Ethanol, 2-ethoxy-, manufacture of, by-products from

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

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

Adequacy of study:

key study

Study period:

1990

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study. Rationale for weight of evidence approach using read across substances is described in the overall section summary.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1990

Report date:

1990

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OTS 798.4900 (Prenatal Developmental Toxicity Study)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

2-(2-(2-methoxyethoxy)ethoxy)ethanol

EC Number:

203-962-1

EC Name:

2-(2-(2-methoxyethoxy)ethoxy)ethanol

Cas Number:

112-35-6

Molecular formula:

C7H16O4

IUPAC Name:

2-[2-(2-methoxyethoxy)ethoxy]ethanol

Details on test material:

- Name of test material (as cited in study report): Triethylene Glycol Monomethyl Ether (TGME)
- Storage condition of test material: Room temperature.
- Other: Documentation of the methods of synthesis, purity, stability and characterization of the test substance are available in the Sponsor’s records.

Test animals

Species:

rat

Strain:

other: Crl:CD(SD)BR

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc., Portage Michigan.
- Age at study initiation: (females) 71 days old; (males) 73 days old
- Weight at study initiation: (females) 174 – 227 g; (males) 238 – 342 g
- Housing: Female rats were housed individually in wire-bottomed stainless-steel cages suspended above absorbent paper liners, except during cohabitation.
- Diet: Certified Rodent Chow #5002 (Ralston Purina) ad libitum
- Water: Local water passes through a reverse osmosis membrane was available ad libitum from an automatic watering system.
- Acclimation period: one week

ENVIRONMENTAL CONDITIONS
- Temperature: 70 - 78°F
- Humidity: 35 - 65%
- Air changes: minimum 10 changes/hr 100% fresh air that had been passed through 99.97% HEPA filters (Airo Clean~ room).
- Photoperiod: 12 hour light/12 hour dark cycle.

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

unchanged (no vehicle)

Details on exposure:

Test material was administered as received. The control group rats were administered R.O. deionized water at a dosage volume (4.8 mL/kg) that was equivalent to that given to the highest dosage group.

Analytical verification of doses or concentrations:

no

Details on analytical verification of doses or concentrations:

Test material was administered as received.

Details on mating procedure:

- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1:1
- Length of cohabitation: (5 days) Afternoon of September 19, 1989 to the morning of September 24, 1989.
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy:sperm in vaginal smear referred to as day 0 of pregnancy

Duration of treatment / exposure:

Gestation days 6-15

Frequency of treatment:

Once daily on gestation day 6 -15. Each daily dosage given to the rats was administered at approximately the same time each day [between 1242 and
1542 hours EDT (1142 and 1442 hours EST)].

Duration of test:

Animals were sacrificed on day 20 of gestation.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

625 mg/kg bw/day

Doses / concentrations 2

Dose / conc.:

1 250 mg/kg bw/day

Doses / concentrations 3

Dose / conc.:

2 500 mg/kg bw/day

Doses / concentrations 4

Dose / conc.:

5 000 mg/kg bw/day

No. of animals per sex per dose:

25

Control animals:

yes

Details on study design:

- Dose selection rationale: All dosages were adjusted daily on the basis of individual body weights recorded immediately prior to intubation.
- Rationale for animal assignment (if not random): The rats were assigned to the five dosage groups using a computer-generated (weight-ordered) randomization procedure based on day 0 of gestation body weights.
- Other: Animals were chosen for evaluation because it is one mammalian rodent species standardly accepted for use in developmental toxicity, this strain of rat had been demonstrated to be sensitive to developmental toxicants, it has been used throughout industry for nonclinical studies of developmental toxicity, historical control data and experience exist and it is the rodent species specified in the TGME Testing Consent Order.

Examinations

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: Daily. Viability checks were made twice daily.

BODY WEIGHT: Yes
- Time schedule for examinations: Once weekly prior to mating. Day 0 of presumed gestation and daily during the dosage and postdosage periods.

FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Day 0 of presumed gestation and daily during the dosage and postdosage periods.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day # 20
- Organs examined: uterus, ovaries.

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 resorptions: Yes
- Other: Gravid uterine weights were inadvertently not recorded for 8, 5, 4, 5 and 7 dams in the O(Control), 625, 1250, 2500, and 5000 mg/kg/day dosage groups, respectively. As sufficient numbers of values were available to allow adequate interpretation of these data, this deviation did not adversely affect the outcome of the study.

Fetal examinations:

- External examinations: Yes
- Soft tissue examinations: Yes: half per litter.
- Skeletal examinations: Yes: half per litter.
- One-half of fetuses in each litter were examined for soft tissue alterations using an adaptation of Wilson’s sectioning technique. The remaining fetuses in each litter were cleared, stained with alizarin red S and examined for skeletal alterations.

Statistics:

Maternal and foetal incidence were analyzed using the Variance Test for Homegeneity of the Binomial Distribution. Maternal body weight and feed consumption data, organ weight data, and litter averages for percent male fetuses, percent dead or resorbed conceptuses per litter, fetal body weights, fetal ossification sites, and percent fetal alterations were analyzed using Bartlett’s Test of Homogeneity of Variances and the Analysis of Variance, when appropriate. If the Analysis of Variance was significant (P ≤ 0.05), Dunnett’s test was used to identify the statistical significance of individual groups. If the Analysis of Variance was not appropriate, the Kruskal-Wallis Test was used when less than or equal to 75% ties were present; when more than 75% ties were present, the Fisher’s Exact Test was used. In cases where the Kruskal-Wallis Test was statistically significant (P ≤ 0.05), Dunn’s Method of Multiple Comparisons was used to identify the statistical significance of individual groups. All other Caesarean-sectioning data were evaluated using the procedure previously described for the Kruskal-Wallis Test.

Indices:

Gross external or soft tissue variations in the fetuses.

Historical control data:

No further data available.

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
One non-pregnant high dosage group rat was found dead on day 13 of presumed gestation. Ataxia, decreased motor activity, and lost righting reflex occurred for this rat, as well as persistent weight loss and reduced feed consumption beginning after the initial dosage. Necropsy revealed distention of the stomach and intestines with gas; observations that were probably secondary to moderate autolysis. No other deaths occurred. The 5000 mg/kg/day dosage of TGME caused significant numbers (P≤ 0.01l) of rats to have decreased motor activity, excess salivation, ataxia, and impaired righting reflex, as compared with the control group numbers. Lost righting reflex, urine-stained abdominal fur, and rales occurred for a few high dosage group rats. The 5000 mg/kg/day dosage group rat that was found dead (13364) had gaseous distention of the stomach and intestines-that may have been caused by autolysis or anorexia. As described in the following information, all other gross lesions were considered unrelated to the test substance and occurred at incidences that were not dosage dependent. One 1250 mg/kg/day dosage group rat (13310) had common urinary tract lesions. Another 1250 mg/kg/day dosage group rat (13312) had a mass, in the region of the mammary gland that was probably a galactocele C. Significant reductions (P ≤ 0.01) in maternal body weight gains occurred for the high dosage group during the dosage period (calculated as days 6 to 16 of gestation). As a result the mean maternal body weights were significantly reduced (P ≤ 0.05 to P ≤ 0.01) for the high dosage group on days 9, 12 and 16 of gestation, as well as during the post-treatment period, on days 18 and 20 of gestation. The mean gravid uterine weight was significantly lower (P ≤ 0.05) for the high dosage group, and the mean maternal body weight on day 20e (day 20 of gestation body weight corrected for the gravid uterine weight) tended to be lower for the high dosage group, as compared with the control group mean. Reflecting these events, maternal body weight changes from days 6 to 20e and 0 to 20e of gestation were significantly decreased (P ≤ 0.05 to P ≤ 0.01) for the high dosage group. Maternal body weights were not affected by treatment with TGME for any of the other dosage groups. Groups given 2500 and 5000 mg/kg/day of TGME had statistically significantly reduced feed consumption for the entire dosage period (calculated as days 6 to 16 of gestation), as compared with the control group. The 2500 mg/kg/day dosage group had significantly reduced (P ≤ 0.05 to P ≤ 0.01) relative maternal feed consumption from days 6 to 16, 6 to 18, and 12 to 16 of gestation, and the 5000 mg/kg/day dosage group had significantly reduced (P ≤ 0.01) mean absolute and relative maternal feed consumption throughout the entire dosage period. Feed consumption was significantly increased (P ≤ 0.05 to P ≤ 0.01) for the high dosage group during the post dosage period, as compared with the control group values. Despite these rebound phenomena, absolute and relative maternal feed consumption values were significantly decreased (P ≤ 0.01) for the high dosage group on days 6 to 20 of gestation and for the entire gestation period (days 0 to 20 of gestation).

Effect levels (maternal animals)

1

Dose descriptor:

NOEL

Effect level:

625 mg/kg bw/day

Basis for effect level:

other: maternal toxicity

2

Dose descriptor:

NOAEL

Effect level:

1 250 mg/kg bw/day

Basis for effect level:

other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Small, significant increases (P ≤ .05 to P ≤ 0.01) n embryo-fetal lethality occurred in the 5000 mg/kg/day dosage group (litter averages for total resorptions, late resorptions, percentage of resorbed conceptuses and dams with at least one resorption), as compared with the control group values. None of these effects were seen for the other dosage groups. Dosages of 1250 mg/kg/day and higher reduced fetal body weights. However, this effect was statistically significant (P ≤ 0.01) for only the 2500 and 5000 mg/kg/day dosage groups. Dosages of TGME as high as 5000 mg/kg/day did not cause gross external, internal soft tissue or skeletal malformations in the fetuses. These dosages also did not result in increased incidences of gross external or soft tissue variations in the fetuses. There were no dosage-dependent, statistically significant differences in the litter or fetal incidences of these parameters. Groups given 1250 mg/kg/day and higher dosages of TGME had significant increases (P ≤ 0.05 to P ≤ 0.0l) in the litter and/or fetal incidences of reversible delays in fetal ossification, common observations in fetuses with reduced body weights. The 2500 and 5000 mg/kg/day dosage groups also had significant increases (P ≤ 0.05 to P ≤ 0.0l) in the litter and/or fetal incidences of cervical ribs, a common variation in rats.

Effect levels (fetuses)

Dose descriptor:

NOEL

Effect level:

625 mg/kg bw/day

Based on:

test mat.

Sex:

not specified

Basis for effect level:

other: developmental effects

Fetal abnormalities

Abnormalities:

not specified

Overall developmental toxicity

Developmental effects observed:

not specified

Any other information on results incl. tables

The death of one nonpregnant high dosage group (5000 mg/kg/day) rat was considered treatment-related. This rat had clinical signs of treatment with the test substance (ataxia, decreased motor activity, lost righting reflex, persistent weight loss, and reduced feed consumption). At necropsy, gaseous distention of the stomach and intestines may have been interrelated with anorexia and/or moderate autolysis. No other deaths occurred in this study, and no other gross lesions seen during necropsy examination were considered to be effects of TGME.

Applicant's summary and conclusion

Conclusions:

The results from this study indicate that TGME is not a developmental toxicant and does not present a unique hazard to development of rat conceptuses. There were no increases in the incidence of external, internal, soft tissue, or skeletal malformations or in the incidences of external or internal soft tissue variations at doses as high as 5000 mg/kg/day of TGME.

Executive summary:

In a guideline (TSCA) and GLP study, the developmental toxicity of TGME was evaluated in rats following oral (gavage) administration at dosages up to 5,000 mg/kg bw/day on Days 6-15 of gestation. The 1250 mg/kg/day dose level slightly reduced relative feed consumption during the dosing period, but the absolute feed consumption at this level and the average maternal body weights and body weight gains at the 1250 and 2500 mg/kg/day levels were not affected by treatment with TGME. Thus, 1250 mg/kg/day is considered a NOAEL, and 625 mg/kg/day is considered a NOEL for maternal toxicity.

There were no increases in the incidence of external, internal soft tissue, or skeletal malformations or in the incidences of external or internal soft tissue variations at doses as high as 5000 mg/kg/day of TGME. Reversible delays in fetal ossification and slightly lower fetal body weight (not statistically different from the control weight) occurred in the 1250 mg/kg/day dosage group. Thus, 1250 mg/kg/day is considered very near the NOAEL for TGME developmental toxicity, and 625 mg/kg/day is considered the NOEL for developmental toxicity.

For TGEE, the lowest member of the homologous series in this UVCB substance, this would be equivalent in molecular weight terms to 678 and 1357mg/kg/day respectively.