4-(2,6,6-trimethylcyclohex-2-ene-1-yl)-but-3-ene-2-one

Endpoint summary

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Effects on fertility

Description of key information

Reproductive toxicity study

Based on the data available from the various test chemicals, the no observed adverse effect level (NOAEL) was considered to be in range of 30 - 750 mg/kg body weight/day when male and female rats were treated with test chemical orally,Thus, comparing this value with the criteria of CLP regulation test chemical is not likely to classify as reproductive toxicant.

Link to relevant study records

Reference

Endpoint:

toxicity to reproduction

Remarks:

other: Chronic repeated dose toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from peer reviewed journal

Qualifier:

equivalent or similar to guideline

Guideline:

other: OECD 408

Principles of method if other than guideline:

Chronic repeated dose toxicity study of test material orally in rats.

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

other: FDRL

Sex:

male/female

Details on test animals or test system and environmental conditions:

- Source: No data available
- Age at study initiation: (P) x wks; (F1) x wks No data available
- Weight at study initiation: (P) Males: x-x g; Females: x-x g; (F1) Males: x-x g; Females: x-x g
P : Males: 59.5 ±1.5 g, Females: 58.0 ± 1.6 g
- Fasting period before study: No data available
- Housing: Animals were housed individually in wire mesh cages
- Diet (e.g. ad libitum): Purina Laboratory Chow, ad libitum
- Water (e.g. ad libitum): Fresh water, ad libitum
- Acclimation period: No data available

Route of administration:

oral: feed

Vehicle:

cotton seed oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: The test substance was diluted in cotton-seed oil in a concentration sufficient to provide the predetermined dosage in 2% of the diet. The oil solutions were incorporated into a nutritionally adequate basal ration (Purina Laboratory Chow).

DIET PREPARATION
- Rate of preparation of diet (frequency): biweekly
- Mixing appropriate amounts with (Type of food): nutritional adequate basal ration (Purina Laboratory Chow)
- Storage temperature of food: No data available

VEHICLE
- Justification for use and choice of vehicle (if other than water): Purina Laboratory Chow
- Concentration in vehicle: 0 or 10.6 mg/kg body weight/day (expected dose) (0 or 11.8 (males) and 11.1 (females) mg/Kg bw/day)
- Amount of vehicle (if gavage): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available

Details on mating procedure:

- Any other deviations from standard protocol: Reproductive organ weight were observed

Analytical verification of doses or concentrations:

not specified

Duration of treatment / exposure:

90 days

Frequency of treatment:

Daily

Details on study schedule:

No data available

Remarks:

Doses / Concentrations:
Males: 11.8 mg/Kg bw/ day Females: 11.1 mg/Kg bw/ day (0 or 10.6 mg/kg body weight/day (expected dose))
Basis:
actual ingested

No. of animals per sex per dose:

Total: 30 males and 30 females
0 mg/kg bw/day: 15 male, 15 female
10.6 mg/kg bw/day: 15 male, 15 female

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Single dosage levels for each substance were derived from the total estimated daily intake, calculated on a mg/kg body weight basis assuming 50 kg as the average body weight, and multiplying by 100.
- Rationale for animal assignment (if not random): No data available
- Rationale for selecting satellite groups: No data available
- Post-exposure recovery period in satellite groups: No data available
- Section schedule rationale (if not random): No data available

Positive control:

No data available

Parental animals: Observations and examinations:

Body weight, Food consumption, haematology and clinical chemistry were examined.

Oestrous cyclicity (parental animals):

No data available

Sperm parameters (parental animals):

No data available

Litter observations:

No data available

Postmortem examinations (parental animals):

Organ weight, Gross pathological and histopathology were examined.

Postmortem examinations (offspring):

No data available

Statistics:

Statistical analysis were performed by the average values for those individual groups of either males or females which deviated by more than two standard errors from those of the controls.

Reproductive indices:

No data available

Offspring viability indices:

No data available

Clinical signs:

not specified

Dermal irritation (if dermal study):

not specified

Mortality:

not specified

Body weight and weight changes:

no effects observed

Description (incidence and severity):

No effects were observed on body weight and body weight gain of treated male and female rats as compared to control.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

No effects were observed on food consumption of treated male and female rats as compared to control.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

no effects observed

Description (incidence and severity):

No effect was observed on clinical chemistry of treated rats as compared to control.

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

Slight degree foci of myeloid metaplasia were observed in one male and control rats and no histopathologial changes were observed in reproductive gonads of male and female rats as compared to control.

Histopathological findings: neoplastic:

not specified

Other effects:

no effects observed

Reproductive function: oestrous cycle:

not specified

Reproductive function: sperm measures:

not specified

Reproductive performance:

not specified

Body weight:
No effects were observed on body weight and body weight gain of treated male and female rats as compared to control.

Food consumption: No effects were observed on food consumption of treated male and female rats as compared to control.

Test substance intake: 11.8 mg/Kg bw/day for male and 11.1 mg/Kg bw/day for female estimated compound intake

Organ weights: No effect was observed on liver and kidney weights of treated and reproductive gonads of male and female rats as compared to control.

Gross pathology: No significant gross pathological changes were observed in treated rats except occasional pulmonary alterations associated with a respiratory infection.
No gross pathological changes were observed in reproductive gonads of male and female rats as compared to control.

Histopathology: Slight degree foci of myeloid metaplasia were observed in one male and control rats and no histopathologial changes were observed in reproductive gonads of male and female rats as compared to control.

other findings:
Haematology: In female rat, slightly increased Haematocrit level was observed as compared to control.
The observed effect were considered to be non significant.

Clinical chemistry: No effect was observed on clinical chemistry of treated rats as compared to control.

Dose descriptor:

NOAEL

Effect level:

11.8 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male

Basis for effect level:

other: No adverse effect on body weight, food consumption, food efficiency, haematology, clinical chemistry, reproductive organ weight, reproductive gross pathology and histopathology

Remarks on result:

other: No effects observed reproductive organ

Dose descriptor:

NOAEL

Effect level:

11.1 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: No adverse effect on body weight, food consumption, food efficiency, haematology, clinical chemistry, reproductive organ weight, reproductive gross pathology and histopathology

Remarks on result:

other: No effetcs observed on reproductive organ

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Clinical signs:

not specified

Dermal irritation (if dermal study):

not specified

Mortality / viability:

not specified

Body weight and weight changes:

not specified

Food consumption and compound intake (if feeding study):

not specified

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Sexual maturation:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

not specified

Histopathological findings:

not specified

Other effects:

not specified

Behaviour (functional findings):

not specified

Developmental immunotoxicity:

not specified

Dose descriptor:

other: not specified

Generation:

other: not specified

Based on:

not specified

Sex:

not specified

Basis for effect level:

other: not specified

Remarks on result:

not measured/tested

Critical effects observed:

not specified

System:

other: not specified

Organ:

not specified

Reproductive effects observed:

not specified

Treatment related:

not specified

Conclusions:

The no observed adverse effect level (NOAEL) was considered to be 11.8 mg/Kg bw/ day for male and 11.1 mg/Kg bw/ day for female when FDRL male and female rats were treated with test chemical orally by feed for 90 days.

Executive summary:

In a Chronic repeated dose toxicity study, FDRL male and female rats treated with test material orally by feed. The test substance was diluted in cotton-seed oil in a concentration sufficient to provide the predetermined dosage in 2% of the diet. The oil solutions were incorporated into a nutritionally adequate basal ration (Purina Laboratory Chow). No effects were observed on body weight and body weight gain, food consumption and efficiency of food utilization of male and female rats as compared to control. Estimated compound intake for male were 11.8 mg/Kg bw/day and for female were 11.1 mg/Kg bw/day. Slightly increased in haematocrit level was observed in female rat as compared to control. The observed effects were considered to be non significant. Similarly, no effect was observed on liver and kidney weight of treated rats as compared to control. In addition, No significant gross pathological and histopathological changes were observed on reproductive gonads in treated male and female rats as compared to control. Therefore, the no observed adverse effect level (NOAEL) was considered to be 11.8 mg/Kg bw/ day for males and 11.1 mg/Kg bw/ day for females when FDRL male and female rats were treated with test chemical orally by feed for 90 days.

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

750 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

Dats is Klimisch 2 and from peer-reviewed journal

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Reproductive toxicity study

Data available fromvarious test chemical was reviewed to determine the reproductive toxicity of test chemical.The studies are as mentioned below:

In a Chronic repeated dose toxicity study, FDRL male and female rats treated with test material orally by feed. The test substance was diluted in cotton-seed oil in a concentration sufficient to provide the predetermined dosage in 2% of the diet. The oil solutions were incorporated into a nutritionally adequate basal ration (Purina Laboratory Chow). No effects were observed on body weight and body weight gain, food consumption and efficiency of food utilization of male and female rats as compared to control. Estimated compound intake for male were 11.8 mg/Kg bw/day and for female were 11.1 mg/Kg bw/day. Slightly increased in haematocrit level was observed in female rat as compared to control. The observed effects were considered to be non significant. Similarly, no effect was observed on liver and kidney weight of treated rats as compared to control. In addition, No significant gross pathological and histopathological changes were observed on reproductive gonads in treated male and female rats as compared to control. Therefore, the no observed adverse effect level (NOAEL) was considered to be 11.8 mg/Kg bw/ day for males and 11.1 mg/Kg bw/ day for females when FDRL male and female rats were treated with test chemical orally by feed for 90 days. The study does not conclude higher dose value. Hence it is acceptable to take such low NOAEL .

 

 

In another study for reproductive and developmental toxicity, the toxicity of test material was evaluated in Crl:CD® (SD) IGS BR VAF®/Plus female rats during formative stages of pregnancy and its effect on developing embryo-fetus orally by gavage in the concentration of 0, 3, 10 and 30 mg/kg/day. No effects were observed on survival, body weight and body weight gain and food consumption of treated female rats as compared to control. Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats. None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups. Similarly, No significant effect were observed on corpora lutea, implantations, litter sizes, live fetuses, resorptions, percentage of dead or resorbed conceptuses and percentage of live male fetuses and no alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control. In addition, no effects were observed on number of live fetuses, fetus weight and on gross external fetal alterations in treated rats as compared to control. Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7^thcervical vertebra in four foetuses were observed in 30 mg/kg/day and in control. Folded retinas in seven foetuses and aberrant umbilical artery in two fetuses at 10 mg/kg/day and Slight dilation of the renal pelvis in one fetus at 3 mg/kg/day was observed. All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to treatment because (1) neither the fetal nor litter incidences were dosage dependent; (2) the incidences did not significantly differ from the control group values; and/or (3) the incidences were within the ranges observed historically at the Testing Facility. Therefore, the no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for P0 and F1 generation when Crl: CD® (SD) IGS BR VAF®/Plus female rats were treated with test chemical orally by gavage route of exposure for 11 days.

In a dose range finding study, reproductive develpmental toxicity was evaluated in Crl:CD(SD) IGS BR VAF/plus pregnant female rats by using test material in the concentration of  0, 1.25, 2.5, 5 and 10 mg/kg/day orally by gavage in corn oil. No mortality and clinical sign were observed in treated female rats as compared to control. Increase in Body weight gains were observed in treated rats during the entire dosage period, the gestation period following the initiation of dosing and the entire gestation period, but this was not considered to be treatment related. Similarly, No effects were observed on Number and distribution of corpora lutea, implantation sites and uterine contents. No thoracic abdominal and pelvic viscera abnormalites were observed in treated rats as compared to control. In addition, No effect on sex of fetuses and no Gross external alterations were observed in fetuses of treated rats as compared to control. Therefore, the no observed adverse effect level (NOAEL) was considered to be > 10 mg/kg/day for P0 and F1 generation when Crl:CD(SD) IGS BR VAF/plus pregnant female rats were treated with test material orally for 11 days.

 

In yet another subchronic oral toxicity study, Sprague-Dawley Crl:CD IGS BR male and female rats were treated with test material in the concentration of 0, 5, 30 and 500 mg/kg/day orally by gavage in corn oil. No effects were observed on survival, clinical sign and Functional/behavioral parameters were observed in treated rats as compared to control. Statistically significant decrease in body weight gains were observed during week 7 and statistically significantly increased in body weight gains during weeks 9 and 13 were observed in 500 mg/kg/day treated rats but, not significant as compared to control at the end of study and considered to be non-toxic. Similarly, no effect s was observed on food consumption, food efficiency, water consumption, Ophthalmoscopic examination and hematological parameters of treated rats as compared to control. Statistically significant increase in albumin, plasma creatinine, total protein and cholesterol and statistically significant decrease in AST and Chloride level were observed in male rats and statistically significant increase in plasma creatinine, total protein and cholesterol and statistically significant decrease in alkaline phosphatase, AST and Bilirubin level of treated female rats at 500 mg/kg/day as compared to control. All individual values were within the normal range for rats of the strain and age used, and the intergroup difference was considered not to be toxicologically significant. In addition, no reproductive orange gross pathological changes were observed in treated rats as compared to control. Some absolute and relative Adrenals, Heart, Kidneys, Liver and Spleen weigh changes were observed in male and female rats as compared to control. All individual values for observed changes were within the normal range for rats of the strain and age used and in the absence of a dose-related response the differences were considered to be of no toxicological importance. At 500 mg/kg/day, minimal to slight adipose infiltration in Bone and bone marrow , minimal to moderate Globular accumulations of eosinophilic material in Kidneys, minimal generalized and centrilobular Hepatocyte enlargement of Liver and minimal Follicular cell hypertrophy of Thyroid were observed in treated male rats and minimal centrilobular Hepatocyte enlargement of Liver in female rats as compared to control. Kidney changes were also observed in 30 mg/kg/day treated male rats. Liver and Kidneys changes were commonly seen following the administration of xenobiotics and is generally regarded as adaptive in nature in the absence of associated degenerative changes. No effects were observed on mammary glands, ovaries, prostate and testes of treated rats as compared to control. Therefore, no observed adverse effect level (NOAEL) was considered to be 500 mg/kg/day when Sprague-Dawley Crl:CD IGS BR male and female rats were treated with test material orally by gavage for 90 days.

Another reproductive and developmental toxicity of the test chemical was also performed on male and female wistar rats, the test chemical was administered into the rats by oral gavage in the concentrations of 0. 250, 500, 750 and 1000 mg/kg bw/day once on pregnancy day 11. The vehicle used to gavage the test chemical was corn oil. The source for the animals was Center for breeding of laboratory animals of the Oswaldo Cruz Foundation (Cecal-Fiocruz) and weighed around 200 -300 g at the initiation of the study. The rats were kept in the controlled temperature environment wherein the humidity was maintained around 70% and photoperiod was adjusted to 12 hours dark and 12 hours light. The rats were housed in standard plastic cages with stainless steel cover lids and wood shavings as bedding and kept under conditions of controlled temperature. The animals were mated in the ratio of 2:1 (2 females and 1 males) and were cohabited for 2 hours. The pregnancy was confirmed by the presence of sperm in the vaginal smear and was referred to as day '0' of pregnancy. On day 21 of gestation all dams were anesthetized and killed by CO2 inhalation. The gravid uterus was weighed with its contents and the number of implantation sites, living and dead fetuses, and resorptions was recorded. The ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter were substantially increased, while the ratio of live fetuses per implantations per litter was drastically decreased, in dams treated with the highest dose of the test chemical (1000 mg/kg bw/day). In maternal animals, no signs of toxicity was observed till 750 mg/kg bw/day. The animals gained normal weight until sacrifice. After sacrifice and gross necropsy, the gravid uterus weight did not differ significantly till 750 mg/kg bw/day. Although, at 1000 mg/kg bw/day dose group, the number of resorptions were observed to be increased in maternal animals. Also, the gravid uterine weight was observed to be decreased significantly as compared to the control group. Also, the ratio of implantation sites to resorptions was found to be lower as compared to the control group. No externally-visible anomaly was noted in fetuses from dams treated orally with test material on pregnancy day 11. Thus, NOAEL was considered to be 750 mg/kg bw/day and LOAEL was considered to be 1000 mg/kg bw/day. When maie and female rats were treated with test material orally once on pregnancy day 11.

Based on the data available from different test chemicals, the target chemical did not showed reproductive toxicity at dose concentration 750 mg/kg body weight/day, when male and female rats were treated with test material orally. Thus, comparing this value with the criteria of CLP regulation, the test chemical is not likely to classify as reproductive toxicant.

 

Effects on developmental toxicity

Description of key information

The no observed adverse effect level (NOAEL) was considered to be 750 mg/kg/day for F0 and F1 generation when rats were treated with test chemical by gavage route of exposure.

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

Experimental data from various test chemicals

Justification for type of information:

Data for the target chemical is summarized based on data from various test chemicals

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

other: Refer below principle

Principles of method if other than guideline:

WoE for the target CAS is summarized based on data from various test chemicals

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

other: 2. Crl:CD® (SD) IGS BR VAF®/Plus; 3. Wistar

Details on test animals or test system and environmental conditions:

2. TEST ANIMALS
- Source: Charles River Laboratories, Raleigh, NC
- Age at study initiation: 6-8 weeks old
- Weight at study initiation: Females: 213 to 241 g,
- Fasting period before study: No data available
- Housing: Animals were housed individually in cage.
- Diet (e.g. ad libitum): Certified Rodent DietR 5002 (PMI Nutrition International, St. Louis, MO), ad libitum
- Water (e.g. ad libitum): reverse osmosis deionized water (with chlorine added to the processed water as a bacteriostat), ad libitum
- Acclimation period: acclimatizated for short period.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17.7 ± 26.11°C
- Humidity (%): 30% to 70%
- Air changes (per hr):at least 10 changes per hour of 100% fresh air passed through 99.97% HEPA filters.
- Photoperiod (hrs dark / hrs light): 12:12-h light-dark lighting cycle was used

IN-LIFE DATES: From: To: No data available

3. TEST ANIMALS
- Source: Center for Breeding of Laboratory Animals, Oswaldo Cruz Foundation
- Age at study initiation: No data available
- Weight at study initiation: approx. 220 g (200-250 g)
- Fasting period before study: No data available
- Housing: Animals were housed in standard plastic cages with stainless steel cover lids and wood shavings as bedding in a controlled environment.
- Diet (e.g. ad libitum): Nuvital diet, ad libitum
- Water (e.g. ad libitum): Tap water, ad libitum
- Acclimatization period: No data available

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23±1°C
- Humidity (%):70%
- Air changes (per hr): No data available
- Photoperiod (hrs dark / hrs light): 12-hr dark/12-hr light

Route of administration:

oral: gavage

Type of inhalation exposure (if applicable):

not specified

Vehicle:

corn oil

Details on exposure:

2. PREPARATION OF DOSING SOLUTIONS:
Dosing formulations were prepared daily from bulk materials. Samples from each concentration of the dosing suspensions (first and last days of treatment) were analyzed for the test chemical content.3 mg/ml conc.level was found to be stable for 10 days when stored at ambient conditions and protected from light.

DIET PREPARATION
- Rate of preparation of diet (frequency):Not applicable
- Mixing appropriate amounts with (Type of food): Not applicable
- Storage temperature of food: Not applicable

VEHICLE
- Justification for use and choice of vehicle (if other than water): corn oil
- Concentration in vehicle: 0, 3, 10 and 30 mg/kg/day
- Amount of vehicle (if gavage): 10 ml/kg
- Lot/batch no. (if required): 103K0107 and 074K0025
- Purity: Not available

3. PREPARATION OF DOSING SOLUTIONS: The test chemical was mixed with corn oil at dose level of 0, 500, 1000 and 1500 mg/kg body wt
DIET PREPARATION-Not available
VEHICLE
- Justification for use and choice of vehicle (if other than water): Corn oil
- Concentration in vehicle: 0,250, 500, 750 or 1000 mg/kg/day
- Amount of vehicle (if gavage): No data available
- Lot/batch no. (if required): No data available
- Purity: No data available

Analytical verification of doses or concentrations:

not specified

Details on mating procedure:

2. - M/F ratio per cage:1:1
- Length of cohabitation:5 days
- After ... days of unsuccessful pairing replacement of first male by another male with proven fertility:Not available
- Further matings after two unsuccessful attempts:Not available
- Verification of same strain and source of both sexes:Not available
- Proof of pregnancy: The presence of spermatozoa and/or a copulatory plug in situ was designated as gestational day 0 (GD 0).
- Any other deviations from standard protocol:Not available

3. - M/F ratio per cage: 1:2
- Length of cohabitation: 2 h
- Proof of pregnancy: Confirmed by the presence of sperm in the vaginal smear.The day on which spermatozoa were found in the smear was designated as day 0 of pregnancy.
- After … days of unsuccessful pairing replacement of first male by another male with proven fertility:Not available
- Further matings after two unsuccessful attempts: [no / yes (explain)]: Not available
- After successful mating each pregnant female was caged (how): singly
- Any other deviations from standard protocol: Not available

Duration of treatment / exposure:

2. 11 days (gestational days 7 to 17)
3. Once (on Day 11 of pregnancy)

Frequency of treatment:

2. Daily
3. Once

Duration of test:

2. gestational day 21
3. 21 days

Remarks:

Doses / Concentrations: / 2
0, 3, 10, or 30 mg/kg/day
Basis:
no data

Remarks:

Doses / Concentrations: / 3
0, 250, 500, 750 or 1000 mg/kg bw
Basis:
actual ingested

No. of animals per sex per dose:

2. Total:100
Control(0)-25 female pregnant rats
3 mg/kg/day-25 female pregnant rats
10 mg/kg/day-25 female pregnant rats
30 mg/kg/day-25 female pregnant rats

3. Total: 65
0 mg/kg bw : 22 pregnant females
250 mg/kg/day: 12 pregnant females
500 mg/kg/day: 9 pregnant females
750 mg/kg/day: 12 pregnant females
1000 mg/kg/day: 10 pregnant females

Control animals:

yes, concurrent vehicle

Details on study design:

2. - Dose selection rationale: Dosages were selected on the basis of range-finding study. dosages of 0, 1.25, 2.5, 5, or 10 mg/kg/day were administered daily to 8 rats/group on GDs 7 to 17. No adverse test chemical related clinical signs and food consumption observed. During the post dosage period, the body weight gains remained increased. All rats were pregnant and all caesarean sectioning and litter observations were comparable among the five groups. No fetal gross external alterations were observed. Based on these data, a dosage greater than 10 mg/kg/day AIM was recommended for the definitive developmental toxicity study in rats.
- Rationale for animal assignment (if not random): The healthy, mated female rats, weighing 213 to 241 g, were assigned to four dosage groups, 25 rats/group, using a computer-generated (weight-ordered) randomization procedure based on body weights recorded on initiation of study.
- Rationale for selecting satellite groups:No data avaiable
- Post-exposure recovery period in satellite groups: No data avaiable
- Section schedule rationale (if not random):No data avaiable

3. Mating performed (8:00-10:00 h) for 2h.The rats were given single dose of beta-Ionone only on day 11 of pregnancy and on day 21 of gestation dams were anesthesized and killed by CO2 inhalation.

Maternal examinations:

2. CAGE SIDE OBSERVATIONS: yes, survival observed. Animals were observed twice daily for viability and examined for abnormal clinical signs
DETAILED CLINICAL OBSERVATIONS: Yes
BODY WEIGHT: Yes, Body weights were recorded prior to the start of the study and daily during the dosage and postdosage periods.
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes, Feed consumption was recorded on GDs 0, 7, 10, 12, 15, 18, and 21
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
POST-MORTEM EXAMINATIONS: Yes
POST-MORTEM EXAMINATIONS: Yes / No / No data
- Sacrifice on gestation day # On gestational day 21.
- Organs examined: Yes, Organ weight and gross pathology were examined.
OTHER: abortions and premature deliveries-

3. Body weight gain: Yes
The rats were weighed on days 0, 11 and 21 of pregnancy.

Ovaries and uterine content:

2. The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: Yes
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

3. The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Gravid uterus weight: Yes
- Number of corpora lutea: No
- Number of implantations: Yes
- Number of early resorptions: Yes
- Number of late resorptions: Yes

Fetal examinations:

2. - External examinations: Yes
- Soft tissue examinations: Yes
- Skeletal examinations: Yes
- Head examinations: No data

3. -Fetal body weight: Yes
- External examinations (incl. anomalies): Yes
- Head examinations: Yes
The living fetuses were weighed, numbered with a marker pen, examined for externally visible anomalies under a stereomicroscope and fixed in a 5% formalin solution.
The externally visible skull defects were scored for severity.

Statistics:

2. Continuous data were analyzed by using Bartlett’s test of homogeneity of variances and the analysis of variance. Dunnett’s test was used to identify statistical significance of individual groups. If the analysis of variance was not appropriate, the Kruskal-Wallis test or Dunn’s method of multiple comparisons was used to identify the statistical significance of the individual groups. If there were greater than 75% ties, Fisher’s exact test was used to analyze the data.

3. The data were analysed by the one-way analysis of variance (ANOVA) followed by the Scheffe’s test for multiple comparisons or, alternatively, by the Kruskal-Wallis test followed by the Mann-Whitney U-test whenever the variable did not fit a normal distribution. Proportions were analyzed by the chi square -test or, alternatively, by the Fisher’s exact test.

Indices:

2. Dead or resorbed conceptuses indice and live male fetuses indice were examined.
3. Not available

Historical control data:

No data avaialble

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

2. Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats. None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups.

3. No overt signs of maternal toxicity were observed as compared to control.

Dermal irritation (if dermal study):

not specified

Mortality:

no mortality observed

Description (incidence):

No effect was observed on survival of treated female rats as compared to control.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

2. No effects were observed on body weight and body weight gain of treated female rat during dosing and post dosage period (GDs 18 to 21).
3. No dose-related reduction of pregnancy weight gain were observed in treated rats.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

2. Absolute feed consumption was 101.0%, 102.1%, and 102.6% of the control value for 3, 10, and 30 mg/kg/day dose group. No effect was observed on food consumption of treated female rats as compared to control.

Food efficiency:

not specified

Water consumption and compound intake (if drinking water study):

not specified

Ophthalmological findings:

not specified

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Urinalysis findings:

not specified

Behaviour (functional findings):

not specified

Immunological findings:

not specified

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

2. No data
3. Decreased gravid uterus weight were observed in dams exposed to 1000 mg/kg/day.

Gross pathological findings:

no effects observed

Description (incidence and severity):

2. No alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control.

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

not specified

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Number of abortions:

not specified

Pre- and post-implantation loss:

not specified

Total litter losses by resorption:

no effects observed

Description (incidence and severity):

No significant effect were observed litter sizes, live fetuses, resorptions

Early or late resorptions:

no effects observed

Description (incidence and severity):

2. No significant effect were observed
3. When treated with 1000 mg/kg/day of test material the ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter were substantially increased

Dead fetuses:

no effects observed

Description (incidence and severity):

No significant effect were observed percentage of live male fetuses of treated female rats as compared to control.

Changes in pregnancy duration:

no effects observed

Changes in number of pregnant:

not specified

Other effects:

not specified

Details on maternal toxic effects:

2. Maternal toxic effects:no effects

Details on maternal toxic effects:
Mortality: No effect was observed on survival of treated female rats as compared to control.

Clinical sing: Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats.

None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups.

Body weight and body weight gain: No effects were observed on body weight and body weight gain of treated female rat during dosing and post dosage period (GDs 18 to 21).

Food consumption: Absolute feed consumption was 101.0%, 102.1%, and 102.6% of the control value for 3, 10, and 30 mg/kg/day dose group.

No effect was observed on food consumption of treated female rats as compared to control.

Reproductive performence: No significant effect were observed on corpora lutea, implantations, litter sizes, live fetuses, resorptions, percentage of dead or resorbed conceptuses and percentage of live male fetuses of treated female rats as compared to control.

Gross pathology: No alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control.

3. Maternal toxic effects:no effects. Remark: No dose related effects observed

Details on maternal toxic effects:
Clinical sign: No overt signs of maternal toxicity were observed as compared to control.
Body weight gain: No dose-related reduction of pregnancy weight gain were observed in treated rats.
Reproductive performance-When treated with 1000 mg/kg/day of BI the ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter were substantially increased, while the ratio of live fetuses per implantations per litter was drastically decreased.
Organ weights-Decreased gravid uterus weight were observed in dams exposed to 1000 mg/kg/day.

Dose descriptor:

NOAEL

Remarks:

2

Effect level:

>= 30 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

body weight and weight gain

clinical signs

food efficiency

gross pathology

maternal abnormalities

mortality

Remarks on result:

other: No toxic effects were observed

Dose descriptor:

NOAEL

Remarks:

3

Effect level:

750 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

other: maternal toxicity: No effects on Maternal weight gain, reproductive performance and organ weight

Abnormalities:

not specified

Localisation:

not specified

Fetal body weight changes:

no effects observed

Description (incidence and severity):

2. No effect was observed on fetus weight in treated groups as compared to control.

Reduction in number of live offspring:

no effects observed

Description (incidence and severity):

2. No effect was observed on number of live fetuses in treated groups as compared to control.
3. When treated with 1000 mg/kg/day of BI the ratio of live fetuses per implantations per litter was drastically decreased.

Changes in sex ratio:

not specified

Changes in litter size and weights:

not specified

Changes in postnatal survival:

not specified

External malformations:

not specified

Description (incidence and severity):

2. No data
3. No externally-visible anomaly and skull defects were observed in fetuses of treated rats as compared to control.

Skeletal malformations:

effects observed, treatment-related

Description (incidence and severity):

Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7th cervical vertebra in four foetuses of 30 mg/kg/day and in control.

Visceral malformations:

not specified

Other effects:

not specified

Details on embryotoxic / teratogenic effects:

2. Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Mortality: No effect was observed on number of live fetuses in treated groups as compared to control.

Fetus weight: No effect was observed on fetus weight in treated groups as compared to control.

Gross pathology: No gross external fetal alterations were observed in treated rats as compared to control.

Soft tissue malformations:
Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7th cervical vertebra in four foetuses of 30 mg/kg/day and in control.

Folded retinas in seven foetuses and aberrant umbilical artery in two fetuses at 10 mg/kg/day

Slight dilation of the renal pelvis in one fetus at 3 mg/kg/day was observed.

All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to test material
because (1) neither the fetal nor litter incidences were dosage dependent; (2) the incidences did not significantly differ from the control group values; and/or (3) the incidences were within the ranges observed historically at the Testing Facility.

3. Embryotoxic / teratogenic effects:no effects
Details on embryotoxic / teratogenic effects:
Gross pathology:
No externally-visible anomaly and skull defects were observed in fetuses of treated rats as compared to control

Dose descriptor:

NOAEL

Remarks:

2

Effect level:

>= 30 mg/kg bw/day

Based on:

test mat.

Sex:

male/female

Basis for effect level:

reduction in number of live offspring

fetal/pup body weight changes

changes in litter size and weights

skeletal malformations

Remarks on result:

other: All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to test material

Dose descriptor:

NOAEL

Remarks:

3

Effect level:

1 000 mg/kg bw/day

Based on:

test mat.

Sex:

not specified

Basis for effect level:

other: embryotoxicity: No effect on Fetal body weight, gross pathology and histopathology

Key result

Abnormalities:

no effects observed

Localisation:

other: not specified

Developmental effects observed:

not specified

Treatment related:

not specified

Conclusions:

The no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when rats were treated with test chemical by gavage route of exposure.

Executive summary:

Data available for various test chemicals was reviewed to determine the developmental toxic nature of the test chemical. The studies are as mentioned below:

Developmental study was initiated to study the toxicity of test material . In this study, Crl:CD® (SD) IGS BR VAF®/Plus male and female rats were mated for 5 days and the presence of spermatozoa and/or a copulatory plug in situ was designated as gestational day 0 (GD 0). The test material in a corn oil vehicle was administered orally via gavage to four groups of presumed pregnant rats on GDs 7 to 17 at dosages of 0, 3, 10, or 30 mg/kg/day. No effects were observed on survival, body weight and body weight gain and food consumption of treated female rats as compared to control. Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats. None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups. Similarly, No significant effect were observed on corpora lutea, implantations, litter sizes, live fetuses, resorptions, percentage of dead or resorbed conceptuses and percentage of live male fetuses and no alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control. In addition, no effects were observed on number of live fetuses, fetus weight and on gross external fetal alterations in treated rats as compared to control. Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7^thcervical vertebra in four foetuses were observed in 30 mg/kg/day and in control. Folded retinas in seven foetuses and aberrant umbilical artery in two fetuses at 10 mg/kg/day and Slight dilation of the renal pelvis in one fetus at 3 mg/kg/day was observed. All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to treatment because (1) neither the fetal nor litter incidences were dosage dependent; (2) the incidences did not significantly differ from the control group values; and/or (3) the incidences were within the ranges observed historically at the Testing Facility. Therefore, the no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when Crl: CD® (SD) IGS BR VAF®/Plus female rats were treated with test material orally by gavage for 11 days.

In another teratogenicity study, Wistar female rats were treated with test material in the concentration of 250,500, 750 and 1000 mg/kg body weight orally by gavage in corn oil at day 11 of pregnancy.No overt signs of maternal toxicity and effect on body weight gain were observed in treated rats as compared to control. Substantially increased in ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter, while the ratio of live fetuses per implantations per litter was drastically decreased at 1000 mg/kg bw in treated rats as compared to control. Similarly, Decreased in gravid uterus weight were observed in dams treated with 1000 mg/kg/day. In addition, no significant effect on externally visible anomaly and skull defects were observed in fetuses of treated rats as compared to control. Therefore, the no observed adverse effect level (NOAEL) was considered to be 750 mg/kg bw for P generation and 1000 mg/kg bw for F1 generation when Wistar female rats were treated with test material once orally by gavage in corn oil at day 11 of pregnancy.

Based on the details of the study, the no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when rats were treated with test chemical by gavage route of exposure.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

750 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Data is from Klimisch 2

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

Developmental toxicity:

Data available for various test chemicals was reviewed to determine the developmental toxic nature of the test chemical. The studies are as mentioned below:

Developmental study was initiated to study the toxicity of test material . In this study, Crl:CD® (SD) IGS BR VAF®/Plus male and female rats were mated for 5 days and the presence of spermatozoa and/or a copulatory plug in situ was designated as gestational day 0 (GD 0). The test material in a corn oil vehicle was administered orally via gavage to four groups of presumed pregnant rats on GDs 7 to 17 at dosages of 0, 3, 10, or 30 mg/kg/day. No effects were observed on survival, body weight and body weight gain and food consumption of treated female rats as compared to control. Chromorhinorrhea, excess salivation, sparse hair coat, localized alopecia, urine-stained abdominal fur, soft or liquid feces, or decreased activities were observed in a few animals from each group of rats. None of these clinical signs were attributed to treatment because the number of affected rats was not dosage related and did not significantly differ between control and treated groups. Similarly, No significant effect were observed on corpora lutea, implantations, litter sizes, live fetuses, resorptions, percentage of dead or resorbed conceptuses and percentage of live male fetuses and no alterations were observed on litters with fetuses and % fetuses/litter in treated female rats as compared to control. In addition, no effects were observed on number of live fetuses, fetus weight and on gross external fetal alterations in treated rats as compared to control. Limited to infrequent incidences of skeletal variations as delayed ossification at various bone ossification centers (non–dose dependent) and the presence of cervical ribs at the 7^thcervical vertebra in four foetuses were observed in 30 mg/kg/day and in control. Folded retinas in seven foetuses and aberrant umbilical artery in two fetuses at 10 mg/kg/day and Slight dilation of the renal pelvis in one fetus at 3 mg/kg/day was observed. All fetal gross external, soft tissue, or skeletal alterations (malformations or variations) were considered unrelated to treatment because (1) neither the fetal nor litter incidences were dosage dependent; (2) the incidences did not significantly differ from the control group values; and/or (3) the incidences were within the ranges observed historically at the Testing Facility. Therefore, the no observed adverse effect level (NOAEL) was considered to be 30 mg/kg/day for F0 and F1 generation when Crl: CD® (SD) IGS BR VAF®/Plus female rats were treated with test material orally by gavage for 11 days.

In another teratogenicity study, Wistar female rats were treated with test material in the concentration of 250, 500, 750 and 1000 mg/kg body weight orally by gavage in corn oil at day 11 of pregnancy.No overt signs of maternal toxicity and effect on body weight gain were observed in treated rats as compared to control. Substantially increased in ratio of resorptions per implantations, and the percentage of resorptions per implantations per litter, while the ratio of live fetuses per implantations per litter was drastically decreased at 1000 mg/kg bw in treated rats as compared to control. Similarly, Decreased in gravid uterus weight were observed in dams treated with 1000 mg/kg/day. In addition, no significant effect on externally visible anomaly and skull defects were observed in fetuses of treated rats as compared to control. Therefore, the no observed adverse effect level (NOAEL) was considered to be 750 mg/kg bw for P generation and 1000 mg/kg bw for F1 generation when Wistar female rats were treated with test material once orally by gavage in corn oil at day 11 of pregnancy.

Based on the details of the study, the no observed adverse effect level (NOAEL) was considered to be 750 mg/kg/day for F0 and F1 generation when rats were treated with test chemical by gavage route of exposure. Considering this, the target chemical did not showe developmental toxicity at dose concentration 750 mg/kg body weight/day, when male and female rats were treated with test material orally. Thus, comparing this value with the criteria of CLP regulation, the test chemical is not likely to classify as reproductive toxicant.

Justification for classification or non-classification

Based on the data available for target chemical α-ionone(CAS no 127-41-3) and its closely related test chemicals, the target chemical is likely to be non hazardous for reproductive and developmental toxicity.