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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:
1988
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with national standard methods
Remarks:
Well documented NTP publication which meets basic scientific principles. Study process involves detailed quality assurance.

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1989
Report date:
1989
Reference Type:
publication
Title:
Cardiovascular development (CVD) in F-344 rats following phase-specific exposure to butoxy ethanol.
Author:
Sleet RB, Price CJ, Marr MC, Morrissey RM and Schwetz BA
Year:
1991
Bibliographic source:
Teratology, 43(5), p466.
Reference Type:
publication
Title:
Developmental toxicity: status of the field and contribution of the NTP.
Author:
Schwetz BA, Harris MW
Year:
1993
Bibliographic source:
Env Hlth Persp, 100, 269-82

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
, dosing not for whole gestation period and split into two cohorts, each with only 3 day but different dosing windows
Principles of method if other than guideline:
The study was specifically designed to assess the critical periods of cardiovascular development - a known target of other glycol ethers. Dosing was only over two narrow windows and not the whole gestation period.
GLP compliance:
not specified
Limit test:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
2-butoxyethanol
EC Number:
203-905-0
EC Name:
2-butoxyethanol
Cas Number:
111-76-2
Molecular formula:
C6H14O2
IUPAC Name:
2-butoxyethanol
Details on test material:
- Supplier: Chem Central, Kansas City.
- Purity >99%, verified
- Lot #C092882/B01
- Stability: stable for 2 weeks at 60C. Aqueous solutions (5%) showed no significant loss over 3 weeks storage at room temperature.

Test animals

Species:
rat
Strain:
Fischer 344
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Kingston NY
- Age at study initiation: 8-12 weeks on arrival in laboratory.
- Weight at study initiation: 150-202g (females)
- Housing: females 3 per cage in solid bottomed polycarbonate or polypropylene cages. Males housed individually
- Diet (ad libitum): Purina lab chow 5002 < 5 months old
- Water (ad libitum): deionised/filtered
- Acclimation period: 7 days


ENVIRONMENTAL CONDITIONS
- Temperature (°F): 68.4-72.2
- Humidity (%): ~48
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: distilled water
Details on exposure:
VEHICLE
- Amount of vehicle (if gavage): 5ml/kg
Analytical verification of doses or concentrations:
not specified
Details on mating procedure:
- Impregnation procedure: cohoused when in estrus or proestrus.
- If cohoused:
- M/F ratio per cage: 1:1
- Length of cohabitation: overnight
- After overnight co-habitation, females checked by vaginal smear for presence of sperm. Sperm positive females removed. Sperm negative females returned to group cages
- Proof of pregnancy: sperm in vaginal smear referred to as day 0.
Duration of treatment / exposure:
Gestation days 9–11 or 11-13 (two separate studies)
Frequency of treatment:
Daily (a.m.)
Duration of test:
to GD 20
Doses / concentrationsopen allclose all
Remarks:
0, 30, 100, 200, mg/kgbw, dose given for experiment GD 9-11
Remarks:
0, 30, 100, 200 300, mg/kgbw, dose given for experiment GD 11-13
No. of animals per sex per dose:
298 total, 104 rats served as controls.
Control animals:
yes
Details on study design:
- Dose selection rationale: range finder test.
- Rationale for animal assignment (if not random): method of stratified randomisation or by random number.

Examinations

Maternal examinations:
DETAILED CLINICAL OBSERVATIONS: Yes, daily

BODY WEIGHT: Yes, days gd 0, 6, 9-20 (or 9, 11-20)

FOOD AND WATER INTAKE: Yes, gd 0, 6, 9, 11-15, 17, 20 and on gd 10 for gd 9-11 dose animals.

POST-MORTEM EXAMINATIONS: Yes / No / No data
- Sacrifice of 93 rats occurred on gestation day #12, sacrifice of 207 rats occurred on gestation day #20 (foetuses were also sacrificed at this time), sacrifice of 104 rats occurred on gestation day #14.

HAEMATOLOGY:: Yes, gd 12, 14 and at termination. Animals sacrificed at these time points (CO2 anasthesia)

ORGAN WEIGHTS: Yes
Ovaries and uterine content:
The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Number of implantations: Yes
- Number of resorptions: Yes
- Number of corpora lutea: Yes
- Number of dead and live fetuses: yes
- fetal blood also collected for pooled analysis by litter.
Fetal examinations:
- External examinations: Yes: No data
- Soft tissue examinations: Yes: number unclear
- Skeletal examinations: Yes: all per litter: fixed and double stained (Alician blue and red)
- Head examinations: Yes: half per litter: fixed in Bouin's solution for free hand sectioning and examination.
Statistics:
Parametric procedures were applied to selected measures from the teratology study. Appropriate General Linear Models (GLM) procedures for these Analyses of Variance were used. Prior to GLM analysis, an arcsine—square root transformat ion was performed on a all linear derived percentage data. Bartlett’s test for homogeneity of variance (alpha level = 0. 001) was performed on all data to be analyzed by ANOVA. GLM analysis was used to determine the significance of the dose—response relationship (Test for. Linear Trend), and to determine whether significant dose effects, replicate effects or dose X replicate interactions had occurred for selected measures (ANOVA). When a significant (p<0.05) main effect for dose occurred, William’s Multiple Comparison Test or Dunnett’s Multiple Comparison was used to compare each exposed group to the vehicle control group for that measure. A one-tailed William’s Test and/or DunnetUs Test was employed for all pairwise comparisons except that a two-tailed test was utilized for maternal and fetal body weight parameters. In addition, the data for any measure which showed a significant (p<0.05) dose X replicate interaction in a two-way (dose X replicate) ANOVA were presented as mean ± S . E . M. for each cell in the ANOVA design. and dose effects within each replicate were further evaluated using a one—way ANOVA, Test for Linear Trend, William’s Multiple Comparison Test and/or Dunnett’s Test. Nonsignificant (i.e. , p>0.05) replicate effects or dose X replicate interactions on selected measures warranted pooling data across replicates for nonparametric analysis. When significant (p<0.05) replicate effects or dose X replicate interactions occurred, nominal scale data for related measures were presented separately for each replicate in the study, as well as for all replicates combined.
Indices:
no data
Historical control data:
no data

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
effects observed, treatment-related
Description (incidence and severity):
Stained bedding in mid and high dose groups (6 and 10 dams respectively) and one dam with vaginal bleeding and some animals showing vaginal discharge in high dose group (varying from day to day). Some animals in high dose group reported as pale and with rough coats, although the latter was not considered treatment related due to inconsistent findings. No other significant findings of note
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
GD9-11 treated group: Dosing resulted in an immediate weight loss in the mid and high dose groups, which was at its most pronounced on the last day of treatment (-1.6%, -9.2% respectively, at which point the high dose animals were still loosing weight). The mid dose animals recovered to a similar weight to controls by sacrifice on GD20 but the high dose animals remained 9.4% below the weight of the control animals. Similar results were also seen in the cohort that was treated similarly but sacrificed on day 12 and with the cohorts treated on GD11-13 and sacrificed on days 14 and 20.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Severe but temporary reduction of absolute food consumption on commencement of treatment. Mid dose group halved on first day then recovered to control levels. High dose group showed ~60% reduction over first three days of treatment with a recovery to control levels 5 days after commencing treatment. Consumption relative to bodyweight showed a similar but less marked pattern of effect.
Water consumption and compound intake (if drinking water study):
effects observed, treatment-related
Description (incidence and severity):
Marked but temporary reduction of absolute water consumption on commencement of treatment. Mid dose group showed ~20% reduction on first day then recovered to control levels. High dose group showed ~50% reduction on first day of treatment with a recovery to control levels 2 days after commencing treatment. There still remained some days up to the GD20 sacrifice dates when water consumption was significantly different to controls (increased or decreased) in both dose groups. Consumption relative to bodyweight showed a similar but less marked pattern of effect.
Haematological findings:
effects observed, treatment-related
Description (incidence and severity):
The maternal effects of 2-butoxyethanol given from GD 9 - 11 or from GD11 - 13 at doses of and greater than 100 mg/kg/day included severe haematotoxicity. In particular, dramatic reductions in circulating red blood cells, haematocrit and haemoglobin resulted by 24 hours after treatment. By GD20 the haematotoxic effects were nearly reversed. The changes observed in haematological parameters are typical of haemolytic anaemia and the compensatory haematopoietic response associated with recovery. Typical significant changes as follows (GD 9-11 treated, sacrifice GD12): Hematocrit, mid dose -27%, high dose -44%; Haemoglobin, mid dose -28%, high dose -45%; RBC, mid dose -33%, high dose -55%; Reticulocytes, mid dose +130%, high dose +290%; WBC, mid dose +185%, high dose +360%; Platelet count, mid dose +9.5%, high dose +20%; mean corpuscular volume, high dose +26%; mean corpuscular haemoglobin, mid dose +8%, high dose +22% ;mean corpuscular Hgb, high dose -3%. In the animals left to GD20, there was some degree of recovery but significant differences remained.
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
Relative and absolute liver weight reduction in high dose animals (~-11% for both). Relative and absolute spleen weight increase in mid high dose animals (absolute +12%, +25%, relative +15%, +40% respectively.) Relative kidney weight increase in high dose animals (+20%). Similar results were also seen in the cohort that was treated similarly but sacrificed on day 12 and with the cohorts treated on GD11-13 and sacrificed on days 14 and 20. The changes observed in organ weights in this study are typical secondary consequences of haemolytic anaemia. Gravid uterine weight was significantly reduced in high dose group (-38%).
Gross pathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined

Maternal developmental toxicity

Number of abortions:
not specified
Pre- and post-implantation loss:
no effects observed
Description (incidence and severity):
Slight trend towards increase in high dose group in some but not all cohorts. Never statistically significant. Replicate II showed a significant increaes compared to the concurrent control, but this was unusually low - much lower than the control values in replicate I cohort.
Total litter losses by resorption:
effects observed, treatment-related
Description (incidence and severity):
Significant increased resorptions in high dose group. 11 of the 22 litters in the GD9-11 dose cohort had resorptions, with 10 showing 100% absorptions. There was also an increasing trend with dose. In replicate group I, this trend was more pronounced, with effects also evident at the mid dose group, although not statistically significantly raised compared to the control. (26% versus 4-5% in controls and other dose groups in replicate g).
Absolute resorptions per litter overall (SD in brackets): Control 0.41 (0.23), low dose (0.38 (0.12), mid dose 0.43 (0.20), high dose 3.0 (0.86).
Early or late resorptions:
effects observed, treatment-related
Description (incidence and severity):
Non-live implants, and adversely affected implants per litter were seen in the 200 mg/kg/day group dosed on GD 9 - 11.
Dead fetuses:
no effects observed
Changes in pregnancy duration:
not examined
Changes in number of pregnant:
no effects observed
Other effects:
no effects observed
Description (incidence and severity):
On number of implantation sites per litter, corpora lutea per dam. Slight trend towards changes with increased dose in some but not all cohorts but never statistically significant.

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
30 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
body weight and weight gain
food consumption and compound intake
haematology
organ weights and organ / body weight ratios
water consumption and compound intake

Maternal abnormalities

Abnormalities:
no effects observed

Results (fetuses)

Fetal body weight changes:
no effects observed
Reduction in number of live offspring:
no effects observed
Changes in sex ratio:
no effects observed
Changes in litter size and weights:
effects observed, treatment-related
Description (incidence and severity):
There was a trend towards reduced weight in all dose groups but it was not statistically significant in the lower two dose groups.
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
no effects observed
Other effects:
effects observed, treatment-related
Description (incidence and severity):
Decreased fetal blood platelet count but no embryolethality in the 300 mg/kg/day group dosed on GD 11 - 13.
Details on embryotoxic / teratogenic effects:
No adverse effects were seen on the cardiac system which are typical effects of other glycol ethers that have teratogenic potential. Embryo/fetal viability and growth were not selectively affected by 2-butoxyethanol since the responses observed followed the exposure to doses that were also maternally toxic.

Effect levels (fetuses)

Dose descriptor:
NOAEC
Effect level:
200 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: fetal blood platelet count

Fetal abnormalities

Abnormalities:
no effects observed

Overall developmental toxicity

Developmental effects observed:
yes
Lowest effective dose / conc.:
200 mg/kg bw/day
Treatment related:
yes
Relation to maternal toxicity:
developmental effects as a secondary non-specific consequence of maternal toxicity effects
Dose response relationship:
yes
Relevant for humans:
no

Any other information on results incl. tables

The effects seen are a secondary consequence of haemolytic anaemia caused by the substance. Humans are resistant to this effect.

Applicant's summary and conclusion

Conclusions:
2-butoxyethanol is neither a specific developmental toxicant nor a teratogen at doses that cause marked maternal toxicity.
Executive summary:

An a well reported developmental toxicity study designed specifically to look at adverse developmental effects on the cardiac system, pregnant rats were exposed during GD9 -11 and GD11 -13 in two separate experiments to 2-butoxyethanol by oral gavage. Doses of up to 200mg/kg were used. No increase in foetal malformations; particularly, no cardiovascular malformations were observed. When 200 mg/kg was given from day 9 to 11, an increased foetal lethality without malformations was noted. When 300 mg/kg were given from gd 11 to 13, a decrease platelet count was seen in foetuses. For developmental toxicity the NOAEL is 100 mg/kg when 2-butoxyethanol is administered from GD 9 - 11 and that this is a more vulnerable period for the fetus than GD 11 -13. It should however be noted that marked maternal toxicity was seen at doses of 100mg/kg including body weight gain reductions, organ weight changes and severe haemotoxicity. This leads to the conclusion that developmental toxicity is not a concern and is secondary to maternal toxicity.

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