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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1987
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
comparable to guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1987
Report Date:
1987

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
GLP compliance:
yes
Limit test:
no

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): Hexyl CELLOSOLVE Vapor (ethylene glycol monohexyl ether)
- Physical state: liquid
- Analytical purity: 99%
- Impurities (identity and concentrations): refer to table 1
- Composition of test material, percentage of components: 99% Hexyl CELLOSOLVE Vapor (ethylene glycol monohexyl ether)
- Isomers composition: not specified in the report
- Purity test date: not specified in the report
- Lot/batch No.: Lot No. S074565
- Expiration date of the lot/batch: not specified in the report
- Stability under test conditions: stable under test conditions
- Storage condition of test material: not specified in the report

Test animals

Species:
rat
Strain:
Fischer 344
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories Inc.
- Age at study receipt: males (69 days upon arrival) and females (62 days upon arrival)
- Weight at study initiation: not specified in the report
- Fasting period before study: none
- Housing: mating - (1:1, male:female), post-mating - females housed individually
- Diet (e.g. ad libitum): Certified Ground Rodent Chow provided ad libitum, except during exposure
- Water (e.g. ad libitum): Municipal water provided ad libitum, except during exposure
- Acclimation period: Animals were quarantined for two weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23-28 °C
- Humidity (%): 50-61 %
- Air changes (per hr): standard conditions
- Photoperiod (hrs dark / hrs light): 12 hours (light:dark cycle)

Administration / exposure

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
- Exposure apparatus: 4 chambers made of glass and stainless steel
- Method of holding animals in test chamber: in cages
- Source and rate of air: ambient
- Method of conditioning air: not specified in the report
- Temperature, humidity, pressure in air chamber: mean temperatures - 25.1-26.5 °C, mean RH - 52.9-56.5%
- Air flow rate: 1000 liters/minute
- Air change rate: 14 air changes/hour
- Treatment of exhaust air: Chamber atmospheres containing hexyl CELLOSOLVE were filtered before leaving an exhaust stack

TEST ATMOSPHERE
- Brief description of analytical method used: A Perkin-Elmer Model 3920B gas chromatograph (GC) equipped with a flame ionization detector was used to monitor the hexyl CELLOSOLVE vapor concentrations in the chambers. The GC column was a 10-feet x 1/8 inch (i.d.) stainless steel column packed with 20% SP-2100 on 80/100 mesh Supelcoport (Supelco, Inc., Bellefonte, PA). Calibration of the gas chromatograph was done with dynamically generated gas standards of hexyl CELL0S0LVE prepared by syringe injection of test material into Tedlar (DuPont) gas bags. The series of standards encompassed the entire range of vapor concentrations generated in the exposure chambers. A linear calibration curve was obtained when areas (integration counts) were plotted versus the gas equivalent concentrations of the standards. Each chamber atmosphere was analyzed for hexyl CELLOSOLVE approximately once every 30 minutes during each 6-hour exposure. Daily nominal concentrations (an estimated concentration calculated from the amount of test material delivered and the chamber airflow during the exposure period) were also calculated for each chamber.
- Samples taken from breathing zone: not specified in the report
- Liquid hexyl CELLOSOLVE® was metered from a piston pump into a heated glass evaporator. The temperature in the evaporator was maintained at the lowest level sufficient to vaporize the liquid (33-52°C). The resultant vapor was carried into the chamber by passage of conditioned air through the evaporator. Chamber atmospheres containing hexyl CELLOSOLVE were filtered before leaving an exhaust stack.
The four chambers employed in this study were rectangular in shape, constructed of glass and stainless steel (Wahmann Manufacturing Company, Timonium, MD), and measured approximately 2.1 m x 1 m x 2.1 m (height). Total volume in each chamber was approximately 4320 L. An orifice plate was positioned in the exhaust duct of the chamber and was connected to a Dwyer Magnehelic Pressure Gauge.
Airflow in each chamber was approximately 1000 L/minute (14 air changes per hour) with a t99 (theoretically-derived time required for the chamber to reach 99% of the equilibrium concentration) of approximately 20 minutes.
The chambers were illuminated with artificial room light. Chamber temperature, relative humidity and air flow rate were recorded at least five (5) times during each exposure. Within each chamber, the animal cages were rotated daily to compensate for any possible, but undetected, variation in chamber exposure conditions.

Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
target concentrations - 0, 20, 40 and 85 ppm
analytical concentrations - 0, 20.8 ± 0.90, 41.1 ± 1.77 and 79.2 ± 10.80 ppm
nominal concentrations - 0, 26.1 ± 1.84, 39.2 ± 1.80 and 91.0 ± 6.28 ppm
Details on mating procedure:
- Impregnation procedure: [cohoused]
- If cohoused:
- M/F ratio per cage: 1:1 (male:female)
- Length of cohabitation: 5 days
- Further matings after two unsuccessful attempts: [no]
- Verification of same strain and source of both sexes: [no]
- Proof of pregnancy: [vaginal plug] referred to as [day 0] of pregnancy
Duration of treatment / exposure:
6 hours/day, gd 6 to gd 15
Frequency of treatment:
daily from gd 6 to gd 15
Duration of test:
9 days
Doses / concentrationsopen allclose all
Dose / conc.:
0 ppm
Remarks:
analytical concentrations - 0 ppm, nominal concentrations - 0 ppm
Dose / conc.:
20 ppm
Remarks:
analytical concentrations - 20.8 ± 0.90 ppm, nominal concentrations - 26.1 ± 1.84 ppm
Dose / conc.:
40 ppm
Remarks:
analytical concentrations - 41.1 ± 1.77 ppm, nominal concentrations - 39.2 ± 1.80 ppm
Dose / conc.:
85 ppm
Remarks:
analytical concentrations - 79.2 ± 10.80 ppm, nominal concentrations - 91.0 ± 6.28 ppm
No. of animals per sex per dose:
25 plug positive females/group
Control animals:
yes, sham-exposed
Details on study design:
- Dose selection rationale: based on range-fiding study
- Rationale for animal assignment: random assignment

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily from gd 0 to gd 21

DETAILED CLINICAL OBSERVATIONS: No data

BODY WEIGHT: Yes
- Time schedule for examinations: gd 0, 6, 9, 12, 15 and 21

FOOD CONSUMPTION AND WATER CONSUMPTION: Yes
- Time schedule for examinations: at intervals gd 0-3, 3-6, 6-9, 9-12, 12-15, 15-18 and 18-21

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 21
- Organs examined: all organs

OTHER: Hematological 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 resorptions: Yes
- Number of late resorptions: Yes
- Other: percent live and dead fetuses
Fetal examinations:
- External examinations: Yes: [all per litter]
- Soft tissue examinations: Yes: [half per litter]
- Skeletal examinations: Yes: [half per litter]
- Head examinations: Yes: [half per litter] - fetuses examined for soft-tissue examinations
Statistics:
The unit of comparison was the pregnant female or the litter. Results of the quantitative continuous variables (e.g., maternal body weights, liver weights, etc.) were intercompared for the three exposure groups and a control group for each species by use of Levene's test for equal variances, analysis of variance (ANOVA), and t-tests with Bonferroni probabilities. The t-tests were used when the F value from the ANOVA was significant. When Levene's test indicated homogeneous variances, and the ANOVA was significant, the pooled t-test was used. When Levene's test indicated heterogeneous variances, all groups were compared by an ANOVA for unequal variances followed, when necessary, by the separate variance t-test.
Nonparametric data obtained following laparohysterectomy were statistically treated using the Kruskal-Wallis test followed by the Mann-Whitney U test when appropriate. Incidence data were compared using Fisher's Exact Test. For all statistical tests, the fiducial limit of 0.05 (two-tailed) was used as the criterion for significance.
Indices:
Pre-and post-implantation loss, percent live fetuses, sex ratio
Historical control data:
not applicable

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Treatment related clinical signs of toxicity which included urine stains on fur, lacrimation, ocular wetness and encrustation were observed only at 85 ppm during the exposure period.
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
There were significant decreases in maternal body weight and weight gain at both the 40 and 85 ppm dose group during the exposure period.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
There were significant decreases in food consumption at 85 ppm dose group during the exposure period.
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Description (incidence and severity):
Water consumption was increased at the 85 ppm dose group during the exposure period.
Haematological findings:
no effects observed
Description (incidence and severity):
There were no changes in the hematological examination
Organ weight findings including organ / body weight ratios:
no effects observed
Description (incidence and severity):
There were no changes on maternal organ weights, gravid uterine weight, gestational body weight and absolute/relative liver weight
Gross pathological findings:
no effects observed
Description (incidence and severity):
At scheduled necropsy, there were no treatment related findings at maternal gross examination.

Maternal developmental toxicity

Details on maternal toxic effects:

.

Effect levels (maternal animals)

Key result
Dose descriptor:
NOEC
Effect level:
20 ppm
Based on:
test mat.
Basis for effect level:
body weight and weight gain

Results (fetuses)

Fetal body weight changes:
no effects observed
Description (incidence and severity):
There were no effects of exposure on fetal body weight
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Description (incidence and severity):
There were no effects of exposure on sex ratio
Changes in litter size and weights:
no effects observed
External malformations:
no effects observed
Description (incidence and severity):
The incidence of fetal malformations and variations were comparable between the treatment groups.
Skeletal malformations:
no effects observed
Description (incidence and severity):
The incidence of fetal malformations and variations were comparable between the treatment groups.
Visceral malformations:
no effects observed
Description (incidence and severity):
The incidence of fetal malformations and variations were comparable between the treatment groups
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
There were no effects of exposure on gestational parameters including number of corpora lutea per dam, number of total, non-viable or live implantations per litter, pre- and post-implantation loss, sex ratio and fetal body weight. The incidence of fetal malformations and variations were comparable between the treatment groups.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEC
Effect level:
85 ppm
Sex:
male/female
Basis for effect level:
other: no adverse effects

Fetal abnormalities

Abnormalities:
no effects observed

Overall developmental toxicity

Key result
Developmental effects observed:
no

Applicant's summary and conclusion

Conclusions:
Exposure to ethylene glycol hexyl ether vapor during organogenesis in Fischer 344 rats resulted in transient maternal toxicity at 85 and 40 ppm, and no embryofetal toxicity (including teratogenicity) at any exposure concentrations employed. The "no observable effect level" (NOEL) for maternal toxicity was 20 ppm for rats and the developmental toxicity NOEL was at the ambient temperature generated vapor concentration of about 85 ppm.
Executive summary:

Timed-pregnant Fischer 344 rats, 25 per group, were exposed to ethylene glycol hexyl ether vapor for six hours/day on gestational days (gd) 6 through 15 for rats at target concentrations of 0, 20, 40 or 85 ppm (analytical concentrations of 20.8 ± 0.90, 41.1 ± 1.77 and 79.2 ± 10.80 ppm for the 20, 40 and 85 ppm groups, respectively). Eighty-five ppm was close to the saturated vapor concentration of the material at 20°C. Maternal body weights were measured on gd 0, 6, 9, 12, 15 and 21 (rats). Food and water consumption was measured, for rats only, for three-day intervals throughout gestation. At scheduled sacrifice on gd 21 (rats), maternal body weight, gravid uterine weight, and liver weight were taken. Maternal blood samples were taken examined for hematologic changes including differential leukocyte counts. Ovarian corpora lutea of pregnancy were counted and all uterine implantation sites were identified and recorded: resorptions (early or late), dead fetuses and live fetuses.All live fetuses were examined for external malformations, including cleft palate, and variations. About 50% of the rat fetuses in each litter were examined for visceral malformations and variations, 50% of the fetuses in each litter were examined for craniofacial defects and the other 50% (intact fetuses) were examined for skeletal malformations and variations.

In rats, maternal toxicity was observed at 40 and 85 ppm, including transient reductions in maternal body weight and weight gain during the exposure period and clinical signs of toxicity at 85 ppm, and reduced weight gain during the exposure period at 40 ppm. Maternal food consumption was reduced at 85 ppm during the exposure period (and increased in the postexposure period). Water consumption was increased at 85 ppm during and after the exposure period. There were no treatment-related clinical signs, gross necropsy observations or hematologic changes. Gestational parameters, including corpora lutea, total, nonviable or live implantations per litter, pre- or postimplantation loss, sex ratio or fetal body weights (males, females or total) per litter, were unaffected by exposures. There were no effects of treatment on the incidence of malformations or variations when examined by individual finding, findings by category or total findings.

Exposure to the test substance vapor during organogenesis in Fischer 344 rats resulted in transient maternal toxicity at 85 and 40 ppm, and no embryofetal toxicity (including teratogenicity) at any exposure concentrations employed. The "no observable effect level" (NOEL) for maternal toxicity was 20 ppm for rats and the NOEL for developmental toxicity was 85 ppm.