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EC number: 202-790-4 | CAS number: 99-82-1
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data
Developmental toxicity / teratogenicity
Administrative data
- Endpoint:
- developmental toxicity
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 11 November 2015 to 4 December 2015
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 016
- Report date:
- 2016
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 414 (Prenatal Developmental Toxicity Study)
- Qualifier:
- equivalent or similar to guideline
- Guideline:
- other: - OECD Guidance No. 43 on Mammalian Reproductive Toxicity Testing and Assessment, 24th July 2008
- GLP compliance:
- yes (incl. QA statement)
- Limit test:
- no
Test material
- Reference substance name:
- 1-isopropyl-4-methylcyclohexane
- EC Number:
- 202-790-4
- EC Name:
- 1-isopropyl-4-methylcyclohexane
- Cas Number:
- 99-82-1
- Molecular formula:
- C10H20
- IUPAC Name:
- 1-methyl-4-(propan-2-yl)cyclohexane
- Test material form:
- liquid
- Details on test material:
- Identification: para-Menthane
Chemical name Cyclohexane,1-methyl-4-(1-methylethyl)/1-isopropyl-4-methylcyclohexane
Appearance: colourless liquid
Constituent 1
Test animals
- Species:
- rat
- Strain:
- Wistar
- Details on test animals or test system and environmental conditions:
- TEST ANIMALS
- Strain: Hsd. Han: WIST Rats
- Source: TOXI-COOP ZRT. 1103 Budapest, Cserkesz u. 90.
- Age at study initiation: females 10 - 11 weeks (males for mating 11 - 12 weeks)
- Weight at study initiation: 166 - 217 g
- Fasting period before study: none
- Housing: pre-mating period: 1-3 females /cage, 2 males/cage; during mating hours:1 male with 1- 3 females; during pregnancy: 2-3 sperm positive females /cage
- Diet: ssniff® SM R/M-Z+H complete diet (ssniff Spezialdiäten GmbH, D-59494 Soest Germany) ad libitum
- Water: tap water ad libitum
- Acclimation period: 7 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 - 22 °C
- Humidity (%): 34 - 47%
- Air changes (per hr): >10/hour
- Photoperiod (hrs dark / hrs light): 12/12
Administration / exposure
- Route of administration:
- oral: gavage
- Vehicle:
- corn oil
- Details on exposure:
- PREPARATION OF DOSING SOLUTIONS: test item formulations was made with a frequency of up to 7 days, using a magnetic stirrer and stored at room temperature
VEHICLE: corn oil
- Justification for use and choice of vehicle (if other than water): low solubility of the test item in water
- Concentration test substance in vehicle: 0, 50, 150 and 500 mg/mL
- Dosing volume (if gavage): 2 mL
- Lot/batch no.: MKBV2080V - Analytical verification of doses or concentrations:
- yes
- Details on analytical verification of doses or concentrations:
- Five samples were taken from different places in the dosing formulation on day 7 and 18 of the study. Samples were diluted with n-hexane in duplicate except for the control. A volume of 0.5 mL of the internal standard solution was added before the samples were filled up to 10 mL final volume. Samples were stored overnight and analysed the next day.
GC/FID
Column: Rtx-502.2, 60 m x 0.25 mm x 1.4 µm, No: 1023994
Temperature program: 125 °C to 150 °C at 2 °C/min
2 min hold
150 °C to 180 °C at 20 °C/min
2 min hold
Injector: 200 °C
Detector: FID, 200 °C
Carrier gas: nitrogen, 2 mL/min, constant flow
Split ratio: 20
Injection volume: 0.5 µL
Retention times: para-Menthane 13.5 - 13.6 and 14.4 - 14.5 min, 1-Octanol: 16.9 - 17.0 min
Evaluation: Sum of the areas of para-Menthane peaks is calculated and divided by 1-octanol peak area. This ratio is used for the computations.
Linearity over 0.1 – 3 mg/mL (r² >= 0.998); repeatability 0.6 - 1.0% (7 injections at 500 and 1 mg/L)
Recovery in corn oil: 96 ±4% at 1 mg/L; at 500 mg/L 95 ±5%
Stability over 14 days: 101 - 104% (room temperature) 98 - 104% (at 5 ±3 °C) measured at 1 and 500 mg/L in corn oil
LOQ: 0.1 mg/L - Details on mating procedure:
- - Impregnation procedure: cohoused
- M/F ratio per cage: 1 male/1 - 3 females
- Length of cohabitation: 2 - 4 hours
- Proof of pregnancy: vaginal plug / sperm in vaginal smear referred to as day 0 of pregnancy - Duration of treatment / exposure:
- day 5 - 19 of gestation
- Frequency of treatment:
- daily
- Duration of test:
- 20 days
Doses / concentrationsopen allclose all
- Dose / conc.:
- 100 mg/kg bw/day (actual dose received)
- Remarks:
- measured concentration 94% of nominal
- Dose / conc.:
- 300 mg/kg bw/day (nominal)
- Remarks:
- measured concentration 100 - 103% of nominal
- Dose / conc.:
- 1 000 mg/kg bw/day (nominal)
- Remarks:
- measured concentration 96 - 100% of nominal
- No. of animals per sex per dose:
- 22 sperm positive females/group
- Control animals:
- yes, concurrent vehicle
- Details on study design:
- Dose selection rationale: based on the results of a DRF test where the substance at the dose level of 1000 mg/kg bw/day caused clinical signs such as digging in the bedding and salivation but no effects on the body weight and food consumption data of the animals and the intrauterine development of the fetuses. The treatment at the dose levels of 80, 200 and 500 mg/kg bw/day caused no maternal or fetal effects (TOXI-COOP ZRT 590-410-0850).
Examinations
- Maternal examinations:
- CAGE SIDE OBSERVATIONS: mortality/morbidity
- Time schedule: twice daily
DETAILED CLINICAL OBSERVATIONS: Yes
- Time schedule: daily
BODY WEIGHT: Yes
- Time schedule for examinations: pre-mating and on day 0, 3, 5, 8, 11, 14, 17 and 20 of gestation (in addition on day 20 corrected for gravid uterus weight)
FOOD CONSUMPTION: Yes
-Time schedule for examinations: day 0 to 3, 3 to 5, 5 to 8, 8 to 11, 11 to 14, 14 to 17 and 17 to 20 by re-weighing the non-consumed diet
WATER CONSUMPTION: No
POST-MORTEM EXAMINATIONS: Yes gross necropsy
- Sacrifice on gestation day 20
- Organs examined: uterus with cervix and left ovary were weighed
OTHER: On gestation days 13 and/or 14 the sperm positive females were checked for the presence of vaginal bleeding which indicated the implantation of conceptuses - Ovaries and uterine content:
- The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantation sites: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes
- Number of live fetuses: Yes
- Fetal death: yes - Fetal examinations:
- - Fetal viability: Yes
- Fetal and placenta weight: Yes
- Fetal gender: Yes (measurement of anogenital distance)
- External examinations: Yes: all per litter
- Soft tissue examinations: Yes: half per litter (fixed in Sanomiva mixture before dissection)
- Skeletal examinations: Yes: half per litter (examined under dissection microscope after fixation in alcian-blue-acetic acid-ethanol mixture and in isopropanol, the skeletons were stained by KOH-Alizarin red-S method)
- Head examinations: Yes: half per litter (fetuses selected for visceral examination Wilson's technique) - Statistics:
- Bartlett's homogeneity of variance test or one-way ANOVA, chi-quare test
positive findings : distribution analysis and if normal concomitant ANOVA (Duncan's Multiple Range test) or Kolmogorov-Smirnov test.
non-normal distributions: Kruskal-Wallis One-Way analysis of variance - Indices:
- Pre-implantation loss
Post-implantation loss
Sex distribution
External abnormalities/litter
Visceral abnormalities/litter
Skeletal abnormalities/litter
calculated as %, group mean - Historical control data:
- included in the report
Results and discussion
Results: maternal animals
General toxicity (maternal animals)
- Clinical signs:
- no effects observed
- Description (incidence and severity):
- There was no mortality and no clinical signs were recorded during the in-life phase.
The number of sperm positive females was 88 in the study. The number of evaluated litters was 79 (20 both in the control and 100 mg/kg bw/day, 22 in the 300 and 17 in the 1000 mg/kg bw/day dose groups respectively).
(sum, %)
DESCRIPTION DOSES: control 100 300 1000 mg/kg bw/day
No. of
animals: 20 20 22 17
CLINICAL SYMPTOMS
- none N 20 20 22 17
% 100 100 100 100
NECROPSY FINDINGS
- no macroscopic alterations N 20 20 22 17
% 100 100 100 100 - Mortality:
- no mortality observed
- Description (incidence):
- There was no mortality and no clinical signs were recorded during the in-life phase.
DATA OF FEMALES MORTALITY(sum, %)
Dose groups mg/kg bw/day Control 100 300 1000
No of preg. fem. died 0 0 0 0 - Body weight and weight changes:
- no effects observed
- Description (incidence and severity):
- The body weight and corrected body weight (including corrected body weight gain) of the females was similar in all groups during the in-life phase.
From gestation day 5 to 8 the mean body weight gain of the dams in the 1000 mg/kg bw/day group was slightly but statistically significantly lower (p<0.01) versus control which was attributed to the treatment with the test item. The body weight gain of the dams in the 300 and 100 mg/kg bw/day group was at the current control level.
Detailed information about the body weight developments are given in the tables below or can be found in the attached tables in pdf format.
The slight but significant decrease in body weight gain during day 5 - 8 at 1000 mg/kg bw is considered incidental and not related to treatment
APPENDIX III SUMMARY OF BODY WEIGHT AND BODY WEIGHT GAIN OF DAMS
Body Weight DOSE GROUPS (mg/kg bw/day)
TIME Gestational days Control 100 300 1000
0 Mean 195,1 195,1 194 192,5
SD 12,66 10,04 10,85 9,04
n 20 20 22 17 NS
3 Mean 208,3 210,4 207,5 206,6
SD 14,44 10,17 11,85 10,32
n 20 20 22 17 NS
5 Mean 217,9 220 216,7 214,6
SD 15,58 13,36 14,48 10,87
n 20 20 22 17 NS
8 Mean 229,7 232,4 229,4 222,5
SD 17,06 14,93 16,75 12,3
n 20 20 22 17 NS
11 Mean 245,3 248,8 246,8 235,8
SD 19,21 18,31 18,78 12,71
n 20 20 22 17 NS
14 Mean 258,7 261,4 260,2 248,9
SD 20,65 19,01 19,61 13,16
n 20 20 22 17 NS
17 Mean 282,6 285,7 286 237,5
SD 23,24 21,47 21,98 14,09
n 20 20 22 17 NS
20 Mean 319,2 320,7 321,3 307,6
SD 28,44 26,72 23,28 12,72
n 20 20 22 17 NS
REMARKS :
NS = Not Significant
* = p < 0.05
** = p < 0.01
U = Mann-Whitney U - test Versus Control
DN = Duncan's multiple range test
Body weight gain DOSE GROUPS (mg/kg bw/day)
TIME Gestational days Control 100 300 1000
0-3 Mean 13,3 15,3 13,5 14,1
SD 3,16 5,03 3,11 3,93
n 20 20 20 17 NS
3-5 Mean 9,6 9,6 9,2 8,1
SD 3,17 3,72 4,18 2,44
n 20 20 22 17 NS
5-8 Mean 11,8 12,4 12,7 7,8
SD 2,76 3,87 3,71 2,72
n 20 20 22 17 **DN
8-11 Mean 15,6 16,5 17,4 12,6
SD 4,33 4,67 3,89 9,23
n 20 20 22 17 NS
11-14 Mean 13,4 12,6 13,4 24,5
SD 3,44 3,94 4,64 5,36
n 20 20 22 17 NS
14-17 Mean 23,9 24,3 25,8 24,5
SD 4,33 5,07 5,31 5,36
n 20 20 22 17 NS
17-20 Mean 36,7 35 35,3 34,2
SD 8,42 7,77 5,17 7,39
n 20 20 22 17 NS
0-20 Mean 124,2 125,6 127,3 115,1
SD 19,4 19,02 15,77 8,08
n 20 20 22 17 NS
REMARKS :
NS = Not Significant
* = p < 0.05
** = p < 0.01
U = Mann-Whitney U - test Versus Control
DN = Duncan's multiple range test
SUMMARY OF GRAVID UTERINE WEIGHT, CORRECTED BODY WEIGHT AND CORRECTED BODY WEIGHT GAIN OF DAMS
DOSE GROUPS (mg/kg bw/day) Control 100 300 1000
Gravid uterine weight (g) Mean 65,2 62,1 63,4 62,1
SD 10,67 8,87 9,79 10,22 NS
N 20 20 22 17
Corrected body weight (g) Mean 254 258,6 257,9 245,5
SD 19,59 22,32 20,36 17,46
N 20 20 22 17 NS
Corrected body weight gain (g) Mean 59 63,5 63,9 53
SD 10,9 15,15 12,83 13,01
N 20 20 22 17 NS
REMARKS : NS = Not Significant
* = p < 0.05 ** = p < 0.01
U = Mann-Whitney U - test Versus Control DN = Duncan's multiple range test - Food consumption and compound intake (if feeding study):
- effects observed, treatment-related
- Description (incidence and severity):
- The mean food consumption of the dams in the 1000 mg/kg bw/day group was slightly but statistically significantly lower (p<0.01) from gestational day 5 to 14 (-23%, -20% and -11% relative to controls between days 5 to 8, 8 to 11 and 11 to 14, respectively) which was considered to be due to the treatment with the test item.
APPENDIX V SUMMARY OF FOOD CONSUMPTION DATA OF DAMS
DOSE GROUPS (mg/kg bw/day)
TIME Gestational days Control 100 300 1000
0-3 Mean 18,7 18,5 17,9 17,8
SD 0,87 1,41 0,92 0,69
n 20 20 22* 17** U
3-5 Mean 20,8 21,3 20,4 20,2
SD 1,06 2,18 1,25 0,59
n 20 20* 22 17 U
5-8 Mean 21,2 21,2 20,6 16,3
SD 0,67 2,29 1,66 1,22
n 20 20 22 17** U
8-11 Mean 22,2 22,7 21,8 17,7
SD 1,12 2,89 1,37 1,12
n 20 20 22 17** U
11-14 Mean 23,2 23,4 23,5 20,7
SD 1,51 2,51 1,22 2,31
n 20 20 22 17** U
14-17 Mean 22,9 23,6 23,7 22
SD 1,57 2,81 1,43 2,17
n 20 20 22 17 NS
17-20 Mean 23 23,9 24,9 22,9
SD 1,62 2,82 1,5 2,43
n 20 20 22** 17 U
REMARKS : NS = Not Significant
* = p < 0.05 ** = p < 0.01
U = Mann-Whitney U - test Versus Control DN = Duncan's multiple range test - Food efficiency:
- not examined
- Water consumption and compound intake (if drinking water study):
- not examined
- Ophthalmological findings:
- not examined
- Haematological findings:
- not examined
- Clinical biochemistry findings:
- not examined
- Urinalysis findings:
- not examined
- Behaviour (functional findings):
- not examined
- Immunological findings:
- not examined
- Organ weight findings including organ / body weight ratios:
- not examined
- Gross pathological findings:
- no effects observed
- Neuropathological findings:
- not examined
- Histopathological findings: non-neoplastic:
- not examined
- Histopathological findings: neoplastic:
- not examined
- Other effects:
- not examined
Maternal developmental toxicity
- Number of abortions:
- no effects observed
- Description (incidence and severity):
- The whole number of pre-implantation loss was statistically significantly higher in the 1000 mg/kg bw/day group according to the Chi square test but not if the mean percent was evaluated. Moreover, the relative value of 12.2% was within the control background data range (8.2 - 21.4%). Hence, this slight increase was considered to be unrelated from the treatment. There were no significant differences in the mean number of corpora lutea, implantations, viable fetuses and their sex distribution. There was no significant increase of early- and late embryonic death and post-implantation loss in the test item treated groups. Moreover, early embryonic death and post-implantation loss was statistically significantly lower in the 1000 mg/kg bw/day group according to the Chi square analysis.
APPENDIX VI/A INTRAUTERINE MORTALITY, VIABLE FETUSES, SEX DISTRIBUTION
GROUPS (mg/kg bw/day): Control 100 300 1000
NUMBER OF DAMS: 20 20 22 17
Total Intrauterine Mortality % Mean±SD: 15.0 ±9.33 15.3 ±9.89 14.4 ±13.76 15.3 ±13.41 NS - Pre- and post-implantation loss:
- no effects observed
- Description (incidence and severity):
- At 1000 mg/kg bw post-implantation loss was decreased significantly (while pre-implantation loss was slightly, but not-significantly increased).
APPENDIX VI/A INTRAUTERINE MORTALITY, VIABLE FETUSES, SEX DISTRIBUTION
GROUPS (mg/kg bw/day): Control 100 300 1000
NUMBER OF DAMS: 20 20 22 17
Pre-implantation Loss % Mean ±SD: 6.3 ±6.42 6.3 ±8.02 7.7 ±10.32 12.2 ±15.20 NS
Post-implantation Loss % Mean ±SD: 9.3 ±7.53 9.5 ±8.74 7.5 ±9.55 3.1 ±4.85 * DN - Total litter losses by resorption:
- no effects observed
- Early or late resorptions:
- no effects observed
- Description (incidence and severity):
- APPENDIX VI/A INTRAUTERINE MORTALITY, VIABLE FETUSES, SEX DISTRIBUTION
GROUPS (mg/kg bw/day): Control 100 300 1000
NUMBER OF DAMS: 20 20 22 17
Early Embryonic Death % Mean ±SD: 8.1 ±7.05 6.6 ±7.65 6.3 ±7.07 2.7 ±4.75 NS
Late Embryonic Death % Mean ±SD: 1.1 ±3.72 2.5 ±4.24 1.2 ±5.81 0.0 ±0.00 NS - Dead fetuses:
- no effects observed
- Description (incidence and severity):
- no treatment related effects
APPENDIX VI/A INTRAUTERINE MORTALITY, VIABLE FETUSES, SEX DISTRIBUTION
GROUPS (mg/kg bw/day): Control 100 300 1000
NUMBER OF DAMS: 20 20 22 17
Dead Fetuses % Mean ±SD: 0.0 ±0.00 0.4 ±1.60 0.0 ±0.00 0.5 ±1.87 NS - Changes in pregnancy duration:
- not examined
- Description (incidence and severity):
- Effects on pregnancy duration: not examined
- Changes in number of pregnant:
- no effects observed
- Description (incidence and severity):
- The lower pregnancy rate at 1000 mg/kg bw is unrelated to treatment (all females were sperm positive at treatment start)
Females evaluated were 20, 20, 22 and 17 at 0, 100, 300 and 1000 mg/kg bw respectively.
APPENDIX I PREGNANCY DATA OF FEMALES, MORTALITY, MALFORMATIONS (sum, %)
Dose groups Control 100 mg/kg bw/day 300 mg/kg bw/day 1000 mg/kg bw/day
Number of sperm positive females 22 22 22 22
Number of females with no implantation but corpora lutea 0 1 0 1
Number of females with no implantation and no corpora lutea 2 1 0 4
Number and percent of pregnant females (females with implantation) 20 91% 20 91% 22 100% 17 77%
Number of dams with 3 or less implantations 0 0 0 0
Number of dams with total intrauterine death (total postimplantation loss) 0 0 0 0
Number of pregnant females died 0 0 0 0
Number of evaluated litters 20 20 22 17 - Other effects:
- no effects observed
Effect levels (maternal animals)
open allclose all
- Key result
- Dose descriptor:
- NOEL
- Effect level:
- 300 mg/kg bw/day
- Based on:
- test mat.
- Basis for effect level:
- food consumption and compound intake
- other: the effect on body weight (gain) due to the decreased food consumption is negligible
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day
- Based on:
- test mat.
- Basis for effect level:
- other: no effects
Results (fetuses)
- Fetal body weight changes:
- no effects observed
- Description (incidence and severity):
- There was no significant difference in the body weight of the fetuses in the experimental groups. The placental weight was slightly but statistically significantly lower in the 100, 300 and 1000 mg/kg bw/day and the relative placental weight in the 300 and 1000 mg/kg bw/day group if compared to the control. The placental weight values were slightly below the historical control level (654.9 - 747.5) in the 300 and 1000 mg/kg bw/day groups and in case of female fetuses in the 100 mg/kg bw/day group. Considering that the relative placental weight values for combined sexes were within the range of the background data (186.9 - 234.4) and showed no dose related tendency, the lower placental and relative placental weight values were likely not a consequence of the treatment.
APPENDIX VIII LITTER MEANS OF FETAL AND PLACENTAL WEIGHT
DOSE GROUPS (mg/kg bw/day) Control 100 300 1000
M+F M F M+F M F M+F M F M+F M F
Fetal weight (g) Mean 3,5 3,5 3,4 3,4 3,4 3,3 3,5 3,5 3,4 3,4 3,4 3,3
SD 0,25 0,25 0,25 0,2 0,27 0,19 2,1 0,22 0,22 0,28 0,29 0,3
N 20 20 20 20 20 20 22 22 22 17 17 17
N S NS NS
Placental weight (mg) Mean 716,5 729,6 699 656,5 660,7 652,6 648,3 651,1 643,3 634,4 645,1 623,3
SD 67,54 64,86 76,09 44,96 51,62 46,72 47,22 65,63 48,78 62,66 58,29 69,64
N 20 20 20 20** 20** 20* 22** 22** 22** 17** 17** 17**
DN DN DN
Relative placental weight (mg/g) Mean 207,7 207,3 207,3 197,9 194,6 200,5 188,8 184,6 191,4 190,2 188,5 193,1
SD 20,61 21,06 21,06 17,5 19,03 18,45 16,68 19,91 18,16 21,79 22,62 23,91
N 20 20 20 20 20 20 22** 22** 22* 17** 17** 17*
DN DN DN
Remarks: M = Male
F = Female
NS = Not Significant
* = p < 0.05
** = p < 0.01
U = Mann-Whitney U - test Versus Control
DN = Duncan's multiple range test
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): no effects observed
Field "Description (incidence and severity)" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.DescriptionIncidenceAndSeverityFetalPupBodyWeightChanges): There was no significant difference in the body weight of the fetuses in the experimental groups. The placental weight was slightly but statistically significantly lower in the 100, 300 and 1000 mg/kg bw/day and the relative placental weight in the 300 and 1000 mg/kg bw/day group if compared to the control. The placental weight values were slightly below the historical control level (654.9-747.5) in the 300 and 1000 mg/kg bw/day groups and in case of female fetuses in the 100 mg/kg bw/day group. Considering that the relative placental weight values for combined sexes were within the range of the background data (186.9-234.4) (Appendix XXIV/A) and showed no dose related tendency, the lower placental and relative placental weight values were likely not a consequence of the treatment. - Reduction in number of live offspring:
- no effects observed
- Changes in sex ratio:
- no effects observed
- Description (incidence and severity):
- There were no significant differences in the mean sex distribution.
APPENDIX VI/A SEX DISTRIBUTION
GROUPS (mg/kg bw/day): Control 100 300 1000
NUMBER OF DAMS: 20 20 22 17
Male fetuses % Mean ±SD: 56.5 12.49 46.6 ±15.91 47.1 ±15.87 53.1 ±17.23 NS
Female fetuses % Mean ±SD: 43.5 ±12.49 53.4 ±15.91 52.9 ±15.87 46.9 ±17.23 NS - Changes in litter size and weights:
- no effects observed
- Changes in postnatal survival:
- not examined
- External malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- The number of examined fetuses was 231, 222, 249 and 195 in the control, 100, 300 and 1000 mg/kg bw/day groups respectively.
Malformations
One fetus was found with an umbilical hernia and malrotated left hindlimb in the 100 mg/kg bw/day dose group. The flexion of the hindlimb was not confirmed at skeletal examination. Considering that this was a single fetus and in the low dose this was judged to have likely no relationship with the test item. Moreover, umbilical hernia occurs sporadically with low incidence unrelated to the treatment according to the experience with this species in this laboratory which is in line with historical control data of another strain of rats, i.e. CrL:CD(R) BR rats (Lang, 1993) (11). There were no external malformations recorded in the 300 and 1000 mg/kg bw/day groups.
Variations
The incidence of body weight retarded fetuses and litters (below 2.89 g for males and below 2.77 g for females) was statistically significantly (p<0.05) higher in the 1000 mg/kg bw/day group according to the Chi square test. There was no significance detected if the incidence of affected litters or the percentile litter means were evaluated and no clear dose relationship was observed. Hence, this was considered to be unrelated from the treatment. There was no statistical significance indicated in the other groups.
Placental abnormalities
There were no treatment related placental abnormalities found during the external examination. Two placentas were fused in the control group.
APPENDIX VII/A RESULTS OF EXTERNAL, VISCERAL AND SKELETAL EXAMINATIONS
(percentile litter means and SD)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
EXTERNAL EXAMINATION
Litters examined N 20 20 22 17
Fetuses examined N 231 222 249 195
Fetuses with abnormalities Mean 4,4 8 3,6 9,4
SD 10,2 13,18 7,83 14,55
Variation Mean 4,4 7,5 3,6 9,4
SD 10,2 13,29 7,83 14,55
Malformation Mean 0 0,5 0 0
SD 0 2,24 0 0
Retarded in body weight Mean 4,4 8 3,6 9,4
SD 10,2 11,78 7,83 14,55
Remarks: * = p < 0.05
** = p < 0.01
U = Mann-Whitney U - test Versus Control
DN = Duncan's multiple range test
APPENDIX VII/A RESULTS OF EXTERNAL, VISCERAL AND SKELETAL EXAMINATIONS
(sum, %)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
EXTERNAL EXAMINATION
Litters examined N 20 20 22 17
Fetuses examined N 231 222 249 195
Fetuses with abnormalities N 9 17 9 18*
% 4 8 4 9
Litters N 6 9 6 8
% 30 45 27 47
Variation N 9 16 9 18*
% 4 7 4 9
Litters N 6 8 6 8
% 30 40 27 47
Malformation N 0 1 0 0
% 0 0 0 0
Litters N 0 1 0 0
% 0 5 0 0
Retarded in body weight N 9 17 9 18*
% 4 8 4 9
Litters N 6 9 6 8
% 30 45 27 47
Remarks:
* = p < 0.05; CH2
** = p < 0.01; CH2
APPENDIX IX TYPES OF EXTERNAL ABNORMALITIES
(sum, %)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
Number of Dams N 20 20 22 17
Fetuses examined N 231 222 249 195
Fetuses with abnormalities N 9 17 9 18*
% 4 8 4 9
Litters N 6 9 6 8
% 30 45 27 47
Variation N 9 16 9 18*
% 4 7 4 9
Litters N 6 8 6 8
% 30 40 27 47
Malformation N 0 1 0 0
% 0 0 0 0
Litters N 0 1 0 0
% 0 5 0 0
FETAL VARIATIONS
Retarded in body weight N 9 17 9 18*
% 4 8 4 9
Litters N 6 9 6 8
% 30 45 27 47
Hydrops fetalis N 0 1 0 0
% 0 0 0 0
Litters N 0 1 0 0
% 0 5 0 0
FETAL MALFORMATIONS
Multiple malformed N 0 1 0 0
% 0 0 0 0
Litters N 0 1 0 0
% 0 5 0 0
Placental abnormalities
Fused N 1 0 0 0
% 0 0 0 0
Litters N 1 0 0 0
% 5 0 0 0
Remarks:
* = p < 0.05; CH2
** = p < 0.01; CH2 - Skeletal malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- The number of examined fetuses was 118, 113, 125 and 97 in the control, 100, 300 and 1000 mg/kg bw/day groups respectively.
Malformations:
There were two malformations found in the control group, one fetus had bent scapula (unilateral) and one with a dumb-bell shaped cartilage of a thoracic vertebral centrum. Three malformed fetuses were found in the 100 mg/kg bw/day group. One of these fetuses had a split sternum (this was the same fetus with umbilical hernia at external examination), bifurcate rib was recorded for a second and short femur, tibia and fibula for a third one. There were no malformations found in the 300 mg/kg bw/day dose group. Abnormal articulation of the fibula with calcaneus and talus was recorded for one fetus in the 1000 mg/kg bw/day group.
Split sternum may occur without any test item relationship with a low incidence according to the laboratory’s Historical Control Data (Appendix XXIV/B) of Toxi-Coop Zrt. This is in line with historical control data of another strain of rats, i.e. CrL:CD(R) BR rats (Lang, 1993) by which also bifurcate (branched) ribs and hypoplastic long bones of the hind limbs may occur in control animals (11). Considering the low incidence (one single fetus in the high dose group) of these different type of malformations the occurrence was likely not attributed to the treatment.
Variations:
Retardation of the skull, incomplete ossification or lack of ossification of the skull bones, hyoid, sternebra, vertebrae and metacarpal/metatarsal, bipartite supraoccipital, as well as bipartite sternal bodies, wavy ribs, dumb-bell shaped and bipartite and asymmetric thoracic or lumbar centra (with or without a slightly dumb-bell shaped cartilage), unossified SII arches, asymmetric ossification of metacarpal/metatarsal, asymmetric sacral/lumbar pelvic articulation and incompletely or not ossified pubic were evaluated as variations during the skeletal examination.
The incidence of 3 or less ossified sternebra increased significantly (p<0.05) in the 1000 mg/kg bw/day dose group however stayed within the historical control range (0 - 8.16%) and was not attributed to an effect of the test item. There were no significant differences in the different type of variations among the experimental groups.
APPENDIX VII/A RESULTS OF EXTERNAL, VISCERAL AND SKELETAL EXAMINATIONS
(percentile litter means and SD)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
SKELETAL EXAMINATION
Litters examined N 20 20 22 17
Fetuses examined N 118 113 125 97
Fetuses with abnormalities Mean 10 12,1 19,2 13,4
SD 16,58 12,29 22,09 16,79
Variation Mean 8,3 9,1 19,2 12,6
SD 15,76 11,17 2,09 17,1
Malformation Mean 1,7 2,9 0 0,8
SD 5,13 7,29 0 3,46
Remarks: * = p < 0.05
** = p < 0.01
U = Mann-Whitney U - test Versus Control
DN = Duncan's multiple range test
APPENDIX VII/A RESULTS OF EXTERNAL, VISCERAL AND SKELETAL EXAMINATIONS
(sum, %)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
SKELETAL EXAMINATION
Litters examined N 20 20 22 17
Fetuses examined N 118 113 125 97
Fetuses with abnormalities N 11 14 24* 13
% 9 12 19 13
Litters N 7 11 12 9
% 35 55 55 53
Variation N 9 11 24** 12
% 8 10 19 12
Litters N 6 9 12 8
% 30 45 55 47
Malformation N 2 3 0 1
% 2 3 0 1
Litters N 2 3 0 1
% 10 15 0 6
Remarks:
* = p < 0.05; CH2
** = p < 0.01; CH2
APPENDIX XI TYPES OF SKELETAL ABNORMALITIES
(sum, %)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
Number of Dams N 20 20 22 17
Fetuses examined N 118 113 125 97
Fetuses with abnormalities N 11 14 24* 13
% 9 12 19 13
Litters N 7 11 12 9
% 35 55 55 53
Variation N 9 11 24** 12
% 8 10 19 12
Litters N 6 9 12 8
% 30 45 47
Malformation N 2 3 0 1
% 2 3 0 1
Litters N 2 3 0 1
% 10 15 0 6
FETAL VARIATIONS
Skull
retarded N 0 1 0 0
% 0 1 0 0
Litters N 0 1 0 0
% 0 5 0 0
incomplete ossification (more than 3 bones) N 1 1 1 0
% 1 1 1 0
Litters N 1 1 1 0
% 5 5 5 0
incomplete ossification, marked (one bone or more) N 5 5 10 5
% 4 4 8 5
Litters N 4 3 6 3
% 20 15 27 18
supra occipital bipartite N 0 1 0 2
% 0 1 0 2
Litters N 0 1 0 2
% 0 5 0 12
hyoid not ossified N 0 0 0 1
% 0 0 0 1
Litters N 0 0 0 1
% 0 0 0 6
Sternebra
3 or less ossified N 1 3 3 6*
% 1 3 2 6
Litters N 1 2 3 5*
% 5 10 14 29
bipartite N 0 1 0 1
% 0 1 0 1
Litters N 0 1 0 1
% 0 5 0 6
Ribs
wavy N 5 0* 2 2
% 4 0 2 2
Litters N 3 0 2 2
% 15 0 9 12
Vertebrae
Thoracic centra
- dumb-bell shaped and/or asymmetric (more than 3) N 0 0 1 0
% 0 0 1 0
Litters N 0 0 1 0
% 0 0 5 0
bipartite or bipartite and asymmetric N 0 0 1 2
% 0 0 1 2
Litters N 0 0 1 2
% 0 0 5 12
dumb-bell shaped or bipartite N 1 1 1 1
and cartilage slightly dumb-bell shaped % 1 1 1 1
Litters N 1 1 1 1
% 5 5 5 6
Thoracic and lumbar centra and/or arches
not ossified (more than 3) N 0 0 0 1
% 0 0 0 1
Litters N 0 0 0 1
% 0 0 0 6
Lumbar/sacral arches
asymmetric pelvic articulation N 0 1 7** 1
% 0 1 6 1
Litters N 0 1 5* 1
% 0 5 23 6
SII arches N 0 0 1 0
not ossified % 0 0 1 0
Litters N 0 0 1 0
% 0 0 5 0
Pelvic girdle
pubic incomplete ossification or not ossified N 0 1 0 2
% 0 1 0 2
Litters N 0 1 0 2
% 0 5 0 12
Metacarpal or metatarsal
asymetric ossification N 1 2 0 0
% 1 2 0 0
Litters N 1 2 0 0
% 5 10 0 0
less than 3/3.5 ossified N 0 2 2 3
% 0 2 2 3
Litters N 0 2 2 3
% 0 10 9 18
FETAL MALFORMATIONS
Sternebra N 0 1 0 0
split % 0 1 0 0
Litters N 0 1 0 0
% 0 5 0 0
Ribs N 0 1 0 0
bifurcate % 0 1 0 0
Litters N 0 1 0 0
% 0 5 0 0
Thoraric centrum
dumb-bell shaped or bipartite (or bipartite asymmetric) and
cartilage dumb-bell shaped N 1 0 0 0
% 1 0 0 0
Litters N 1 0 0 0
% 5 0 0 0
Pectorale girdle N 1 0 0 0
scapula bent % 1 0 0 0
Litters N 1 0 0 0
% 5 0 0 0
Hindlimb N 0 1 0 0
femur, tibia, fibula short % 0 1 0 0
Litters N 0 1 0 0
% 0 5 0 0
fibula-calcanaeus, fibula-talus abnormal articulation N 0 0 0 1
% 0 0 0 1
Litters N 0 0 0 1
% 0 0 0 6
Remarks: * = p < 0.05; CH2
* = p < 0.05; CH2
** = p < 0.01; CH2 - Visceral malformations:
- effects observed, non-treatment-related
- Description (incidence and severity):
- The number of examined fetuses was 113, 109, 124 and 98 in the control, 100, 300 and 1000 mg/kg bw/day groups respectively.
Malformations:
One fetus was found with microphthalmy and one with situs inversus totalis in the control group. One fetus had multiple malformations (microphthalmy (bilateral); small kidney, small and malpositioned adrenal (unilateral) in the 100 mg/kg bw/day group. The right ovary was absent in this fetus and a technical error was not excluded. Considering that no other malformations were found in the test item treated groups at visceral examination this was judged to be likely unrelated from the treatment.
Variations:
Hydroureter and malpositioned testes (upper) was found with a very low incidence also in the control group and was judged to be unrelated from the treatment.
APPENDIX VII/A RESULTS OF EXTERNAL, VISCERAL AND SKELETAL EXAMINATIONS
(percentile litter means and SD)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
VISCERAL EXAMINATION
Litters examined N 20 20 22 17
Fetuses examined N 113 109 124 98
Fetuses with abnormalities Mean 3,5 1,8 0 0,8
SD 7,48 5,67 0 3,46
Variation Mean 2 0,8 0 0,8
SD 6,29 3,73 0 3,46
Malformation Mean 1,5 1 0 0
SD 4,78 4,47 0 0
Remarks: * = p < 0.05
** = p < 0.01
U = Mann-Whitney U - test Versus Control
DN = Duncan's multiple range test
APPENDIX VII/A RESULTS OF EXTERNAL, VISCERAL AND SKELETAL EXAMINATIONS
(sum, %)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
VISCERAL EXAMINATION
Litters examined N 20 20 22 17
Fetuses examined N 113 109 124 98
Fetuses with abnormalities N 4 2 0* 1
% 4 2 0 1
Litters N 4 2 0* 1
% 20 10 0 6
Variation N 2 1 0 1
% 2 1 0 1
Litters N 2 1 0 1
% 10 5 0 6
Malformation N 2 1 0 0
% 2 1 0 0
Litters N 2 1 0 0
% 10 5 0 0
Remarks:
* = p < 0.05; CH2
** = p < 0.01; CH2
APPENDIX X TYPES OF VISCERAL ABNORMALITIES
(sum, %)
DOSE GROUPS (mg/kg bw/day Control 100 300 1000
Number of Dams N 20 20 22 17
Fetuses examined N 113 109 124 98
Fetuses with abnormalities N 4 2 0* 1
% 4 2 0 1
Litters N 4 2 0* 1
% 20 10 0 6
Variation N 2 1 0 1
% 2 1 0 1
Litters N 2 1 0 1
% 10 5 0 6
Malformation N 2 1 0 0
% 2 1 0 0
Litters N 2 1 0 0
% 10 5 0 0
FETAL VARIATIONS
Ureters N 1 0 0 1
% 1 0 0 1
Litters N 1 0 0 1
% 5 0 0 6
Gonads N 1 1 0 0
% 1 1 0 0
Litters N 1 1 0 0
% 5 5 0 0
FETAL MALFORMATIONS
Eyes N 1 0 0 0
microphthalmy % 1 0 0 0
Litters N 1 0 0 0
% 5 0 0 0
General N 1 0 0 0
situs inversus totalis % 1 0 0 0
Litters N 1 0 0 0
% 5 0 0 0
Multiple malformed*** N 0 1 0 0
% 0 1 0 0
Litters N 0 1 0 0
% 0 5 0 0
Remarks:
* = p < 0.05; CH2 ***=microphthalmy (bilateral); kidney small, (right); adrenal small, half of left (right) and malpositioned, ovary absent (right) /ovary technical error is not excluded
** = p < 0.01; CH2 - Other effects:
- effects observed, non-treatment-related
- Description (incidence and severity):
- significantly increased growth retardation in fetuses at 1000 mg/kg bw (variation no dose response)
incomplete ossification in all dose groups (significantly increased at 300 mg/kg bw); no dose response
Effect levels (fetuses)
- Key result
- Dose descriptor:
- NOAEL
- Effect level:
- 1 000 mg/kg bw/day
- Based on:
- test mat.
- Sex:
- male/female
- Basis for effect level:
- other: no treatment related adverse effects observed at any dose level
Overall developmental toxicity
- Key result
- Developmental effects observed:
- no
- Treatment related:
- no
Any other information on results incl. tables
Maternal effects
GROUPS (mg/kg bw/day): |
Control |
100 |
300 |
1000 |
NUMBER OF DAMS: |
20 |
20 |
22 |
17 |
Corpora Lutea |
13.6 |
13.2 |
13.3 |
13.6 |
Preimplantation Loss % |
6.3 |
6.3 |
7.7 |
12.2 |
Implantation |
12.7 |
12.3 |
12.2 |
11.9 |
Early Embryonic Death % |
8.1 |
6.6 |
6.3 |
2.7 |
Late Embryonic Death % |
1.1 |
2.5 |
1.2 |
0.0 |
Dead Fetuses % |
0.0 |
0.4 |
0.0 |
0.5 |
Postimplantation Loss % |
9.3 |
9.5 |
7.5 |
3.1* |
Total Intrauterine Mortality % |
15.0 |
15.3 |
14.4 |
15.3 |
Viable fetuses |
11.6 |
11.1 |
11.3 |
11.5 |
Male fetuses % |
56.5 |
46.6 |
47.1 |
53.1 |
Female fetuses % |
43.5 |
53.4 |
52.9 |
46.9 |
Fetal effects
EXTERNAL EXAMINATION |
Control |
100 |
300 |
1000 |
Litters examined |
20 |
20 |
22 |
17 |
Fetuses examined |
231 |
222 |
249 |
195 |
Fetuses with abnormalities |
4.4 |
8.0 |
3.6 |
9.4 |
Variation |
4.4 |
7.5 |
3.6 |
9.4 |
Malformation |
0.0 |
0.5 |
0.0 |
0.0 |
Retarded in body weight |
4.4 |
8.0 |
3.6 |
9.4 |
VISCERAL EXAMINATION |
|
|
|
|
Litters examined |
20 |
20 |
22 |
17 |
Fetuses examined |
113 |
109 |
124 |
98 |
Fetuses with abnormalities |
3.5 |
1.8 |
0.0 |
0.8 |
Variation |
2.0 |
0.8 |
0.0 |
0.8 |
Malformation |
1.5 |
1.0 |
0.0 |
0.0 |
SKELETAL EXAMINATION |
|
|
|
|
Litters examined |
20 |
20 |
22 |
17 |
Fetuses examined |
118 |
113 |
125 |
97 |
Fetuses with abnormalities |
10.0 |
12.1 |
19.2 |
13.4 |
Variation |
8.3 |
9.1 |
19.2 |
12.6 |
Malformation |
1.7 |
2.9 |
0.0 |
0.8 |
Applicant's summary and conclusion
- Conclusions:
- Based on the results of the study no developmental toxicity was observed at 1000 mg/kg bw. Maternal effects (decreased food consumption during day 5 - 14) were observed at 300 mg/kg bw.
- Executive summary:
Twenty two sperm positive female rats were exposed orally to the substance from day 5 to 19 of gestation at 0, 100, 300 and 1000 mg/kg bw in a test according to OECD 414. Maternal effects were limited to a decreased food consumption during day 5 to 14 of the test period. No mortality, clinical signs and macroscopic changes were observed in the dams. Evaluations for developmental and fetal effects are based on 20, 20, 22 and 17 pregnancies. No effects on numbers of corpora lutea, implantations, resorptions, live fetuses, sex ratio and fetal weight were observed. No treatment related effects were found in external, visceral and skeletal examinations of the fetuses. The NOAEL for maternal toxicity is set at 1000 mg/kg bw (NOEL 300 mg/kg bw) and the NOAEL for developmental effects is 1000 mg/kg bw.
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