[No public or meaningful name is available]

Toxicity to reproduction

Administrative data

Endpoint:

one-generation reproductive toxicity

Remarks:

based on test type (migrated information)

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Study period:

1993

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted according to valid methods and considered reliable, adequate and relevant.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 415 [One-Generation Reproduction Toxicity Study (before 9 October 2017)]

GLP compliance:

yes

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Soybean oil, epoxidized

EC Number:

232-391-0

EC Name:

Soybean oil, epoxidized

Cas Number:

8013-07-8

IUPAC Name:

8013-07-8

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): Soybean oil, epoxidized; ESBO; Reoplast 39
- Molecular formula (if other than submission substance): C57-H98-O12 (typical structure example)
- Substance type: UVCB
- Physical state: Amber light liquid
- Analytical purity: 100 % w/w

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: Soybean Oil (SBO)

Details on mating procedure:

- M/F ratio per cage: 1/1
- Length of cohabitation: Each male was paired with one female of the same treatment group for the night. The female was placed with the same male until mating occurred or 10 days had elapsed.
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
- After 10 days of unsuccessful pairing and 3 days rest period replacement of first male by another male with proven fertility. (If no evidence of mating was observed after 10 days, the female was placed after 3 days rest period with another male that had already successfully mated, until mating occurred or 11 days had elapsed.)
- Further matings after two unsuccessful attempts: Not provided.
- After successful mating each pregnant female was caged (how): Not provided.

Duration of treatment / exposure:

Pretreatment: 71 days in males; 15 days in female; then F0 male and female rats paired. Treatment continued in females during the mating, pregnancy and lactation until day 21 post-partum (PP) and in males until day 21 PP of F1 litters.

Frequency of treatment:

Daily, 7 times a week

Doses / concentrations

Remarks:

Doses / Concentrations:
100, 300 and 1000 mg/kg bw/day
Basis:
actual ingested

No. of animals per sex per dose:

28

Control animals:

yes, concurrent vehicle

Examinations

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: F0 animals were observed daily for clinical signs.

DETAILED CLINICAL OBSERVATIONS: No data
- Time schedule:/

BODY WEIGHT: Yes
- Time schedule for examinations: Body weight was recorded for each males on the first day of treatment (day 1) and then once a week until sacrifice. Body weight was recorded for each females on the first day of treatment (day 1), once a week before mating and during mating periods, on days 0, 7, 14 and 20 of pregnancy, and on days 1, 7, 14 and 21 postpartum.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study):
The quantity of food consumed by each male was recorded once a week over a 7-day period of treatment until sacrifice. The quantity of food consumed by each female was recorded as follows:
during the premating period, once a week over a 7-day period,
during pregnancy, at the intervals day 0-day 7, day 7-day 14 and day 14-day 20,
during lactation, at the intervals day 1 pp-day 7 pp, day 7pp-day 14 pp and day 14 pp-day 21 pp.
However, food consumption was not measured during the mating period. Food intake per animal and per day was calculated using the amount of food given and left in each feeder.


WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No data
- Time schedule for examinations: /

OTHER:
Any animal showing signs of poor clinical condition, especially if death appeared imminent, was asphyxiated by carbon dioxide and killed by exsanguination. Any animal found dead or killed due to poor clinical condition was subjected to macroscopic examination and a full spectrum of tissues was preserved whenever possible.

About 24 hours after the last administration, hematology and blood chemistry investigations were performed in 5 males and 5 females of each group.
At the end of the study, macroscopic examination of all F0 males and females (and F1 pups) was performed. In all the parents, reproductive organs and macroscopic lesions were sampled and additionally in 5 males and 5 females of each group ileum, kidneys and liver were sampled.
Microscopic examination of the reproductive organs was performed in all animals of the control and 1000 mg /kg bw/day groups and in animals suspected of infertility, those that died or were sacrificed in the 100 and 300 mg/kg bw/day groups. In addition the livers of one male of the 1000 mg/kg
bw/day group and of the control group were also examined microscopically.

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes. On day 4 post-partum, the size of each litter was adjusted by eliminating extra pups by random selection, as nearly as possible, at 4 males and 4 females per litter. Whenever, the number of male or female pups prevented having at least 4 of each sex per litter, partial adjustment (for example, 5 males and 3 females) was made. Adjustments were not made for litters of less than 8 pups.
The number of pups in each litter exhibiting the following characteristics was recorded:
on day 5 post-partum: pinna unfolding, hair growth, surface righting reflex
on day 11 post-partum: cliff avoidance
on day 13 post-partum: incisor eruption
on day 17 post-partum: eye opening, auricular duct opening, air righting reflex
- If yes, maximum of 8 pups/litter (4/sex/litter as nearly as possible); excess pups were killed and discarded.

PARAMETERS EXAMINED
The following parameters were examined in F1 / offspring: The F1 litters were examined daily for clinical signs, viability, physical and reflex development until day 21 post-partum. Pup body weights were recorded on days 1, 4, 7, 14 and 21 post-partum (each pup was identified by tattoo).
[number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities, other:]

GROSS EXAMINATION OF DEAD PUPS:
yes, at the end of the study, macroscopic examination of all F1 pups was performed.

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals at the end of the study. About 24 hours after the last administration, hematology and blood chemistry investigations were performed in 5 males (and 5 females) of each group.
- Maternal animals: All surviving animals at the end of the study. About 24 hours after the last administration, hematology and blood chemistry investigations were performed in (5 males and) 5 females of each group.

GROSS NECROPSY
- Gross necropsy consisted of [external and internal examinations including the cervical, thoracic, and abdominal viscera.]
In all the parents, reproductive organs and macroscopic lesions were sampled and additionally in 5 males and 5 females of each group ileum, kidneys and liver were sampled.

HISTOPATHOLOGY / ORGAN WEIGHTS
Microscopic examination of the reproductive organs was performed in all animals of the control and 1000 mg /kg bw/day groups and in animals suspected of infertility, those that died or were sacrificed in the 100 and 300 mg/kg bw/day groups. In addition the livers of one male of the 1000 mg/kg bw/day group and of the control group were also examined microscopically.

Statistics:

Statistical analysis: Mean values were compared by one-way variance analysis and Dunnett's test. Percentage values were compared by Fisher's exact probability test.
The following sequence was used for the statistical analysis of haematology and blood biochemistry data:
The normality of the distribution of the values in each group was checked by Komolgorov Smirnov's test (1948). If the distribution was normal, the homogeneity of variances between the groups was assessed by Bartlett's test (1937)(more than 2 groups) or Fisher's test (1934) (2 groups). If no significant heterogeneity of the variances was established, the comparison between treated and control groups was performed by Dunnett's test (1955). If the variances were heterogeneous, the comparison between treated and control groups was performed by Dunn's test (1964) (more than 2 groups) or by Mann Whitney's test (1947) (2 groups).
If the distribution of values in the group was not normal, the analysis was repeated after logarithmic transformation of the values. If this logarithmic transformation fails to normalise the distribution of the values, comparison of treated and control groups was performed by Dunn's test using original values.

Reproductive indices:

Reproductive data: The mating and fertility indices of males and females were similar in the control and treated groups.
Mating
Males: The mating index of males was similar in the control group (96.4%) and the 100 (89.3%), 300 (96.4%) and 1000 (96.4%) mg/kg bw/day groups.
Females: The mating index of females was 100 % in the control group and the 100 and 300 mg/kg bw/day groups. It was 96.4% in the 1000 mg/kg bw/day group as 1 female did not mate. This is not related to the treatment.
Fertility
Males: The fertility index of males was similar in the control group (81.5%) and the 100 (88.5%), 300 (92.6%) and 1000 (100.0%) mg/kg bw/day groups.
Females: The fertility index of females was similar in the control group (82.1%) and the 100 (92.9%), 300 (92.9%) and 1000 (100.0%) mg/kg bw/day groups.

Offspring viability indices:

Litter data:
The gestation index and the mean duration of gestation were similar in all groups. The live birth index was 100% in all groups. The viability indices on day 4 and day 21 post-partum, the physical and reflex development of pups and the mean pup body weight were similar in the control and treated groups.
-Gestation: The mean number of implantation sites was similar in the control and treated groups. The gestation index was 100% in the control, 300 and 1000 mg/kg bw/day groups. In the 100 mg/kg bw/day group, it was 96.2% as one pregnant female was sacrificed on day 9 of pregnancy (due to a misdosing). The mean duration of gestation was similar in all groups and within the normal range of 21-22 days. The live birth index was 100 % in all groups.
-Pups: The viability index on day 4 post-partum was similar in the control (94.2%) and the 100 (91.1 %), 300 (97.9%) and 1000 (93.5%) mg/kg bw/day groups. On day 21 post-partum, the lactation index was also similar in the control (97.8%) and the 100 (98.9%), 300 (97.9%) and 1000 (94.4%) mg/kg bw/day groups. The physical development and reflex development were similar in the control and treated groups. No clinical signs attributed to the treatment were observed in pups of any group. No treatment-related macroscopic changes were noted in pups sacrificed at the end of the study. In pups culled on day 4 post-partum, no macroscopic changes were noted in the control and the 100 mg/kg bw/day groups. In the 300 and 1000 mg/kg bw/day groups I out of 208 (0.5%) pups and 6 out of 203 (3.0%; p < 00.5) pups had a dilated renal pelvis. In the 1000 mg/kg bw/day group, 5 out of the 6 affected pups provided from the same litter whose mother was found dead on day 7 postpartum. This litter had a mean body weight lower than that of the other litters of the same group. Consequently the dilatation of renal pelvis of these pups is related to their slight delayed development related itself to the clinical conditions of the mother.

Results and discussion

Results: P0 (first parental generation)

General toxicity (P0)

Clinical signs:

no effects observed

Description (incidence and severity):

No treatment related mortalities or clinical signs were observed.

Body weight and weight changes:

no effects observed

Description (incidence and severity):

The mean food consumption and body weight gain of males and females were similar in the control and treated groups.

Food consumption and compound intake (if feeding study):

no effects observed

Description (incidence and severity):

The mean food consumption and body weight gain of males and females were similar in the control and treated groups.

Histopathological findings: non-neoplastic:

no effects observed

Description (incidence and severity):

The microscopic examination of the animals did not reveal any changes attributed to the treatment.

Other effects:

not examined

Description (incidence and severity):

Test substance intake: oral gavage

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:

no effects observed

Description (incidence and severity):

Regular oestrous cycle as evidenced from the morphological changes in the ovaries uterus and vagina, was noted in the control and treated female rats, except in 2 females given 100 mg/kg bw/day. Weak incidence is not considered of toxicological importance

Reproductive performance:

no effects observed

Description (incidence and severity):

The mating and fertility indices of males and females were similar in the control and treated groups.

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
No treatment related mortalities or clinical signs were observed.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
The mean food consumption and body weight gain of males and females were similar in the control and treated groups.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS): oral gavage

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
Reproductive data: The mating and fertility indices of males and females were similar in the control and treated groups.Details:
-Mating
Males: The mating index of males was similar in the control group (96.4%) and the 100 (89.3%), 300 (96.4%) and 1000 (96.4%) mg/kg bw/day groups.
Females: The mating index of females was 100 % in the control group and the 100 and 300 mg/kg bw/day groups. It was 96.4% in the 1000 mg/kg bw/day group as 1 female did not mate. This is not related to the treatment.
-Fertility
Males: The fertility index of males was similar in the control group (81.5%) and the 100 (88.5%), 300 (92.6%) and 1000 (100.0%) mg/kg bw/day groups.
Females: The fertility index of females was similar in the control group (82.1%) and the 100 (92.9%), 300 (92.9%) and 1000 (100.0%) mg/kg bw/day groups.

ORGAN WEIGHTS (PARENTAL ANIMALS)

GROSS PATHOLOGY (PARENTAL ANIMALS)
The macroscopic examination of the animals did not reveal any changes attributed to the treatment.
Macroscopic examination: Liver enlargement, without any relevant histopathological findings, was noted in 1 out of 28 males given 1000 mg/kg bw/day. Paleness of the liver was found in 2 out of 28 males given 300 mg/kg bw/day. These findings were considered to be of spontaneous nature and irrelevant to the treatment with the test substance. The other macroscopic findings encountered were those which are commonly recorded spontaneously in the rat of this strain and age and were considered to be of no toxicological importance.

HISTOPATHOLOGY (PARENTAL ANIMALS)
The microscopic examination of the animals did not reveal any changes attributed to the treatment.
Microscopic examination: Regular oestrous cycle, as evidenced from the morphological changes in the ovaries, uterus and vagina, was noted in the control and treated female rats, except in 2 females given 100 mg/kg bw/day. In these two animals deciduomatosis was noted in one female; severe metritis and degenerated placenta were noted in the other female. These changes can be found in untreated rats, therefore this weak incidence is not considered of toxicological importance. No treatment-related abnormalities were found in the male genital organs of the animals examined

OTHER FINDINGS (PARENTAL ANIMALS)
The variations of hematologic or blood chemistry parameters were not of toxicological importance.
The statistically significant differences noted between control and treated animals in some haematological parameters, namely total white and red blood cell count in males, were considered to be of no toxicological importance as they were minor, not dose-related and the individual values were within the range of our background data (white blood cells: 5.9 - 15.6 G/l; red blood cells: 8.05 - 9.80T/1). No perceptible differences between control and treated animals in the other parameters.

Effect levels (P0)

Dose descriptor:

NOEL

Remarks:

reproduction

Effect level:

1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks:

100% purity > 100% active ingredient

Sex:

male/female

Basis for effect level:

other: the highest tested dose tested

Results: F1 generation

General toxicity (F1)

Clinical signs:

no effects observed

Description (incidence and severity):

The physical development and reflex development were similar in the control and treated groups. No clinical signs attributed to the treatment were observed in pups of any group.

Mortality / viability:

no mortality observed

Description (incidence and severity):

The viability index on day 4 post-partum was similar in the control (94.2%) and the 100 (91.1 %), 300 (97.9%) and 1000 (93.5%) mg/kg bw/day groups.

Gross pathological findings:

no effects observed

Description (incidence and severity):

See details

Details on results (F1)

VIABILITY (OFFSPRING)
The live birth index was 100% in all groups. The viability index on day 4 post-partum was similar in the control (94.2%) and the 100 (91.1 %), 300 (97.9%) and 1000 (93.5%) mg/kg bw/day groups.

CLINICAL SIGNS (OFFSPRING)
The physical development and reflex development were similar in the control and treated groups. No clinical signs attributed to the treatment were observed in pups of any group.

BODY WEIGHT (OFFSPRING)

SEXUAL MATURATION (OFFSPRING)

ORGAN WEIGHTS (OFFSPRING)

GROSS PATHOLOGY (OFFSPRING)
No treatment-related macroscopic changes were noted in pups sacrificed at the end of the study. In pups culled on day 4 post-partum, no macroscopic changes were noted in the control and the 100 mg/kg bw/day groups. In the 300 and 1000 mg/kg bw/day groups I out of 208 (0.5%) pups and 6 out of 203 (3.0%; p < 00.5) pups had a dilated renal pelvis. In the 1000 mg/kg bw/day group, 5 out of the 6 affected pups provided from the same litter whose mother was found dead on day 7 postpartum. This litter had a mean body weight lower than that of the other litters of the same group. Consequently the dilatation of renal pelvis of these pups is related to their slight delayed development related itself to the clinical conditions of the mother. This dilatation of renal pelvis is not attributed to the treatment.

HISTOPATHOLOGY (OFFSPRING)

OTHER FINDINGS (OFFSPRING)
On day 21 post-partum, the lactation index was also similar in the control (97.8%) and the 100 (98.9%), 300 (97.9%) and 1000 (94.4%) mg/kg bw/day groups.

Effect levels (F1)

Dose descriptor:

NOAEL

Remarks:

reproduction

Generation:

F1

Effect level:

>= 1 000 mg/kg bw/day (actual dose received)

Based on:

test mat.

Remarks:

100% purity > 100% active ingredient

Overall reproductive toxicity

Reproductive effects observed:

not specified

Any other information on results incl. tables

No treatment related mortalities or clinical signs were observed. The mean food consumption and body weight gain of males and females were similar in the control and treated groups. The variations of hematologic or blood chemistry parameters were not of toxicological importance. The macroscopic and microscopic examination of the animals did not reveal any changes attributed to the treatment.

Reproductive data: The mating and fertility indices of males and females were similar in the control and treated groups.

Litter data: The gestation index and the mean duration of gestation were similar in all groups. The live birth index was 100% in all groups. The viability indices on day 4 and day 21 post-partum, the physical and reflex development of pups and the mean pup body weight were similar in the control and treated groups.

Details: Mating Males: The mating index of males was similar in the control group (96.4%) and the 100 (89.3%), 300 (96.4%) and 1000 (96.4%) mg/kg bw/day groups.

Females: The mating index of females was 100 % in the control group and the 100 and 300 mg/kg bw/day groups. It was 96.4% in the 1000 mg/kg bw/day group as 1 female did not mate. This is not related to the treatment.

Fertility Males: The fertility index of males was similar in the control group (81.5%) and the 100 (88.5%), 300 (92.6%) and 1000 (100.0%) mg/kg bw/day groups.

Females: The fertility index of females was similar in the control group (82.1%) and the 100 (92.9%), 300 (92.9%) and 1000 (100.0%) mg/kg bw/day groups.

Gestation: The mean number of implantation sites was similar in the control and treated groups. The gestation index was 100% in the control, 300 and 1000 mg/kg bw/day groups. In the 100 mg/kg bw/day group, it was 96.2% as one pregnant female was sacrificed on day 9 of pregnancy (due to a misdosing). The mean duration of gestation was similar in all groups and within the normal range of 21-22 days. The live birth index was 100 % in all groups.

Pups: The viability index on day 4 post-partum was similar in the control (94.2%) and the 100 (91.1 %), 300 (97.9%) and 1000 (93.5%) mg/kg bw/day groups. On day 21 post-partum, the lactation index was also similar in the control (97.8%) and the 100 (98.9%), 300 (97.9%) and 1000 (94.4%) mg/kg bw/day groups. The physical development and reflex development were similar in the control and treated groups. No clinical signs attributed to the treatment were observed in pups of any group. No treatment-related macroscopic changes were noted in pups sacrificed at the end of the study. In pups culled on day 4 post-partum, no macroscopic changes were noted in the control and the 100 mg/kg bw/day groups. In the 300 and 1000 mg/kg bw/day groups I out of 208 (0.5%) pups and 6 out of 203 (3.0%; p < 00.5) pups had a dilated renal pelvis. In the 1000 mg/kg bw/day group, 5 out of the 6 affected pups provided from the same litter whose mother was found dead on day 7 postpartum. This litter had a mean body weight lower than that of the other litters of the same group. Consequently the dilatation of renal pelvis of these pups is related to their slight delayed development related itself to the clinical conditions of the mother. This dilatation of renal pelvis is not attributed to the treatment.

Haematology: The statistically significant differences noted between control and treated animals in some haematological parameters, namely total white and red blood cell count in males, were considered to be of no toxicological importance as they were minor, not dose-related and the individual values were within the range of our background data (white blood cells: 5.9 - 15.6 G/l; red blood cells: 8.05 - 9.80T/1). No perceptible differences between control and treated animals in the other parameters.

PATHOLOGY:

Macroscopic examination: Liver enlargement, without any relevant histopathological findings, was noted in 1 out of 28 males given 1000 mg/kg bw/day. Paleness of the liver was found in 2 out of 28 males given 300 mg/kg bw/day. These findings were considered to be of spontaneous nature and irrelevant to the treatment with the test substance. The other macroscopic findings encountered were those which are commonly recorded spontaneously in the rat of this strain and age and were considered to be of no toxicological importance.

Microscopic examination: Regular oestrous cycle, as evidenced from the morphological changes in the ovaries, uterus and vagina, was noted in the control and treated female rats, except in 2 females given 100 mg/kg bw/day. In these two animals deciduomatosis was noted in one female; severe metritis and degenerated placenta were noted in the other female. These changes can be found in untreated rats, therefore this weak incidence is not considered of toxicological importance. No treatment-related abnormalities were found in the male genital organs of the animals examined.

Applicant's summary and conclusion

Conclusions:

The test substance ESBO administered daily by the oral route (gavage) to male and female Sprague-Dawley rats at the 100, 300 and 1000 mg/kg bw/day dose levels 71 days before mating in males and 15 days before mating in females until day 21 post-partum of F1 litters did not induce any toxic effects in parent males and females, did not disturb their capacity of reproduction and did not impair the development of the F1 offspring.
Under the experimental conditions, the highest tested dose of 1000 mg/kg bw/day was found to be the No Observed Effect Level.

Executive summary:

The test substance Epoxidized Soybean Oil (ESBO) was given daily 7 times a week to 3 groups of 28 F0 male and 28 F0 female Sprague-Dawley rats by gavage at the dose levels of 100, 300 and 1000 mg/kg bw/day. The F0 control animals (28 males and 28 females) received the vehicle Soybean Oil (SBO).

After 71 days of treatment in males and 15 days of treatment in females, the F0 male and female rats were paired.

Each male was paired with one female of the same treatment group for the night. The female was placed with the same male until mating occurred or 10 days had elapsed. If no evidence of mating was observed after 10 days, the female was placed after 3 days rest period with another male that had already successfully mated, until mating occurred or 11 days had elapsed. Each morning, a vaginal lavage was performed in order to detect the presence of spermatozoa. The day when spermatozoa were found was designated as day 0 of pregnancy.

Treatment was continued in females during the mating and pregnancy and lactation periods until day 21 post-partum and in males during the mating period until day 21 post-partum

of F1 litters.

No treatment related mortalities or clinical signs were observed. The mean food consumption and body weight gain of males and females were similar in the control and treated groups. The variations of hematologic or blood chemistry parameters were not of toxicological importance. The macroscopic and microscopic examination of the animals did not reveal any changes attributed to the treatment.

Reproductive data: The mating and fertility indices of males and females were similar in the control and treated groups.

Litter data: The gestation index and the mean duration of gestation were similar in all groups. The live birth index was 100% in all groups. The viability indices on day 4 and day 21 post-partum, the physical and reflex development of pups and the mean pup body weight were similar in the control and treated groups.

Details: Mating Males: The mating index of males was similar in the control group (96.4%) and the 100 (89.3%), 300 (96.4%) and 1000 (96.4%) mg/kg bw/day groups.

Females: The mating index of females was 100 % in the control group and the 100 and 300 mg/kg bw/day groups. It was 96.4% in the 1000 mg/kg bw/day group as 1 female did not

mate. This is not related to the treatment.

Fertility Males: The fertility index of males was similar in the control group (81.5%) and the 100 (88.5%), 300 (92.6%) and 1000 (100.0%) mg/kg bw/day groups.

Females: The fertility index of females was similar in the control group (82.1%) and the 100 (92.9%), 300 (92.9%) and 1000 (100.0%) mg/kg bw/day groups.

Gestation: The mean number of implantation sites was similar in the control and treated groups. The gestation index was 100% in the control, 300 and 1000 mg/kg bw/day groups. In the 100 mg/kg bw/day group, it was 96.2% as one pregnant female was sacrificed on day 9 of pregnancy (due to a misdosing). The mean duration of gestation was similar in all groups and within the normal range of 21-22 days. The live birth index was 100 % in all groups.

Pups: The viability index on day 4 post-partum was similar in the control (94.2%) and the 100 (91.1 %), 300 (97.9%) and 1000 (93.5%) mg/kg bw/day groups. On day 21 post-partum, the lactation index was also similar in the control (97.8%) and the 100 (98.9%), 300 (97.9%) and 1000 (94.4%) mg/kg bw/day groups. The physical development and reflex development were similar in the control and treated groups. No clinical signs attributed to the treatment were observed in pups of any group. No treatment-related macroscopic changes were noted in pups sacrificed at the end of the study. In pups culled on day 4 post-partum, no macroscopic changes were noted in the control and the 100 mg/kg bw/day groups. In the 300 and 1000 mg/kg bw/day groups I out of 208 (0.5%) pups and 6 out of 203 (3.0%; p < 00.5) pups had a dilated renal pelvis. In the 1000 mg/kg bw/day group, 5 out of the 6 affected pups provided from the same litter whose mother was found dead on day 7 postpartum. This litter had a mean body weight lower than that of the other litters of the same group. Consequently the dilatation of renal pelvis of these pups is related to their slight delayed development related itself to the clinical conditions of the mother. This dilatation of renal pelvis is not attributed to the treatment.

Haematology: The statistically significant differences noted between control and treated animals in some haematological parameters, namely total white and red blood cell count in males, were considered to be of no toxicological importance as they were minor, not dose-related and the individual values were within the range of our background data (white blood cells: 5.9 - 15.6 G/l; red blood cells: 8.05 - 9.80T/1). No perceptible differences between control and treated animals in the other parameters.

PATHOLOGY:

Macroscopic examination: Liver enlargement, without any relevant histopathological findings, was noted in 1 out of 28 males given 1000 mg/kg bw/day. Paleness of the liver was found in 2 out of 28 males given 300 mg/kg bw/day. These findings were considered to be of spontaneous nature and irrelevant to the treatment with the test substance. The other macroscopic findings encountered were those which are commonly recorded spontaneously in the rat of this strain and age and were considered to be of no toxicological importance.

Microscopic examination: Regular oestrous cycle, as evidenced from the morphological changes in the ovaries, uterus and vagina, was noted in the control and treated female rats, except in 2 females given 100 mg/kg bw/day. In these two animals deciduomatosis was noted in one female; severe metritis and degenerated placenta were noted in the other female. These changes can be found in untreated rats, therefore this weak incidence is not considered of toxicological importance. No treatment-related abnormalities were found in the male genital organs of the animals examined.

CONCLUSION:

The test substance ESBO administered daily by the oral route (gavage) to male and female Sprague-Dawley rats at the 100, 300 and 1000 mg/kg bw/day dose levels 71 days before mating in males and 15 days before mating in females until day 21 post-partum of F1 litters did not induce any toxic effects in parent males and females, did not disturb their capacity of reproduction and did not impair the development of the F1 offspring.

Under the experimental conditions, the highest tested dose of 1000 mg/kg bw/day was found to be the No Observed Effect Level.