1,2-epoxybutane

Endpoint summary

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Effects on fertility

Description of key information

No study on reproductive toxicity of 1,2-epoxybutane are available. No indications of reproductive toxicity were seen in repeated dose toxicity studies, in the dominant lethal test in vivo and in developmental toxicity studies. However, reproduction toxicity data are available for the structural analogue substance propylene oxide (CAS 75-56-9), which are used as read-across. No adverse effects on fertility were found in a two-generation study (GLP and similar to OECD guideline 416) in which groups of 30 male and 30 female F344 rats were exposed to 0, 70, 240 and 710 mg/m3, propylene oxide vapour, whole body, for 6 hours/day, 5 days/week for 14 weeks prior to mating. The NOAEC for reproductive toxicity is 710 mg/m3 (300 ppm).

Link to relevant study records

Reference

Endpoint:

two-generation reproductive toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1983-04-04 to 1985-06-19

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 416 (Two-Generation Reproduction Toxicity Study)

GLP compliance:

yes

Remarks:

Mammalian and Environmental Toxicology Research Laboratory Health and Environmental Sciences, Dow Chemical, USA. Midland, Michigan 48640

Limit test:

no

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratory, Kingston NY
- Age at study initiation: 7 weeks (P), 5 weeks (F1)
- Weight at study initiation: weight range of 128.3-129.6 g (males P) and
100.8-101.6 g (females P) ; weight range of 97.4-104.2g (males F1) and 86.1-88.1 g (females F1)
- Fasting period before study: no data
- Housing: singly in wire mesh-bottemed, stainless steel cages
- Diet: ad libitum (except during the exposure periods)
- Water: ad libitum (except during the exposure periods)
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22
- Humidity (%): 40 - 60
- Air changes (per hr): 6 h/day exposure to test material
- Photoperiod (hrs dark / hrs light): 12 hour photocycle

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

unchanged (no vehicle)

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
Exposures were conducted in 14.5 cubic meter chambers with stainless steel pyramidal shaped ceilings and epoxy resin coated floors and walls. All chambers were operated under dyhamic airflow conditions at a slight negative pressure relative to the surrounding area. Control animals were placed in an identical chamber. Air supplied to the chambers was controlled by a system designed to control temperature (approximately 22°C) and relative humidity (approximately 50%). These data were recorded each exposure day.

Propylene oxide test atmospheres were generated by vaporizing the liquid at controlled rates using glass J-tubes (Miller et al., 1980). The vapors were swept from the J-tubes with compressed air into the air inlet ducts of the chambers where there was further dilution to the desired concentration. The compressed air was preheated wlth a flameless heat torch (Master, Model FHT-4) at approximately 34 degrees C, in order to facilitate complete vaporization of the liquid test material. Total chamber airflow was maintained at approximately 2200 liters per minute.

The concentration of the test material in each chamber was determined at least once per hour by a MIRAN I infrared spectrophotometer at a wavelength of 11.95 microns. The nominal concentration (ratio of the amount of test material to the total anount of air through the chamber) was calculated for each chamber on a daily basis.

Details on mating procedure:

- M/F ratio per cage: 1:1
- Length of cohabitation: two 5 day cohabitation periods
- Proof of pregnancy: sperm in vaginal smear referred to as day 0 of pregnancy
- After successful mating each pregnant female was caged: individually and during late gestation the cages contained ground corn cob bedding
- Any other deviations from standard protocol: avoiding brother-sister matings

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Once an hour by MIRAN 1 infrared spectrophotometer at a wavelength of 11.95 microns. The nominal concentration was calculated for each chamber on a daily basis.

Duration of treatment / exposure:

6 hours a day

Frequency of treatment:

5 days a week during the premating period
7 days a week during mating, gestation and lactation

Details on study schedule:

- F1 parental animals not mated until 17 weeks after selected from the F1 litters.
- Selection of parents from F1 generation when pups were 4 weeks of age.
- Age at mating of the mated animals in the study: 21 weeks

Dose / conc.:

30 ppm (nominal)

Remarks:

70 mg/m3 (analytical)

Dose / conc.:

100 ppm (nominal)

Remarks:

240 mg/m3 (analytical)

Dose / conc.:

300 ppm (nominal)

Remarks:

710 mg/m3 (analytical)

No. of animals per sex per dose:

30

Control animals:

yes

Details on study design:

- Dose selection rationale: based on the results of the chronic study in Wistar Rats (Reuzel and Kuper, 1982)
- Rationale for animal assignment: random

Positive control:

none

Parental animals: Observations and examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: at least once a day

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: weekly, and for the sperm positive females on day 1, 7, 14 and 21 of lactation, body weights of maternal animals were recorded on Days 1, 4, 7, 21, and 28 postpartum

Oestrous cyclicity (parental animals):

no information

Sperm parameters (parental animals):

no information

Litter observations:

STANDARDISATION OF LITTERS
- Performed on day 4 postpartum: yes
- 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 / F2 offspring:
number and sex of pups, stillbirths, live births, postnatal mortality, presence of gross anomalies, weight gain, physical or behavioural abnormalities,

GROSS EXAMINATION OF DEAD PUPS:
yes, for external and internal abnormalities; possible cause of death was determined for pups born or found dead

Postmortem examinations (parental animals):

SACRIFICE
- Male animals: All surviving animals as soon as the last litter of F1 or F2 pups were weaned
- Maternal animals: All surviving animals as soon as the last litter of F1 or F2 pups were weaned

GROSS NECROPSY
- Gross necropsy consisted of external and internal examinations including the cervical, thoracic, and abdominal viscera.

HISTOPATHOLOGY / ORGAN WEIGHTS: yes

Postmortem examinations (offspring):

SACRIFICE
- The F1 offspring not selected as parental animals and all F2 offspring were sacrificed at 28 days of age.

Statistics:

Body weights were evaluated by Barlett's test and ANOVA, Littersize by ANOVA,
Mating and conception indices by Fisher Exact Probability test, Evaluation of neonatal sex ratio by Binomial distribution test, Survival indices by censored Wilcoxon test

Reproductive indices:

Mating index: number of females with a vaginal plug or sperm positive vaginal smear expressed as percentage of the total number of mated females
Conception index: number of females delivering a litter expressed as a percentage of the number of mated (sperm positive) females.
Gestation index: Number of females delivering a live litter expressed as a percentage of the number of females delivering a litter.

Offspring viability indices:

gestation survival index: percentage of newborn pups that were alive at birth.
1-28 day survival index: percentage of liveborn pups that survived for 1-28 days.

Clinical signs:

no effects observed

Description (incidence and severity):

No treatment-related alterations in demeanor or physical appearance were observed.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

One P0 male rat in the 30 ppm group was found dead after exposure on Day 72 of the study. A second P0 male in this group was found dead on Day 166. The findings on these animals did not indicate a treatment-related effect.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Decreases in body weight were observed in all 3 male exposure groups from week 17 until sacrifice. There was also an decrease in bodyweight in the 300 ppm females beginning after week 1.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not specified

Clinical biochemistry findings:

not specified

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not specified

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Histopathological findings: neoplastic:

not examined

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

effects observed, non-treatment-related

Description (incidence and severity):

Mating indices: 90, 87, 97 and 100% respectively in the 0, 30, 100 and 300 ppm groups,
Conception indices: 81, 85, 59 and 70 % respectively in the 0, 30, 100 and 300 ppm groups
Gestation survival index: was 99.6 - 100%
The conception index in the 100 ppm group was lower than the control (59% vs
81%) though the difference was not significant. However, the conception index in the 300 ppm group was 70% and thus there was no apparent dose-response.

Key result

Dose descriptor:

NOAEC

Remarks:

systemic toxicity

Effect level:

100 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

Key result

Dose descriptor:

NOAEC

Remarks:

fertility

Effect level:

> 300 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No effects on fertility were observed at the highest dose. 300 ppm is considered equivalent to 710 mg/m3.

Key result

Critical effects observed:

no

Clinical signs:

no effects observed

Mortality:

no mortality observed

Description (incidence):

No deaths occurred during treatment.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

A significant decrease in body weight was observed in males and females exposed to 300 ppm beginning at Week 3 and continuing until the end of the study. Body weights of the males exposed to 100 ppm were also lower, though
the difference was significant only at Week10. Significant decreases in body weight were observed in males exposed to 30 ppm beginning at Week 5 and continuing throughout the rest of the study.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Endocrine findings:

not examined

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, non-treatment-related

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Histopathological findings: neoplastic:

not examined

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

effects observed, non-treatment-related

Description (incidence and severity):

Mating indices: 87, 93, 93 and 100% respectively in the 0, 30, 100 and 300 ppm groups,
Conception indices: 77, 86, 82 and 93 % respectively in the 0, 30, 100 and 300 ppm groups
Gestation survival index: was 98.6 - 100%

Key result

Dose descriptor:

NOAEC

Remarks:

systemic toxicity

Effect level:

100 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

Key result

Dose descriptor:

NOAEC

Remarks:

fertility

Effect level:

> 300 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No effects on fertility were observed at the highest dose. 300 ppm is considered equivalent to 710 mg/m3.

Key result

Critical effects observed:

no

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, non-treatment-related

Description (incidence and severity):

The litter size on day 1 was lower in the 100 ppm group than the control value: 7.5 vs. 10.5. However, the litter size in the 300 ppm group was comparable to control group.
The survival indices on days 1, 4, 7, 14, 21 and 28 for the 3 exposure ranged from 94 to 100 %

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

No significant differences were observed in mean pup body weights on days 1, 4, 7, 21, or 28 postpartum.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not specified

Anogenital distance (AGD):

not specified

Nipple retention in male pups:

not specified

Organ weight findings including organ / body weight ratios:

not specified

Gross pathological findings:

effects observed, non-treatment-related

Histopathological findings:

effects observed, non-treatment-related

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

Key result

Dose descriptor:

NOAEC

Generation:

F1

Effect level:

> 300 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No adverse effects observed at highest dose.

Key result

Critical effects observed:

no

Clinical signs:

no effects observed

Mortality / viability:

mortality observed, non-treatment-related

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

No significant differences were observed in mean pup body weights on days 1, 4, 7, 21, or 28 postpartum.

Food consumption and compound intake (if feeding study):

not examined

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Urinalysis findings:

not examined

Sexual maturation:

not specified

Anogenital distance (AGD):

not specified

Nipple retention in male pups:

not specified

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

effects observed, non-treatment-related

Histopathological findings:

effects observed, non-treatment-related

Behaviour (functional findings):

not examined

Developmental immunotoxicity:

not examined

Key result

Dose descriptor:

NOAEC

Generation:

F2

Effect level:

> 300 ppm (nominal)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No adverse effects observed at highest dose.

Key result

Critical effects observed:

no

Key result

Reproductive effects observed:

no

Effect on fertility: via oral route

Endpoint conclusion:

no study available

Effect on fertility: via inhalation route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

300 ppm

Species:

rat

Quality of whole database:

similar to OECD 416 (read across)

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

No data for reproductive toxicity of 1,2-epoxybutane are available. No indications of reproductive toxicity were seen in repeated dose toxicity studies, in the dominant lethal test in vivo and in developmental toxicity studies. However, reproduction toxicity data are available for the structural analogue substance propylene oxide (CAS 75-56-9), which are used as read-across. Please refer to section 13 for Read-Across Justification.

In this two-generation reproduction study similar to OECD TG 416, groups of 30 male and 30 female F344 rats were exposed to 0, 30, 100, 300 ppm (0, 70, 240 and 710 mg/m3) propylene oxide vapour. Exposure was performed for 6 hours/day, 5 days/week for 14 weeks prior to mating (whole body). After weaning, 30 F1 pups/sex/group were similarly exposed to propylene oxide for 17 weeks and then mated to produce F2. Reproductive parameters examined included fertility, litter size, neonatal growth and survival. All adults and weanlings were examined for gross and microscopic lesions. No deaths occurred, and there were no treatment-related alterations in demeanour or physical appearance in any of the animals during the pre-mating periods. Toxicity was evident as reduced body weight gain in F0 (8%) and F1 (16%) rats at 300 ppm, however there was no evidence of treatment-related adverse effect on fertility in F0 or F1 matings. Growth and survival of F1 and F2 offspring was not adversely affected by exposure of either generation of parents at any dose. Mating and conception were not significantly affected in either F0 or F1 matings. Litter size was not adversely affected. Detailed pathology examination of adults and weanlings revealed no changes considered attributable to propylene oxide exposure. The results indicate that inhalation exposure to levels up to 300 ppm over two generations did not produce any adverse effect on reproductive function (Dow, 1985).

Effects on developmental toxicity

Description of key information

The available studies on developmental toxicity for 1,2 -epoxybutane (rat and rabbit) revealed no teratogenicity.

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

GLP compliance:

not specified

Limit test:

no

Species:

rat

Strain:

Wistar

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Hilltop Lab Animals, Inc. , Scottdale, PA
- Age at study initiation: 4-5 weeks (females), 8 weeks (males)
- Weight at study initiation: 100-125 g (females), 300-324 g (males)
- Fasting period before study:
- Housing: The animals were housed in stainless steel cages before, during, and after inhalation exposure. Caging of experimental females was individual within the exposure chamber except during the mating period. Cage assignment was not random and the locations were not rotated.
- Diet: Wayne Lab-Blox was provided ad libitum, except during the daily exposure period when the food was removed.
- Water: Water was supplied by an automatic watering system which was disconnected and drained prior to initiation of each exposure
- Acclimation period: ca 4 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): not controlled
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

other: room air

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
Rats were exposed in stainless steel chambers. A polydispersed particulate aerosol attained equilibirum in 15 - 20 min after introduction into the chamber. Concentrations measured above the six catch pans showed a 3 - 5% variation. The design of these chambers allows for up to 192 adult rats to be both exposed and housed within each chamber.
Three chambers were employed: one for the high-level, one for the low-level, and one for the pregestational filtered air exposure of rats. The same chambers were used for gestational exposure of rats. Animals were transferred to washed and sterilized chamber-caging units at least once a week.
HPE filtered air was continually suppled to the exposure chambers and the vapours under study were introduced into the filtered air streams. Chamber air flows were maintained at 10 cfm, which provided seven air changes per hour.
Methodology for generating the desired butylene oxide atmospheres was developed using an exposure chamber set up in the aerosol physic laboratory. Liquid butylene oxide was placed in a 3 -necked, 500 -ml distillation flask with a coarse fritted glass bubbler extending from the center neck into the liquid. One of the remaining necks was stoppered while the other had a tube extending into the inlet air line to the exposure chamber. Dry N2 gas was used at 0.225 -1.5 liters per minute to avoid explosive vapor concentrations in the generator. The liquid bath was heated to the temperatures necessary to generate the required amount of vapor. For the first three weeks of exposure, the butylene oxide was periodically replenished; subsequently it was replaced daily.
The generators were located in a fume hood, and connected to the exposure chambers by 3 -inch diameter flexible hose. The chamber was connected to a transvector air flow amplifier which introduced the exhaust air to another flexible hose which was connected to a room exhaust port. The total flow into the room exhaust hose was approx. 125 cfm of which only 10 cfm came from the chamber. To pump 10 cfm required about 3 cfm of drining air for a total output of 13 cfm. The system was adjusted to provide vapor concentrations in the chambers of 1000 and 250 ppm of butylene oxide.

Analytical verification of doses or concentrations:

yes

Remarks:

gas chromatography

Details on analytical verification of doses or concentrations:

Concentrations within the chamber were uniform to within 10% and remained within this range over prolonged generation periods.

Details on mating procedure:

Pregestational exposures ordinarily began on a Monday and were continued for 7 hours per day, 5 days a week, for 3 weeks. The females were transferred to a standard rack united and caged with males (2:1). Vaginal lavages were performed the following mornings and examined for the presence of sperm. Sperm-positive females were randomly assigned to gestational exposure groups. Gestational exposures were started on the day on which sperm were detected which was denoted as day 1 of gestation (d.g. 1). Mating and initiation of gestational exposures continued for 7 - 9 days, until about 36 sperm-positive rats were assigned to each experimental group. Gestational exposures were performed 8 hours per day, 7 days per week, through d.g. 19. All exposed mated rats were sacrificed at d.g. 21.
Based upon the combination of pregestational and gestational exposures, 1 control and 6 experimental groups were formed. These groups will be identified by the pregestational and gestational exposure in the presentation of results:
Air-air (control) - 3-week pregestational exposure to filtered air followed by exposure to filtered air during days 1 - 19 of gestation. Air-low - 3-week pregestational exposure to filtered air followed by low level exposure during days 1-19 of gestation. Air-high - 3-week pregestational exposure to filtered air followed by high level exposure during days 1-19 of gestation. Low-air - 3-week pregestational low level exposure followed by exposure to
filtered air during days 1-19 of gestation. Low-low - 3-week pregestational low level exposure followed by low level exposure during days 1-19 of gestation. High-air - 3-week pregestational high level exposure followed by exposure to filtered air during days 1-19 of gestation. High-high - 3-week pregestational high level exposure followed by high level exposure during days 1-19 of gestation.

Duration of treatment / exposure:

day 1 - 19 of gestation, additionally groups with pregestational exposure of 21 days

Frequency of treatment:

7 hours/day, 5 days/week

Duration of test:

until day 21 (cesarian section)

Dose / conc.:

250 ppm (nominal)

Remarks:

0.75 mg/mL (nominal conc.)

Dose / conc.:

1 000 ppm (nominal)

Remarks:

3.0 mg/mL (nominal conc.)

No. of animals per sex per dose:

total: 380

Control animals:

yes

Maternal examinations:

CAGE SIDE OBSERVATIONS: not specified

BODY WEIGHT: Yes
- Time schedule for examinations: The female rats were weighted at least twice weekly during pregestation exposure and on days 1, 7, 14 and 21 of gestation.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes

WATER CONSUMPTION AND COMPOUND INTAKE: No

POST-MORTEM EXAMINATIONS: Yes
The animals were sacrificed by introduction of carbon dioxide into an euthanisa chamber. Necropsy were performed on all animals; liver, lung, and kidneys were weighed. Internal abnormalities of the pregnant and non-pregnant animals were also recorded. Samples of the ovaries, uterus, liver, lungs with trachea, and kidneys were perserved in 10% neutral buffered formalin. Histopathological examination was performed on tissues from 25% of the pregnant animals, selected at random. The residual tissues and the tissues from the remaining 75% of the animals and from the non-pregnant animals have been preserved for possible future examination.

Ovaries and uterine content:

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

Blood sampling:

not specified

Fetal examinations:

- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [half per litter]
- Skeletal examinations: Yes: [all per litter ]
- Head examinations: Yes: [half per litter]
- Anogenital distance of all live rodent pups: no

Statistics:

Fisher's Exact Probability Test was used on the maternal and fetal data; Bonferronni's method was used to adjust for the problem associated with multiple comparisons against a control group. In all instances, differences from the control group at probability levels of 0.05 or less were accepted as statistically significant.

Indices:

no data

Historical control data:

no data

Clinical signs:

not specified

Mortality:

mortality observed, non-treatment-related

Description (incidence):

No mortality without pregestational exposure (0/38). High pregestational exposure caused one death (1/42). However, this death was not regarded as substance-related (the surviving animals showed no signs of severe toxicity; no further details are given in the report).

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

Pregestational exposure to 1000 ppm of butylene oxide produced a slight, but statistically significant, reduction in the body weight of the rats relative to the controls at most time periods. The differences were transient and were not statistically significant at the end of the pregestation exposure. By 7 days of gestational exposure, the rats exposed at the high levels were significantly lighter than the controls, and remained lighter throughout the study.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

No remarkable changes in food consumption were produced by pregestational or gestational exposure of rats.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Endocrine findings:

not specified

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

The relative organ weight of liver, kidney and lung were not significantly changed compared to control. No statistically significant differences in placental weight were detected.

Gross pathological findings:

not specified

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

Lung: A variety of pulmonary lesions, generally classified as inflammations, were frequently seen in the rats. In nine rats there were focal alveolar inflammations (accumulations of lymphocytes and plasma cells) which were not directly associated with arterioles or bronchioles, and small granulomas were observed in 2 rats. Other changes in the lungs were seen in all experiments, but did not appear to be related to exposure. These include alveolar histiocytosis, a focal accumulation of large histiocytes in alveoli. Most rats showed slight mononuclear peribronchiolitis and perivasculitis-lymphoid nodules and plasma cells around the bronchioles and associated arterioles. These lesions are common for rats, and may reflect previous exposure to pathogenic organisms.
Kidney: Several rats of each dose group were considered to have interstitial nephritis. This diagnosis was used when any lymphocytes or plasma cells were seen in the interstitial tissues. These infiltrates were very minimal and considered normal. Evidence of glomerulonephrosis was not detected.
Uterus: Slight to mild mononuclear endometritis, i.e. any infiltrate of lymphocytes and/or plasma cells in the endometrium, was a consistent observation. These changes were, for the most part, considered within the normal range, particularly since all rats were pregnant. Apparently normal corpora lutea were noted in the sections of ovary from each rat.

Histopathological findings: neoplastic:

not examined

Number of abortions:

no effects observed

Pre- and post-implantation loss:

effects observed, non-treatment-related

Total litter losses by resorption:

effects observed, non-treatment-related

Early or late resorptions:

effects observed, non-treatment-related

Dead fetuses:

no effects observed

Changes in pregnancy duration:

not examined

Changes in number of pregnant:

effects observed, non-treatment-related

Description (incidence and severity):

The exposure led to a slightly reduced percentage of sperm-positive rats which were pregnant (36/37, 33/37, 28/35, 33/42, 38/44, 33/40, 31/39 for air-air, air-low, air-high, low air, low-low, high-air and high-high respectively) although no clear dose relationship was seen and the decrease was not statistically significant.

Key result

Dose descriptor:

NOAEC

Effect level:

1 000 ppm

Based on:

test mat.

Basis for effect level:

other: No adverse effect observed up to highest dose.

Key result

Abnormalities:

effects observed, non-treatment-related

Fetal body weight changes:

effects observed, non-treatment-related

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

effects observed, non-treatment-related

Description (incidence and severity):

The exposure had no statistically significant effect on either the weight or length of the fetuses in rats.

Anogenital distance of all rodent fetuses:

not examined

Changes in postnatal survival:

effects observed, non-treatment-related

External malformations:

effects observed, non-treatment-related

Skeletal malformations:

effects observed, non-treatment-related

Visceral malformations:

effects observed, non-treatment-related

Key result

Dose descriptor:

NOAEC

Effect level:

> 1 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No teratogenicity observed up to highest dose tested.

Key result

Abnormalities:

effects observed, non-treatment-related

Key result

Developmental effects observed:

no

Reference 2

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

GLP compliance:

not specified

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Scotts Rabbitry, Vashon Island, Washington
- Age at study initiation: 4-5 months (females), 6 months (males)
- Weight at study initiation: ca. 3 kg
- Fasting period before study: no
- Housing: caged in individual stainless steel cages
- Diet, Wayne Rabbit Diet ad libitum, except during exposure
- Water: ad libitum, except during exposure
- Acclimation period: min. 18 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): no data
- Humidity (%): not controlled
- Air changes (per hr): no data
- Photoperiod (hrs dark / hrs light): 12/12

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

other: room air

Details on exposure:

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
Rabbits were exposed in stainless steel chambers. A polydispersed particulate aerosol attained equilibirum in 15 - 20 min after introduction into the chamber. Concentrations measured above the six catch pans showed a 3 - 5% variation.
Three chambers were employed: one for the high-level, one for the low-level, and one for filtered air exposure.
HPE filtered air was continually suppled to the exposure chambers and the vapours under study were introduced into the filtered air streams. Chamber air flows were maintained at 10 cfm, which provided seven air changes per hour.
Methodology for generating the desired butylene oxide atmospheres was developed using an exposure chamber set up in the aerosol physic laboratory. Liquid butylene oxide was placed in a 3 -necked, 500 -ml distillation flask with a coarse fritted glass bubbler extending from the center neck into the liquid. One of the remaining necks was stoppered while the other had a tube extending into the inlet air line to the exposure chamber. Dry N2 gas was used at 0.225 -1.5 liters per minute to avoid explosive vapor concentrations in the generator. The liquid bath was heated to the temperatures necessary to generate the required amount of vapor. For the first three weeks of exposure, the butylene oxide was periodically replenished; subsequently it was replaced daily.
The generators were located in a fume hood, and connected to the exposure chambers by 3 -inch diameter flexible hose. The chamber was connected to a transvector air flow amplifier which introduced the exhaust air to another flexible hose which was connected to a room exhaust port. The total flow into the room exhaust hose was approx. 125 cfm of which only 10 cfm came from the chamber. To pump 10 cfm required about 3 cfm of drining air for a total output of 13 cfm. The system was adjusted to provide vapor concentrations in the chambers of 1000 and 250 ppm of butylene oxide.

Analytical verification of doses or concentrations:

yes

Remarks:

gas chromatography

Details on analytical verification of doses or concentrations:

Concentrations within the chamber were uniform to within 10% and remained within this range over prolonged generation periods.

Details on mating procedure:

The rabbits were artificially inseminated over the course of three successive days, using pooled semen samples (Hafez, 1970; Andrew and Staples, 1977) from bucks which had been previously trained to serve an artificial vagina (AV). Each AV was filled with water at about 45°C before use, and semen was collected into the external reservoir at room temperature. The pooled semen samples from three males were diluted with a buffered extender to a concentration of approx. 0.5 mL was used to inseminate each doe within 1 - 2 hours of semen collection. Ovulation was induced by nearly simultaneous i.f. injections of 2.5 mg of pituitary luteinizing hormone. The morning following insemination was defined as GD1.

Duration of treatment / exposure:

day 1 - 24 of gestation

Frequency of treatment:

7 hours/day, 5 days/week

Duration of test:

until day 30 (cesarian section)

Dose / conc.:

250 ppm (nominal)

Remarks:

0.75 mg/L (nominal)

Dose / conc.:

1 000 ppm (nominal)

Remarks:

3.0 mg/L (nominal)

No. of animals per sex per dose:

24

Control animals:

yes, concurrent no treatment

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes

BODY WEIGHT: Yes
The adult rabbits were weighted before exposure, prior to the daily exposures on days 1, 9, 16 and 23 of gestation, and prior to sacrifice at day 30.

FOOD CONSUMPTION AND COMPOUND INTAKE: Yes
Food consumption was measured before the exposure period began and every two days throughout exposure.

WATER CONSUMPTION AND COMPOUND INTAKE : Yes

POST-MORTEM EXAMINATIONS: Yes

Ovaries and uterine content:

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

Fetal examinations:

- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [all per litter]
- Skeletal examinations: Yes: [half per litter]
- Head examinations: Yes: [half per litter]

Statistics:

Fisher's Exact Probability Test was used on the maternal and fetal data. Bonferronni's method was used to adjust for the problem associated with multiple comparisons against a control group. In all instances, differences from the control group at probability levels of 0.05 or less were accepted as statistically significant.

Indices:

no data

Historical control data:

no data

Clinical signs:

not specified

Mortality:

mortality observed, treatment-related

Description (incidence):

The mortality incidences of 1 of 23 and 5 of 25 rabbits exposed to 250 ppm were not significantly different from each other. Fourteen of 24 (58%) exposed to 1000 ppm died during exposure to butylene oxide; a significant increase. Supperative pneumonia was the usual necropsy finding in rabbits which died during exposure. Positive cultures for Pasturella multocida were obtained from the lungs of many of these ani- mals.it therefore appears that the development of overt infection was related to the butylene oxide exposure.

Body weight and weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

Body weight was not significantly affected by exposure.

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Description (incidence and severity):

Adequate data for butylene oxide exposed rabbits are available only for the low level, where no effect was detected. Incomplete measurements in a first experiment, in which animals have been exposed to 250 ppm, confirm this lack of effect at the low dose but suggest that there may have been an effect at the high level.

Food efficiency:

not examined

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

not examined

Ophthalmological findings:

not examined

Haematological findings:

not examined

Clinical biochemistry findings:

not examined

Endocrine findings:

not specified

Urinalysis findings:

not examined

Behaviour (functional findings):

not examined

Immunological findings:

not examined

Organ weight findings including organ / body weight ratios:

effects observed, non-treatment-related

Description (incidence and severity):

The exposure did not have a major effect on lung weights; kidney weights were unaffected. No statistically significant differences in placental weight were detected.

Gross pathological findings:

not specified

Neuropathological findings:

not examined

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

Lungs: A variety of inflammatory lesions were also seen in the rabbits and may have been influenced by an endemic Pasteurella infection.

Histopathological findings: neoplastic:

not examined

Details on results:

Gestational exposure did not have statistically significant effects on any of the measures of reproductive success.

Number of abortions:

not specified

Pre- and post-implantation loss:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment related effects at dose level of 250 ppm. At highest dose number of examined does/litter was to small for statistically evaluations due to high mortality.

Total litter losses by resorption:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment related effects at dose level of 250 ppm. At highest dose number of examined does/litter was to small for statistically evaluations due to high mortality.

Early or late resorptions:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment related effects at dose level of 250 ppm. At highest dose number of examined does/litter was to small for statistically evaluations due to high mortality.

Dead fetuses:

effects observed, non-treatment-related

Description (incidence and severity):

No treatment related effects at dose level of 250 ppm. At highest dose number of examined does/litter was to small for statistically evaluations due to high mortality.

Changes in pregnancy duration:

not examined

Changes in number of pregnant:

effects observed, non-treatment-related

Description (incidence and severity):

The exposure led to a reduced percentage of sperm-positive rabbits, based on the results at scheduled sacrifice. Although this might be considered to be influenced by preimplantation mortality of does, nine of 13 does in the high group which died prior to scheduled sacrifice were pregnant. This gives an overall fraction of 11/23 or 48%, as compared to the 42% pregnant in the air group.

Key result

Dose descriptor:

NOAEC

Effect level:

250 ppm

Based on:

test mat.

Basis for effect level:

mortality

Key result

Abnormalities:

effects observed, treatment-related

Fetal body weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

The exposure had no statistically significant effect on either the weight or length of the fetuses. The fetuses of the few (2 and 1, respectively) surviving litters of high dose rabbits were markedly smaller than were those of the control or low dose groups.

Reduction in number of live offspring:

effects observed, non-treatment-related

Changes in sex ratio:

no effects observed

Description (incidence and severity):

Statistically significant differences in the sex ratios were not observed.

Changes in litter size and weights:

effects observed, non-treatment-related

Changes in postnatal survival:

not examined

External malformations:

effects observed, non-treatment-related

Skeletal malformations:

effects observed, non-treatment-related

Visceral malformations:

effects observed, non-treatment-related

Details on embryotoxic / teratogenic effects:

There were only two litters in the high dose group available for examination. In the two litters exposed to the high level (for which statistical analysis could not be performed) there was a suggestion of a decreased number of live fetuses per litter and an increase in the frequency of resorptions (6.7, 7.0, 4.0 live fetuses per litter in the control, low and high dose group, respectively and 0.85, 0.47, 2.5 resorptions per litter in the control, low and high dose group, respectively). Alterations of the nature or incidence of morphologic changes related to exposure were not noted. The authors concluded: The few detectable embryotoxic effects of butylene oxide occurred in the rabbit at the concentration which was maternally lethal.

Key result

Dose descriptor:

NOAEC

Effect level:

> 1 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: No teratogenicity observed up to highest dose tested.

Key result

Abnormalities:

effects observed, non-treatment-related

Key result

Developmental effects observed:

no

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no study available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

1 000 ppm

Study duration:

subacute

Species:

rat

Quality of whole database:

similar to OECD TG 414

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

A study similar to OECD TG 414 in rats is available. Wistar rats have been exposed from day 1 to 19 of gestation or in additional groups with pregestational expose of 21 days to test item vapours (whole body). Animals were exposed to concentrations of 250 and 1000 ppm (i.e, 0.75, 3.0 mg/L air nominal) for 7 h/day for 5 day/week. A negative control group received air only. Pregestational exposure to 1000 ppm of butylene oxide produced a slight, but statistically significant, reduction in the body weight of the rats relative to the controls at most time periods. The differences were transient and were not statistically significant at the end of the pregestation exposure. No mortality was observed in the group without pregestational exposure. High pregestational exposure caused one death. However, this death was not regarded as substance-related. No remarkable changes in food consumption were produced by pregestational or gestational exposure of rats. The exposure did not adversely affect liver weight or lung weight. No statistically significant differences in placental weight were detected. Apparently normal corpora lutea were noted in the sections of ovary from each rat. The exposure led to a slightly reduced percentage of sperm-positive rats which were pregnant although no clear dose relationship was seen and the decrease was not statistically significant. The exposure had no statistically significant effect on either the weight or length of the fetuses in rats. Statistically significant differences in the sex ratios were not observed. Pregestational and/or gestational exposure did not have statistically significant effects on any of the measures of reproductive success. Alterations of the nature or incidence of morphologic changes related to exposure were not noted. The NOAEC for maternal toxicity was 1000 ppm and the NOAEC for developmental toxicity and teratogenicity also was 1000 ppm (NIOSH,1981).

A further study similar to OECD TG 414 in rabbits is available. New Zealand White rabbits have been exposed from day 1 to 24 of gestation to test item vapours at concentrations of 250 and 1000 ppm (i.e, 0.75, 3.0 mg/L air nominal) for 7 h/day for 5 days/week. A negative control group received air only. Fourteen of 24 (58%) exposed to 1000 ppm died during exposure; a significant increase. Supperative pneumonia was the usual necropsy finding in rabbits which died during exposure. The exposure did not have a major effect on lung weight and kidney weights were unaffected. No statistically significant differences in placental weight were detected. The exposure led to a reduced percentage of sperm-positive rabbits, based on the results at scheduled sacrifice. Although this might be considered to be influenced by preimplantation mortality of does, nine of 13 does in the high group which died prior to scheduled sacrifice were pregnant. This gives an overall fraction of 11/23 or 48%, as compared to the 42% pregnant in the air group. The exposure had no statistically significant effect on either the weight or length of the fetuses. The fetuses of the few (2 and 1, respectively) surviving litters of high dose rabbits were markedly smaller than were those of the control or low dose groups. Statistically significant differences in the sex ratios were not observed. The NOAEC for maternal toxicity was found to be 250 ppm and the NOAEC for teratogenicity was 1000 ppm (NIOSH, 1981).

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on available data the test item is not classified for toxicity to reproduction according to Regulation (EC) No 1272/2008 (CLP).