2-tert-butylhydroquinone

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

Reproductive Toxicity Study:

The reproductive toxicity study summaries for the test chemical is as follows:

The study was performed to evaluate the cumulative toxic effects of test chemical on theF344/N rats.The test chemical was fed to female F344/N rats (F0 generation) at a doses of  0, 2500, 5000, 10000, 20000, or 40000 ppm (Approx 250, 500, 1000, 2000 or 4000 mg/kg bw) in diet for approximately 10 weeks (from 2 weeks prior to cohabitation until weaning of F1 pups). Clinical findings, body weights, and feed consumption were recorded weekly for F0 females during the first 2 weeks and during lactation.No effect was observed on gestation length, the average number of pups born per litter, or the number of dams with stillborn pups for dams up to 1000 mg/kg. The dams exposed to 2000 or 4000 mg/kg did not litter. In the500 and 1000 mg/kg group, more number of pup deaths and reduction in mean body weights of pups at weaning was observed. The average number of surviving pups per litter in the 1000 mg/kg group was lower as compared to the controls.Male and female F344/N rats for the 13-week base study were offspring (F1 generation) of the breeders from the perinatal phase of the study. Rats were approximately 34 days old: on the first day of the study.. Groups of 10 male and 10 female F1 rats were fed diets containing 0, 250, 500 and 1000 mg/kg of test chemical for 13 weeks after weaning. (No F1 offspring resulted from F0 females fed diets containing 2000 and 4000mg/kg doses so these exposure levels were not used in the 13-week rat study). Clinical findings were recorded and the animals were weighed initially, weekly, and at the end of the study; feed consumption was recorded as an average of grams per animal per day. Blood samples was collected at week 13 from core study (F1) for clinical pathology analyses .At the end of the study, samples from 0, 250, 500 and 1000mg/kg group F1 rats were collected for sperm motility and vaginal cytology evaluations. A necropsy was performed on all core study F1 animals. The results (F0 generation) of the study revealed no effect on gestation length, the average number of pups born per litter, the number of dams with stillborn pups(F0 generation) and dams exposed to 2000 or 4000 mg/kg did not litter. The results of the F1 generation rats revealed, hair discoloration in all except 250mg/kg treated rats. More number of pup deaths in 500 and 1000 mg/kg group than that of the control animals. Serum bile acid levels (in 500 and 1000mg./kg male and females and Serum alanine aminotransferase activity levels (in 250, 500 and 1000 mg/kg group males and female) were increased as comapred to control but not considered to be biologically significant. Increased incidence of hyperplasia of nasal respiratory epithelium (500 and 1000mg/kg) and increased incidences of splenic pigmentation (500 mg/kg and 1000 mg/kg)Thus, based on all the conclusions, the NOAEL for test chemical inF344/N rats was determined to be approximately 250 mg/kg bw.

Link to relevant study records

Reference

Endpoint:

one-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

experimental study

Adequacy of study:

key study

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 a NTP report.

Qualifier:

according to guideline

Guideline:

other:

Principles of method if other than guideline:

The 13-week study was performed to evaluate the cumulative toxic effects of test chemical with exposure to the chemical in Male and female F344/N rats and also to determine the dose levels to be used in the long-term study.

GLP compliance:

no

Limit test:

no

Specific details on test material used for the study:

- Molecular weight (if other than submission substance): 166.22 g/mol
- Substance type:Organic
- Physical state: solid
- Impurities (identity and concentrations):99%

Species:

rat

Strain:

other: F344/N

Details on species / strain selection:

No Data Available

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
Males acquired for the reproductive toxicity phase were used for breeding purposes only and were not considered part of the study.
- Source: Taconic Farms (Germantown, NY).
- Age at study initiation: F: 57 day old
- Housing: Polycarbonate cages with bedding
- Diet (e.g. ad libitum): NIH-07 open formula mash diet, ad libitum
- Water (e.g. ad libitum): Tap water via automatic watering system; available ad libitum
- Acclimation period: 19 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18.7°C to 24.2°C
- Humidity (%): 35.8 %-79.3 %
- Air changes (per hr): minimum of 10 changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours/day

Route of administration:

oral: feed

Vehicle:

other: NIH-07 open formula mash diet

Details on exposure:

Fo - Approximately 10 weeks
F1 - 87-89 days (clinical pathology study F1 rats)
93-94 days (core study F1 rats)

Details on mating procedure:

- M/F ratio per cage: 1:2
- Proof of pregnancy: Vaginal smears were taken daily from breeder females to determine the presence of sperm.
- After successful mating each pregnant female was caged (how): Females were housed individually when pregnancy was confirmed

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

The dose formulations for the 13-week study were prepared weekly. Homogeneity and stability analyses of the dose formulations were conducted by the analytical chemistry laboratory using high-performance liquid chromatography. Homogeneity was confirmed, and the stability of the dose formulations was confirmed for at least 3 weeks when stored in sealed containers in the dark at 5 ° C. For the 13-week studies, dose formulations were analyzed at the beginning, in the middle, and at the end of the studies. Of the dose formulations used in the 13-week studies, 98% were within 10% of the target concentration with no value greater than 16% from the target concentration.

Duration of treatment / exposure:

F0 generation exposed approximately for 10 weeks

F1 generation exposed to test chemical for 87-89 days (clinical pathology study F1 rats)
and 93-94 days (core study F1 rats)

Frequency of treatment:

Daily

Details on study schedule:

Fo generation - 10 females
F1 generation
Core study - I0 males and 10 females
Clinical pathology study - 10 males and 10 females were used.
Since Males acquired for the reproductive toxicity phase were used for breeding purposes only males not considered as part of the study

Remarks:

Doses / Concentrations: 0, 250, 500, 1000, 2000 or 4000 mg/kg bw

Basis:
nominal in diet Approx 0, 2500, 5000, 10000, 20000, or 40000 ppm

No. of animals per sex per dose:

Control: 10 females
2500 ppm (250 mg/kg): 10 females
5000 ppm (500 mg/kg): 10 females
10000 ppm (1000 mg/kg): 10 females
20000 ppm (2000 mg/kg): 10 females
40000 ppm (4000 mg/kg): 10 females

Control animals:

yes, concurrent vehicle

Details on study design:

Groups of 10 female rats (F0 generation) were fed diets containing Approx 0, 250, 500, 1000, 2000 or 4000 mg/kg of test chemial for approximately 10weeks. During cohabitation two F0 females were housed with one breeder male.Clinical findings and body weights were also recorded weekly during lactation. Rats that did not litter by day 25 were killed and uteri were stained with ammonium sulfide and examined for implantation sites. After parturition, on day 4 postpartum, litters were randomly culled to a maximum of eight rat pups per litter; pup weights were recorded on days 4, 11, 18, and 28. Pups were weaned on day 28.

Male and female F344/N rats for the 13-week base study were offspring (F1 generation) of the breeders from the perinatal phase of the study. Rats were approximately 34 days old: on the first day of the study.. Groups of 10 male and 10 female F1 rats were fed diets containing 0, 250, 500 and 1000mg/kg of test chemical for 13 weeks after weaning. (No F1 offspring resulted from F 0 females fed diets containing 2000 and 4000mg/kg doses so these exposure levels were not used in the 13-week rat study). Clinical findings were recorded and the animals were weighed initially, weekly, and at the end of the study; feed consumption was recorded as an average of grams per animal per day.

Blood samples was collected at week 13 from core study (F1) for clinical pathology analyses .At the end of the study, samples from 0, 250, 500 and 1000mg/kg group F1 rats were collected for sperm motility and vaginal cytology evaluations. A necropsy was performed on all core study F1 animals.

Positive control:

No Data Available

Parental animals: Observations and examinations:

F0 females were observed twice daily.
Clinical findings, body weights, and feed consumption were recorded weekly during the first 2 weeks of the study (prior to cohabilation; clinical findings and body weights were also recorded weekly during lactation.


F1 generation were observed twice daily.
Clinical findings were recorded and the animals were weighed initially, weekly, and at the end of the study; feed consumption was recorded as an average of grams per animal per day.

Blood samples was at 13 week from core study (F1), sperm motility and vaginal cytology evaluations and necropsy was performed on all core study F1 animals.

Oestrous cyclicity (parental animals):

From F1 generation, for 7 consecutive days prior to scheduled terminal sacrifice, the vaginal vaults of the females were moistened with saline and samples were collected Relative numbers of leukocytes, nucleated epithelial cells and large squamous epithelial cells were determined and used to ascertain estrous cycle stage. (i.e.,diestrus, proestrus, estrus, and metestrus).

Sperm parameters (parental animals):

Male rats from F1 generation were evaluated for sperm count and motility.

Litter observations:

After parturition in F0 generation, pups were examined and the number and sex of pups and the litter weight were recorded. Pup weights were recorded on days 4, 11, 18, and 28.

Postmortem examinations (parental animals):

F0 generation rats that did not litter by day 25 were killed and uteri were stained with ammonium sulfide and examined for implantation sites.

Postmortem examinations (offspring):

A necropsy was performed on all core study F1 animals. The heart, right kidney, liver, lungs, right testes, and thymus were weighed. Tissues for microscopic examination were fixed and preserved in 10% neutral buffered formalin, processed and trimmed, embedded in paraffin, sectioned to a thickness of 5 to 6 um, and stained with hematoxylin and eosin. A complete histopathologic examination was performed on all control and 1000mg/kg rats. Additionally, the following tissues from selected exposure groups were examined: the nose of all exposed groups of male rats and 500mg/kg female rats; the spleen of 500 and 1000 mg/kg group male rats and all exposed groups of female rats; the mesenteric lymph node of 500mg/kg female rats; and the kidneys of 250 and 1000mg/kg female rats.

Reproductive indices:

No data

Offspring viability indices:

No data

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Hair discoloration of all the treated groups were observed.

Body weight and weight changes:

not examined

Food consumption and compound intake (if feeding study):

not examined

Organ weight findings including organ / body weight ratios:

not examined

Histopathological findings: non-neoplastic:

not examined

Other effects:

not examined

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

effects observed, treatment-related

Description (incidence and severity):

Dams exposed to 2000 or 4000 mg/kg did not litter.

No effect on gestation length, the average number of pups born per litter, or the number of dams with stillborn pups for dams up to 1000 mg/kg

Dose descriptor:

NOAEL

Effect level:

ca. 250 mg/kg bw/day (nominal)

Based on:

test mat.

Sex:

female

Basis for effect level:

other: No effect on gestation length, the average number of pups born per litter, the number of dams with stillborn pups, number of pup deaths or mean body weight of pups

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

Hair discoloration was observed in all exposed groups of rats, with the exception of 250mg/kg females.

Mortality / viability:

mortality observed, treatment-related

Description (incidence and severity):

More number of pup deaths in 500 and 1000 mg/kg group than that of the control animals.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Mean body weight reduced in 500 and 1000 mg/kg group compared to control animals.

Haematological findings:

no effects observed

Description (incidence and severity):

Serum bile acid levels were generally significantly increased in 500 and 1000mg./kg male and female rats at day 5, at week 3, and at the end of the study.

Serum alanine aminotransferase activity levels were increased at day 5 in females exposed to 1000mg/kg, at week 3 in males and females exposed to 250, 500, or 1000mg/kg, and at the end of the study in males receiving 250mg/kg. However, because the increases observed in these two parameters were marginal hence not considered as biologicaly significant change.

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

no effects observed

Description (incidence and severity):

Differences in absolute and/or relative organ weights of control and exposed groups occured but these changes were not considered to be terst chemical related.

Gross pathological findings:

not examined

Histopathological findings:

effects observed, treatment-related

Description (incidence and severity):

Increased incidences of hyperplasia of the nasal respiratory epithelium were observed in males exposed to 500mg/kg and males and females exposed to 1000mg/kg, andan increased incidence of nasal exudate was observed in males in the 1000mg/kg group.

Increased incidences of splenic pigmentation were observed in males and females exposed to 500 and 1000mg/kg and incidences of atrophy of the red
pulp were observed in these groups of females.

Other effects:

effects observed, treatment-related

Description (incidence and severity):

The mean spermatid count, spermatid heads per testis and spermatid heads per gram of testis were significantly decreased in F1 males exposed to 500mg/kg and estrous cycles of F1 females exposed to 250 and 500mg/kg were significantly longer than that of the
controls.

Key result

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

250 mg/kg diet

Based on:

test mat.

Sex:

male/female

Basis for effect level:

clinical biochemistry

organ weights and organ / body weight ratios

histopathology: non-neoplastic

Reproductive effects observed:

not specified

Conclusions:

The NOAEL for test chemical in parental female F344/N rats was determined to be approximately 250 mg/kg bw.

Executive summary:

The study was performed to evaluate the cumulative toxic effects of test chemical on the F344/N rats. The test chemical was fed to female F344/N rats (F0 generation) at a doses of  0, 2500, 5000, 10000, 20000, or 40000 ppm (Approx 250, 500, 1000, 2000 or 4000 mg/kg bw) in diet for approximately 10 weeks (from 2 weeks prior to cohabitation until weaning of F1 pups). Clinical findings, body weights, and feed consumption were recorded weekly for F0 females during the first 2 weeks and during lactation. No effect was observed on gestation length, the average number of pups born per litter, or the number of dams with stillborn pups for dams up to 1000 mg/kg. The dams exposed to 2000 or 4000 mg/kg did not litter. In the 500 and 1000 mg/kg group, more number of pup deaths and reduction in mean body weights of pups at weaning was observed. The average number of surviving pups per litter in the 1000 mg/kg group was lower as compared to the controls. Male and female F344/N rats for the 13-week base study were offspring (F1 generation) of the breeders from the perinatal phase of the study. Rats were approximately 34 days old: on the first day of the study.. Groups of 10 male and 10 female F1 rats were fed diets containing 0, 250, 500 and 1000 mg/kg of test chemical for 13 weeks after weaning. (No F1 offspring resulted from F0 females fed diets containing 2000 and 4000mg/kg doses so these exposure levels were not used in the 13-week rat study). Clinical findings were recorded and the animals were weighed initially, weekly, and at the end of the study; feed consumption was recorded as an average of grams per animal per day. Blood samples was collected at week 13 from core study (F1) for clinical pathology analyses .At the end of the study, samples from 0, 250, 500 and 1000mg/kg group F1 rats were collected for sperm motility and vaginal cytology evaluations. A necropsy was performed on all core study F1 animals. The results (F0 generation) of the study revealed no effect on gestation length, the average number of pups born per litter, the number of dams with stillborn pups(F0 generation) and dams exposed to 2000 or 4000 mg/kg did not litter. The results of the F1 generation rats revealed, hair discoloration in all except 250mg/kg treated rats. More number of pup deaths in 500 and 1000 mg/kg group than that of the control animals. Serum bile acid levels (in 500 and 1000mg./kg male and females and Serum alanine aminotransferase activity levels (in 250, 500 and 1000 mg/kg group males and female) were increased as comapred to control but not considered to be biologically significant. Increased incidence of hyperplasia of nasal respiratory epithelium (500 and 1000mg/kg) and increased incidences of splenic pigmentation (500 mg/kg and 1000 mg/kg) Thus, based on all the conclusions, the NOAEL for test chemical in F344/N rats was determined to be approximately 250 mg/kg bw.

 

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

250 mg/kg bw/day

Study duration:

chronic

Species:

rat

Quality of whole database:

K2 level data refered from relaible source of 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:

The reproductive toxicity study summaries for the test chemical is as follows:

Reproductive Toxicity Study 1:

The study was performed to evaluate the cumulative toxic effects of test chemical on theF344/Nrats.The test chemical was fed to female F344/N rats (F0 generation) at a doses of  0, 2500, 5000, 10000, 20000, or 40000 ppm (Approx 250, 500, 1000, 2000 or 4000 mg/kg bw) in diet for approximately 10 weeks (from 2 weeks prior to cohabitation until weaning of F1 pups). Clinical findings, body weights, and feed consumption were recorded weekly for F0 females during the first 2 weeks and during lactation.No effect was observed on gestation length, the average number of pups born per litter, or the number of dams with stillborn pups for dams up to 1000 mg/kg. The dams exposed to 2000 or 4000 mg/kg did not litter. In the500 and 1000 mg/kg group, more number of pup deaths and reduction in mean body weights of pups at weaning was observed. The average number of surviving pups per litter in the 1000 mg/kg group was lower as compared to the controls.Male and female F344/N rats for the 13-week base study were offspring (F1 generation) of the breeders from the perinatal phase of the study. Rats were approximately 34 days old: on the first day of the study.. Groups of 10 male and 10 female F1 rats were fed diets containing 0, 250, 500 and 1000 mg/kg of test chemical for 13 weeks after weaning. (No F1 offspring resulted from F0 females fed diets containing 2000 and 4000mg/kg doses so these exposure levels were not used in the 13-week rat study). Clinical findings were recorded and the animals were weighed initially, weekly, and at the end of the study; feed consumption was recorded as an average of grams per animal per day. Blood samples was collected at week 13 from core study (F1) for clinical pathology analyses .At the end of the study, samples from 0, 250, 500 and 1000mg/kg group F1 rats were collected for sperm motility and vaginal cytology evaluations. A necropsy was performed on all core study F1 animals. The results (F0 generation) of the study revealed no effect on gestation length, the average number of pups born per litter, the number of dams with stillborn pups(F0 generation) and dams exposed to 2000 or 4000 mg/kg did not litter. The results of the F1 generation rats revealed, hair discoloration in all except 250mg/kg treated rats. More number of pup deaths in 500 and 1000 mg/kg group than that of the control animals. Serum bile acid levels (in 500 and 1000mg./kg male and females and Serum alanine aminotransferase activity levels (in 250, 500 and 1000 mg/kg group males and female) were increased as comapred to control but not considered to be biologically significant. Increased incidence of hyperplasia of nasal respiratory epithelium (500 and 1000mg/kg) and increased incidences of splenic pigmentation (500 mg/kg and 1000 mg/kg)Thus, based on all the conclusions, the NOAEL for test chemical inF344/N rats was determined to be approximately 250 mg/kg bw.

Reproductive Toxicity Study 2:

The 13-week study was performed to evaluate the cumulative toxic effects with exposure to the chemical in Male and female B6C3F1mice and also to determine the  dose levels to be used in the long-term study. In the present study groups of 10 male and 10 female mice were fed diets containing 0, 2500, 5000, 10000, 20000, or 40000 ppm equivalent to (440, 880, 1950, 4000 and 8400mg/kg) in males and (500, 1100, 2200, 4600 and 9000mg/kg) of test chemical for 13 weeks. Feed and water were available ad libitum. Mice were housed five per cage. Clinical findings were recorded and the animals were weighed initially, weekly, and at the and of the study; feed consumption was recorded as an average of grams per animal per day. Blood was collected at week 13 from core study mice for selected hematology, clinical chemistry, and coagulation analyses. Additionally, clinical pathology analyses were performed on special groups Of 10 male and 10 female F1 mice fed diets containing 0,440, 1950mg/kg (for male) and 0, 500, 1100, 2200mg/kg (for females) test chemical for 3 weeks after weaning. Hematology and clinical chemistry .parameters were measured for these animals on day 5 and at week 3. For clinical pathology analyses, mice were anesthetized with CO 2 and bled from the retroorbital sinus. Clinical pathology parameters were measured for the 13-week rat study. At the end of the study, samples from 0, 440, 880 and  1950mg/kg (for male) and 0, 500, 1100 and 2200mg/kg (for females)  mice were collected for sperm motility and vaginal cytology evaluations. A necropsy was performed on all core study animals.

The results of the study revealed, hair discoloration of all the treated groups were observed, due to free spillage, these observation might have been due to dermal exposure of test chemical. Mortality were observed in two groups (2200 and 9000mg/kg) but the deaths were not considered due to exposure of test chemical. The body weight results revealed decrease in the body weight in male (1950, 4000 nad 8400mg/kg) and female mice (2200, 4600 and 9000mg/kg) groups to those of the control  groups. There were no change in feed consumption in any group exposure compared to control. The haemattological studies revealed significant increases in segmented neutrophil counts at week 3 and at the end of the study in the 4000 and 8400 mg/kg groups of males and females and in the 2200mg/kg female group. No change in body weigtt were seen at any dose levels. the gross pathology results estimated no lesions in the reproductive system of exposed groups of mice; thus, differences in reproductive parameters were considered to be secondary to body weight changes.Histopathologically increased incidences of mucosal hyperplasia were observed in the forestomach of male mice exposed to 4000 or 8400mg/kg and in female mice exposed to 2200, 4600, or 9000. The severity of this lesion also increased with increasing exposure concentration. Increased incidences of inflammation occurred in the nose and skin of males  (1950, 4000 and 8400mg/kg) and females (2200, 4600 and 9000mg/kg). However, left caudal, left epididymis, and left testis weights of males exposed to 1950 or 8400mg/kg were generally significantly lower than those of the controls. In females, The estrous cycle of 9000mg/kg group was significantly longer than that of the controls. Since, the reproductive effects in treated male and female observed at 1950 nad 9000mg/kg respectively, the lower doses of 880mg/kg in males and 4600mg/kg in females selected as a NOAEL value.

Reproductive Toxicity Study 3:

The safety of test chemical as an oil-soluble food grade antioxidant was evaluated in rat reproductive efficiency and placental transfer studies. A diet containing 0 % or 0.5 % (approx 250 mg/kg bw) of test chemical was fed to groups of 15 male and 15 female Sprague-Dawley rats for three successive generations. Rats were mated to produce two litters per generation. The following parameters were assessed: gonadal functions, estrus cycles, mating, conception rates, gestation rates, parturition, , and lactation.Measurements of these parameters in the treated group were similar to the controls. The NOAEL value of Approx 250 mg/kg bw was determined for the test chemical in a 3 generation study conducted on Sprague Dawley rats.The doses used in these reproductive studies may have been too low to adequately determine the potential of test chemical to have an effect on reproduction or to cause fetal malformations.

Effects on developmental toxicity

Description of key information

Developmental Toxicity Study:

The test chemical is an approved antioxidant used alone or in combination with other antioxidants in food. The teratogenic potential of test chemical is examined in this study.Test chemical was administered via the diet to pregnant Sprague-Dawley rats at concentrations of 0,0.125, 0.25or0.50% (about 0,125, 250 or 500 mg/kg bw) during the sensitive period of gestation. The experimental diets were fed from day 6 to day 16 of gestation. On day 20 of gestation, the females were killed by ether inhalation and examined for teratologic effects.The results of the present study revealled, the mean body weight and feed consumption of the dams were unaffected. The average number of corpora lutea, implantation sites, viable fetuses, resorptions and fetal body weights and mortality did not differ between the control and treated groups.Skeletal examinations revealed a significant number of fetuses with rudimentary ribs; however, the control group showed twice as many animals with this anomaly. Test chemical also did not result in any gross external or internal tissue anomalies in the fetuses.Therefore, it is concluded that test chemical fed to pregnant rats at doses up to 250 times the approved use level for humans caused no abnormalities in rat fetuses.The substance was considered to be non-teratogenic at all dose levels and the NOAEL is determined to be about 500 mg/kg bw in diet.

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

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 a publication.

Qualifier:

according to guideline

Guideline:

other:

Principles of method if other than guideline:

The experimental diets containing 0, 0.125, 0.25 or 0.50% test chemical were fed from day 6 to day 16 of gestation to Sprague-Dawley rats. On day 20 of gestation, the teratologic effects were examined.

GLP compliance:

not specified

Limit test:

no

Specific details on test material used for the study:

- Molecular weight (if other than submission substance):166.22 g/mol
- Substance type:Organic
- Physical state:solid

Species:

rat

Strain:

Sprague-Dawley

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Age at study initiation: Sexually mature Sprague-Dawley rats were used
- Diet (e.g. ad libitum): PURINA Laboratory Chow supplemented with 5.0% Mazola brand corn oil, ad libitum
- Water (e.g. ad libitum): Ad libitum

Route of administration:

oral: feed

Vehicle:

other: PURINA Laboratory Chow supple-mented with 5.0% Mazola brand corn oil

Details on exposure:

DIET PREPARATION
- Mixing appropriate amounts with (Type of food): Test chemical was added to ground PURINA Laboratory Chow supplemented with 5.0% Mazola brand corn oil in concentrations of 0, 0.125, 0.25 or 0.5%

Analytical verification of doses or concentrations:

not specified

Details on analytical verification of doses or concentrations:

No Data Available

Details on mating procedure:

- Impregnation procedure: cohoused
- M/F ratio per cage: 1:2
- Proof of pregnancy: Inseminations were verified by daily vaginal smears
- After successful mating each pregnant female was housed individually

Duration of treatment / exposure:

Day 6 to 16 of gestation period

Frequency of treatment:

daily

Duration of test:

Upto day 20 of gestation

Remarks:

Doses / Concentrations:
0, About 125, 250 or 500 mg/kg
Basis:
nominal in diet
0, 0.125, 0.25 or 0.50%

No. of animals per sex per dose:

Control: 20 females
125 mg/kg: 20 females
250 mg/kg: 20 females
500 mg/kg: 20 females

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: The selected doses were 62.5, 125 and 250 times the approved use level for humans
- Rationale for animal assignment: Inseminated females were randomly assigned to one of four groups (20 rats/group)

Maternal examinations:

Individual body weights were recorded on the day of insemination and daily throughout the period of gestation. Food consumption was monitored from day 0 to 20 of gestation. The mean body weight of the fetuses was calculated by averaging the body weight per litter within each group and calculating the mean of these values to obtain the group mean.

Ovaries and uterine content:

On day 20 of gestation, the females were killed by ether inhalation and the uteri were exposed by laparotomy. Total implantation sites were counted and categorized as consisting of live fetuses, dead fetuses or resorptions.
The ovaries of all females were removed and the total number of corpora lutea was recorded

Fetal examinations:

The fetuses were removed from the placentae, blotted dry, carefully examined for gross anomalies, sexed and weighed. Half of the fetuses were fixed in Bouin's fixative and examined for internal soft tissue anomalies according to the free-hand razor blade technique. The other half were fixed in 95% ethanol, eviscerated, macerated in KOH and stained with alizarin red for skeletal examination.

Statistics:

No Data Available

Historical control data:

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

Food consumption and compound intake (if feeding study):

no effects observed

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:

not specified

Gross pathological findings:

no effects observed

Neuropathological findings:

not specified

Histopathological findings: non-neoplastic:

no effects observed

Histopathological findings: neoplastic:

not specified

Other effects:

not specified

Details on results:

No Data Available

Number of abortions:

no effects observed

Pre- and post-implantation loss:

no effects observed

Total litter losses by resorption:

no effects observed

Early or late resorptions:

no effects observed

Dead fetuses:

no effects observed

Changes in pregnancy duration:

not specified

Changes in number of pregnant:

not specified

Other effects:

not specified

Details on maternal toxic effects:

Maternal toxic effects:no effects
Details on maternal toxic effects:
No effect on the mean body weight gain or food consumption of the dams
Average number of corpora lutea, implantation sites, viable fetuses, resorptions and body weight of the fetuses per litter, fetal mortality and sex ratio were unaffected at 125, 250 or 500 mg/kg levels of treatment.

Dose descriptor:

NOAEL

Effect level:

500 mg/kg bw/day

Based on:

test mat.

Basis for effect level:

body weight and weight gain

dead fetuses

early or late resorptions

food consumption and compound intake

gross pathology

histopathology: non-neoplastic

maternal abnormalities

number of abortions

pre and post implantation loss

total litter losses by resorption

Remarks on result:

other: teratogenicity was measured

Abnormalities:

not specified

Fetal body weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

Some sporadic cases were observed wherein the animals in 500 mg/kg were found to be of lesser weight whn compared to control.

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

no effects observed

Changes in postnatal survival:

no effects observed

External malformations:

no effects observed

Skeletal malformations:

no effects observed

Visceral malformations:

no effects observed

Other effects:

not specified

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects
Details on embryotoxic / teratogenic effects:
A significant number of rudimentary ribs were seen in all groups. However, the incidence of this variation was two times greater in the control group than in any treatment group (125, 250 or 500 mg/kg)
The ingestion of test chemical did not cause any gross external or internal tissue anomalies.

Dose descriptor:

NOAEL

Effect level:

500 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: teratogenicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Conclusions:

The NOAEL value for the test chemical to Sprague-Dawley rats was determined to be about 500 mg/kg bw in diet. The substance did not produce any teratogenic effects at this dose level.

Executive summary:

The test chemical is an approved antioxidant used alone or in combination with other antioxidants in food. The teratogenic potential of test chemical is examined in this study. Test chemical was administered via the diet to pregnant Sprague-Dawley rats at concentrations of 0,0.125, 0.25or0.50% (about 0,125, 250 or 500 mg/kg bw) during the sensitive period of gestation. The experimental diets were fed from day 6 to day 16 of gestation. On day 20 of gestation, the females were killed by ether inhalation and examined for teratologic effects. The results of the present study revealled, the mean body weight and feed consumption of the dams were unaffected. The average number of corpora lutea, implantation sites, viable fetuses, resorptions and fetal body weights and mortality did not differ between the control and treated groups. Skeletal examinations revealed a significant number of fetuses with rudimentary ribs; however, the control group showed twice as many animals with this anomaly. Test chemical also did not result in any gross external or internal tissue anomalies in the fetuses. Therefore, it is concluded that test chemical fed to pregnant rats at doses up to 250 times the approved use level for humans caused no abnormalities in rat fetuses. The substance was considered to be non-teratogenic at all dose levels and the NOAEL is determined to be about 500 mg/kg bw in diet.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LOAEL

500 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Quality of whole database:

K2 level data refered from reliable source of journal

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 Studies:

The summaries of the developmental toxicity study is:

Developmental Toxicity Study 1:

The test chemical is an approved antioxidant used alone or in combination with other antioxidants in food. The teratogenic potential of test chemical is examined in this study.Test chemical was administered via the diet to pregnant Sprague-Dawley rats at concentrations of 0,0.125, 0.25or0.50% (about 0,125, 250 or 500 mg/kg bw) during the sensitive period of gestation. The experimental diets were fed from day 6 to day 16 of gestation. On day 20 of gestation, the females were killed by ether inhalation and examined for teratologic effects.The results of the present study revealled, the mean body weight and feed consumption of the dams were unaffected. The average number of corpora lutea, implantation sites, viable fetuses, resorptions and fetal body weights and mortality did not differ between the control and treated groups.Skeletal examinations revealed a significant number of fetuses with rudimentary ribs; however, the control group showed twice as many animals with this anomaly. Test chemical also did not result in any gross external or internal tissue anomalies in the fetuses.Therefore, it is concluded that test chemical fed to pregnant rats at doses up to 250 times the approved use level for humans caused no abnormalities in rat fetuses.The substance was considered to be non-teratogenic at all dose levels and the NOAEL is determined to be about 500 mg/kg bw in diet.

Developmental Toxicity Study 2:

The study was performed to evaluate the cumulative toxic effects of test chemical on female rats.The test chemical was fed to female F344/N rats at a doses of  0, 2500, 5000, 10000, 20000, or 40000 ppm (Approx 250, 500, 1000, 2000 or 4000 mg/kg bw) in diet for approximately 6 weeks (from 2 weeks prior to cohabitation until weaning of F1 pups). Clinical findings, body weights, and feed consumption were recorded weekly for F0 females during the first 2 weeks and during lactation.In the 500 and 1000 mg/kg group, more number of pup deaths and reduction in mean body weights of pups at weaning was observed. The average number of surviving pups per litter in the 1000 mg/kg group was lower as compared to the controls.No effect was observed on gestation length, the average number of pups born per litter, or the number of dams with stillborn pups for dams up to 1000 mg/kg.The dams exposed to 2000 or 4000 mg/kg did not litter.The LOAEL (Lowest Observed Adverse Effect Level) value for F344 rats was found to be 500 mg/kg/day for the F1 generation.

Justification for classification or non-classification

Based on above results it can be concluded that the test chemical is considered to be not toxic to reproduction as well as developmental toxicity and does not qualify as toxicant according to CLP classification.