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Toxicity to reproduction

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Administrative data

Endpoint:
two-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:
key study
Study period:
1988
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Well documented publication and full study report that meets basic scientific principles. Rationale for using a read across substance is included in overall remarks section.

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Association of sperm, vaginal cytology and reproductive organ weight data with results of continuous breeding reproduction studies in Swiss CD1 Mice.
Author:
Morrissey RE , Lamb JC, Schwetz BA, Teague JL and Morris RW
Year:
1988
Bibliographic source:
Fundamental and Applied Toxicology, 11, p359-371.
Reference Type:
publication
Title:
Results and evaluations of 48 continuous breeding reproduction studies conducted in mice.
Author:
Morrissey RE , Lamb JC, Morris RW, Chapin RE, Gulati DK and Heindel JJ
Year:
1989
Bibliographic source:
Fundamental and Applied Toxicology, 13, p747-777.
Reference Type:
publication
Title:
Assessment of Ethylene Glycol Monobutyl and monophenol Ether reproductive toxicity using a continuous breeding protocol in Swiss CD-1 mice.
Author:
Heindel JJ , Gulati DK, Russel VS, Reel JR, Lawton AD and Lamb JC
Year:
1990
Bibliographic source:
Fundamental and Applied Toxicology, 15, p683-696.
Reference Type:
study report
Title:
Unnamed
Year:
1985
Report Date:
1985

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: National Toxicology Programme Continuous Breeding Protocol
Deviations:
yes
Remarks:
, F1 tests were only carried out with the control and low dose groups
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): ethylene glycol butyl ether
- Analytical purity: >99% (confirmed by GC)
- Supplier: Midwest research inst, Kansas City.

Test animals

Species:
mouse
Strain:
CD-1
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Kingston NY
- Age at study initiation: (P) x 11wks; (F1): 74+/-10 days
- Housing: by sex in solid bottom polypropylene or polycarbonate cages with stainless steel wire lids, then subsequently in breeding pairs. Ad-Sorb-Dri bedding used.
- Diet (ad libitum): NIH-07
- Water (ad libitum): tap
- Acclimation period: 2 weeks

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21-25
- Photoperiod (hrs dark / hrs light): 10/14

Administration / exposure

Route of administration:
oral: drinking water
Vehicle:
water
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: Each dosing solution formulated directly. No loss of compound after 3 days at room temperature and only 0.9% after 21 days lost. Consequently, fresh solutions prepared every 2 weeks.
Details on mating procedure:
- M/F ratio per cage: 1:1
- Length of cohabitation: 7-day pre-mating period and a 98-day cohabitation period.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Analysis on days 1, 6, 10 and 14. All doses found to be within 97-104% of expected values.
Duration of treatment / exposure:
14 weeks
Frequency of treatment:
continuous
Details on study schedule:
At the completion of the continuous breeding phase, the F0 breeding pairs were separated and housed individually and exposure to 2-butoxyethanol continued. When the last litter was weaned from the continous breeding phase F0 males and females from the 1 % dose group were mated with male and female control animals in a one-week crossover mating study to determine any sex-related reproductive effects of 2-butoxyethanol.

Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0 0.5, 1.0 or 2.0 %
Basis:
nominal in water
Remarks:
Doses / Concentrations:
0, 720, 1340 and 2050 mg/kg bw
Basis:
actual ingested
No. of animals per sex per dose:
20. 40 in control
Control animals:
other: Yes. Control males and females were also mated for comparative purposes.
Details on study design:
-The study itself consisted of three phases:
Phase 1 = continous breeding phase
Phase 2 = F0 males and females mated with control animals
Phase 3 = F1 males and females mated

Exposure to 2-butoxyethanol was discontinued during the one-week mating period and then reintroduced at 1 % dose level (estimated daily intake 1830 mg/kg bw). Control males and females were also mated for comparative purposes. The proportion of successful copulations from the breeding pairs was similar in all groups. However, the number of fertile females was significantly reduced in the group where treated females were mated with control males.

A final phase was conducted to assess the fertility and reproductive effects of 2-butoxyethanol in second generation (F1) pups. The pups selected were those born after the CBP and when the maternal animals were individually housed. As there were insufficient pups in the 1 and 2 % dose groups, only the pups from the 0.5 % dose group were used. The F1 generation pups were nursed, weaned and reared to sexual maturity. After weaning, the mice received 0.5 % 2-butoxyethanol in their drinking water (estimated daily intake 950 mg/kg bw). At 74 ± 10 days of age, the F1 animals from different litters were mated. The animals were necropsied after delivery.
Positive control:
No

Examinations

Parental animals: Observations and examinations:
CAGE SIDE OBSERVATIONS: Yes

DETAILED CLINICAL OBSERVATIONS: Yes

BODY WEIGHT: Yes

WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): Yes
Oestrous cyclicity (parental animals):
F0 Females Mated with Control Males
-When evaluated over the 7 day period prior to necropsy, proportionally more females (7/13) in the 1 % treated group than controls (9/38) had oestrus cycles longer than 7 days.
Sperm parameters (parental animals):
Parameters examined in P and F1 male parental generations
Litter observations:
Parameters examined in P and F1 generations
Postmortem examinations (parental animals):
Parameters examined in P and F1 generations
Postmortem examinations (offspring):
Parameters examined in F1 and F2 generations
Statistics:
Cochran-Armitage test for dose related trends in fertility. Chi-square test for differences in fertility among groups. Pairwise comparisons between dose groups and control using Fisher's exact test. Reproductive indices between groups compared using Kruskal-Wallis test and ordered differences using Jonckheere's test. Pairwise comparisons of treatment groups perfomed using Wilcoxon-Mann-Whitney U test. Pup number per litter corrected for when calculating average pup weight.
Reproductive indices:
The numbers of fertile pairs from the surviving pairs of the continuous breeding phase were 38/39, 19/19, 13/14 and 5/7 at 0, 0.5, 1.0 and 2.0 % dose levels, respectively.
Offspring viability indices:
no further information

Results and discussion

Results: P0 (first parental animals)

General toxicity (P0)

Clinical signs:
effects observed, treatment-related
Body weight and weight changes:
effects observed, treatment-related
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
not specified
Histopathological findings: non-neoplastic:
no effects observed
Other effects:
effects observed, treatment-related

Reproductive function / performance (P0)

Reproductive function: oestrous cycle:
no effects observed
Reproductive function: sperm measures:
not examined
Reproductive performance:
effects observed, treatment-related

Details on results (P0)

CLINICAL SIGNS AND MORTALITY (PARENTAL ANIMALS)
-During the 98 day cohabitation period, deaths occurred in the female mice: 13/20 in the 2 % group, 6/20 in the 1 % dose group, 1/20 in the 0.5 % dose group and 1/40 in the control group. In the male mice, no deaths occurred.

BODY WEIGHT AND FOOD CONSUMPTION (PARENTAL ANIMALS)
-The average body weights in the female 2 % dose group were consistently lower than the controls. Male groups experienced weight loss (1-2 % of initial body weight) in the two highest doses.
-At necropsy, male and female mice from the 1 % dose group had significantly lower body weights.

ORGAN WEIGHTS
-The 1% dose group exhibited increased relative kidney weights, and female also had significant increases in relative liver weight.
-A significant increase in relative kidney weight in the females, and a significant increase in relative liver weight in both the males and females were observed in the F1 generation.

TEST SUBSTANCE INTAKE (PARENTAL ANIMALS)
-Reduced fluid consumption was observed at all dose levels in both sexes (22 %, 18 % and 36 % reduction relative to controls at 0.5 %, 1.0 % and 2.0 %, respectively after 14 weeks of dosing).

REPRODUCTIVE FUNCTION: ESTROUS CYCLE (PARENTAL ANIMALS)
-In the 1% group no significant differences were observed between the control and treated animals for the average oestrous cycle length and frequency.
-No treatment-related changes in the oestrous cycle length and frequency were noted in the F1 generation.

REPRODUCTIVE FUNCTION: SPERM MEASURES (PARENTAL ANIMALS)
-In the 1% group no significant differences were observed between the control and treated animals for the weights of reproductive organs, sperm motility or morphology.
-No treatment-related changes in the weights of reproductive organs, sperm motility or morphology were noted in the F1 generation.

REPRODUCTIVE PERFORMANCE (PARENTAL ANIMALS)
-Significant reduction in reproductive performance occurred at 1 and 2 % dose levels as indicated by dose-related decrease in number of litters per fertile pair, litter sizes, pup viability and live pup weight.
-No significant fertility and reproductive effects were observed in the F1 animals as indicated by the proportions of successful copulation and fertile females, litter size, pup viability and live pup weights.

HISTOPATHOLOGY (PARENTAL ANIMALS)
-In the only histopathological examination carried out on the treated females, no treatment related kidneys lesions were observed.

Effect levels (P0)

Dose descriptor:
NOAEL
Effect level:
720 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: Body weight reduction, mortality, reproductive performance

Results: F1 generation

General toxicity (F1)

Clinical signs:
no effects observed
Mortality / viability:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Sexual maturation:
no effects observed
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Gross pathological findings:
no effects observed
Histopathological findings:
no effects observed

Details on results (F1)

BODY WEIGHT (OFFSPRING)
A small but significant reduction (by 3 %) of live F1 pup weight was also observed in the 0.5 % dose group without other significant reproductive effects but no effect was seen in the pups born to the F1 generation.

On the F1 generation (low dose and control only tested) , there were no effects on mating and fertility indices or any other reproductive parameter. There were no significant differences in F2 litter size, pups numbers or weights. There were no effects on sperm parameters in males or estrous cyclicity in females but relative liver and kidney weights were increased in the dose group receiving 2-butoxyethanol (0.5%)

Effect levels (F1)

Dose descriptor:
NOAEL
Generation:
F1
Effect level:
720 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: pup weight

Results: F2 generation

Effect levels (F2)

Dose descriptor:
NOAEL
Generation:
F2
Effect level:
720 mg/kg bw/day
Sex:
male/female
Basis for effect level:
other: No reproductive effects observed at single dose tested

Overall reproductive toxicity

Reproductive effects observed:
not specified

Any other information on results incl. tables

The continuous breeding first generation study showed that treatment significantly affected fertility at the mid and high dose levels and marginally affected pup weight at the lowest level. The cross-breeding results suggest that the fertility effects were only due to effects on the female mice. In the cross-over study there were no effecting on mating index (number with copulatory plugs/number co-habited), however for females that co-habited with control males the fertility index (number fertile/number with copulatory plugs) ws significantly reduced compared to the controls and other cross-over group. Similarly, the number of live pups/litter of the 1% treated females/control males fell. These effects may have been an indirect consequence of the severe general systemic toxicity rather than a direct effect of 2-butoxyethanol on the reproductive organs.

Applicant's summary and conclusion

Conclusions:
In this study, effects were seen on fertility only at doses which were severely toxic to the animals (1340 and 2050 mg/kg bw/d). A NOAEL of 720 mg/kg bw/day can be identified for fertility effects by oral route in mice. Marginal pup weight reductions at this dose were not repeated in the second generation and therefore not regarded as a significant finding.
Executive summary:

In a continuous breeding study carried out to an NTP protocol, mice were exposed for a period of 14 weeks to the oral drinking water routes at doses from 720 to 2050mg/kg of the substance 2 -butoxyethanol. Significant adverse reproductive effects were observed in the females at very high dose levels (1340 mg/kg and above) which also caused severe general toxicity, including death. Under the conditions of the study, the NOAEL for reproductive toxicity of 2-butoxyethanol (fertility) can be set as 720 mg/kg/day. For developmental toxicity, no NOAEL can be derived. A conservative LOAEL of 720 mg/kg/day can be taken as only a very slight decrease in pup weight was observed at this dose.

At the lowest dose studied, the only adverse finding was a marginal statistically significant reduction in pup weight. However, since this reduction was only 3% compared to controls and was not repeated in the F1 generation, it was not considered a significant adverse finding.

No NOAEL or LOAEL can be determined for systemic parental toxicity because this kind of study is not designed to assess systemic toxicity although it is of note that there were effects (reduced fluid consumption) even at the lowest dosage of 720 mg/kg/day

On the basis that the toxicity of this series of ethylene glycol butyl ethers tends to decrease with increasing number of ethylene oxide units, the toxicity to reproduction of 2 -(2 -butoxyethoxy)ethanol is expected to be less that that of butoxyethanol and therefore this is an adequate substance for read across purposes for this end point.