean no. of implantations
Mean no. of resorptions: No. of resorptions / no. of litters
Mean pre implantation loss per group: (mean no. of corpora lutea – mean no. of implantations) * 100 /mean no. of implantations
Mean post implantation loss (%): (Total no. resorptions + total no. of dead fetuses) * 100 / no. of fetuses.
All calculations and statistical tests are documented within the appendix of the report.
Table 5: mean fetal birth weight and weight changes
|
Mean weight [g] |
||||
|
Both sexes |
Males |
Females |
Weight change [%] Versus controls |
|
Dose group |
Males |
Females |
|||
600 mg/kg |
4.76 |
4.91 |
4.65 |
-11.74 |
-11.8 |
300 mg/kg |
5.01 |
5.11 |
4.90 |
-7.8 |
-7.0 |
150 mg/kg |
506 |
5.19 |
4.94 |
-6.3 |
-6.3 |
Vehicle |
5.38 |
5.54 |
5.27 |
|
|
Effect on developmental toxicity: via oral route
- Endpoint conclusion:
- adverse effect observed
- Dose descriptor:
- NOAEL
- 300 mg/kg bw/day
- Study duration:
- subacute
- Species:
- rat
- Quality of whole database:
- Valid OECD 414 guideline study (reliabiliy 1) in Wistar rats
Additional information
In a combined repeated dose/reproduction toxicity study (performed according to OECD TG 422 and under GLP conditions), male and female Wistar rats (12 per sex and dose) received norbornene at dose levels of 125, 250, and 500 mg/kg bw and day by oral gavage. Vehicle controls received corn oil only.
There were no mortalities or clinical signs of toxicity that were attributable to the test substance. The NOAEL for local and systemic toxicity was 500 mg/kg bw and day for both sexes in this study (cf. section 7.5.1).
Regarding reproduction parameters, norbornene treatment had no effect on the following parameters:
Duration of pregnancy; the number of dams with live pups; the mean number of alive pups at day zero and at day four post partum; the mean numbers per dam of corpora lutea, the mean numbers per dam of implantations, as well as live pups. The mean litter mass at day o and day 4 were normal for Wistar rats. A statistically significant increase of corpora lutea was detected in dams of the medium dose group but this was within the normal biological range. Of the five stillborns found in total, two were delivered by dams of the low dose group and three by dams of the medium dose group (Tab. 6, attached document); this was considered to be incidental. The sex-ratio of pups was not affected. The mean pup body weight was within the normal range for Wistar rats, and no statistically significant differences were detected between the dose groups. Data of the numbers of abnormal pups born, or the loss of offspring (pre-implantation, pre-natal and post-natal loss) were normal for rats of this strain and age.
Thus, norbornene had no effect on reproduction and development of male and female Wistar rats (12 rats per sex and dose) exposed to the test substance at 125, 250, and 500 mg/kg bw and day under the conditions of this combined repeated dose/reproduction toxicity screening test which was performed according to OECD TG 422 and under GLP conditions (Vivo, 2010).
This study is considered to be valid and suitable for assessment. The NOAEL for local and systemic toxicity was therefore 500 mg/kg bw and day for both sexes. The NOAEL for toxicity to reproduction and development was also 500 mg/kg bw and day under the conditions of this study.
The developmental toxicity and teratogenicity of Norbornene was studied in time-mated Wistar rats according to the OECD 414 test guideline and under GLP conditions. Norbornene, dissolved in corn oil, was administered by oral gave at dose levels of 0, 150, 300 and 600 mg/kg bw, based on the results of preceding studies [OECD 422 (oral) and 413 (inhalation)] which gave NOAEL values of 500 and 582 mg/kg bw, respectively.
The NOAEL of Norbornene in the OECD 414 study using rats was established at 300 mg/kg bw (gavage), based on highly significant foetal weight reduction at all tested dose levels (150, 300, and 600 mg/kg bw) that reached >10% in the high dose group. Maternal toxicity and teratogenicity were not seen in any dose group. A significant increase of supernummary ribs was seen in fetuses of the high dose group (40% versus 8% in the control) and on litter basis (90% vs. 39% in the vehicle control. Therefore, this finding is considered to be test-item related, but not teratogen.
Overall the NOAEL of Norbornene in the Wistar rat was 600 mg/kg bw for maternal toxicity and teratogenicity, and 300 mg/kg bw for developmental toxicity in the OECD 414 study, which is considered valid and suitable for assessment.
It is noteworthy that, in contrast to the 90 -day inhalation study, in the OECD 414 oral study no effect on the ovaries was seen (no increased weight or increased number of corpora lutea). The reason is presently unclear but may be related to the short exposure period (gestational day 6-20) compared to the 90 day exposure period of the OECD 413 study. Additionally, toxicokinetic differences are expected depending of the route of exposure. Further, as no effect on the foetal birth weight was seen in the OECD 422 study in contrast to the OECD 414 study, it may be speculated that induction of the xenobiotics metabolising enzymes in the OECD 422 study might have led to an increased metabolism and excretion
Justification for selection of Effect on developmental toxicity: via oral route:
Valid OECD 414 guideline study (reliability 1)
Justification for classification or non-classification
The single adverse effect observed in the OECD 414 study was reduced fetal body weight, potentially indicating delayed development. This effect has not been observed in fetuses of the earlier OECD 422 study conducted on the same strain of rats and by the same laboratory, introducing some element of doubt on the significance of the observed effect.
As a precautionary apporach the following classification according to 1272/2008/EC is proposed:
Reproductive toxicity category 2, H361
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