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Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
not specified
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: The study was equivalent or similar to a guideline but was non-GLP.

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1979
Report Date:
1979
Reference Type:
publication
Title:
Unnamed
Year:
1979

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Gas Chromatographic Analysis:
Epichlorohydrin 99.8% (wt.)
2,3-Dichloropropene 0.11% (wt.)
Cis-1,3-Dichloropropene 0.03% (wt.)
Beta Chloroallyl Alcohol 0.01% (wt.)

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
Sprague-Dawley rats (Spartan Research Animals, Haslett, MI) weighing approximately 250 to 300 g were used. The rats were bred by the supplier, day 0 of pregnancy was identified as the day on which sperm were found in a vaginal smear. During the periods between exposures, the animals were individually housed in wiremesh cages in rooms designed to control temperature, relative humidity and light cycle. Test animals were provided with laboratory animal chow (Ralston Purina Company, St. Louis, MO) and tap water ad libitum. During exposures, food and water were not available and animals were group-housed.

Administration / exposure

Route of administration:
inhalation: vapour
Type of inhalation exposure (if applicable):
whole body
Vehicle:
unchanged (no vehicle)
Details on exposure:
Whole-body exposures were carried out in 4.3 cubic meter Rochester-type chambers. The epichlorohydrin atmosphere was generated by metering the liquid compound at a controlled rate into a temperature regulated vaporization flask (75 to 100°C). The vapor was swept with compressed air into the inlet of the exposure chamber where it was further diluted to the desired concentration by tempered air. The flow rate was in the range of 600-900 liters per minute. Control animals were placed in identical chambers and exposed to filtered air.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
Chamber air was assayed for vapor concentration via GC analysis 2-7 times during each 7-hour exposure period. The mean analytical concentrations were 2.46+/-0.32 and 24.6+/-1 .0 ppm for the target levels of 2.5 and 25 ppm, respectively.
Details on mating procedure:
Purchased bred rats by the supplier.
Duration of treatment / exposure:
7 hours/day
Frequency of treatment:
day 6-15 of gestation
Duration of test:
21 days
Doses / concentrationsopen allclose all
Remarks:
Doses / Concentrations:
0, 2.43+/-0.32 or 24.4+/-1.1 ppm
Basis:
nominal conc.
Remarks:
Doses / Concentrations:
0, 2.46+/-0.32 or 24.6+/-1.0 ppm
Basis:
analytical conc.
No. of animals per sex per dose:
Groups of 43-46 pregnant rats (rabbits are discussed under a separate record)
Control animals:
yes, sham-exposed
Details on study design:
No additional information available.

Examinations

Maternal examinations:
Animals were observed daily during the exposure period for signs of toxicity. Body weights were recorded on gestation days 6, 10, 12, and 16. Food and water consumption were also recorded at 3-day intervals beginning on gestation day 6.

Cesarean-sections were performed on all animals on gestation day 21. Liver weights were recorded at that time, and samples of nasal turbinates, trachea, and lungs were preserved for possible histologic examination. Examination of these tissues was not deemed necessary.




Ovaries and uterine content:
At cesarean-section, developmental parameters were recorded: the number and position of live, dead, and resorbed fetuses, the number of corpora lutea, and the weight of the gravid uterus with ovaries attached.
Fetal examinations:
After being weighed and measured for crown-rump length, all fetuses were examined for external alterations and cleft palate. One-third of each litter, randomly selected, was examined for evidence of soft-tissue alterations by fresh dissection. Heads of fetuses undergoing soft-tissue examination were preserved in Bouin's fixative and examined by free-hand serial sectioning. All fetuses were then eviscerated and processed for skeletal examination.
Statistics:
Statistical evaluation of the incidence of fetal alterations and resorptions was made by the Wilcoxon test as modified by Haseman and Hoel (1974). Analysis of other incidence data was made by the Fisher's exact probability test (Siegel, 1956). Analyses of fetal body weights and organ weights, and food and water consumption data were made by an analysis of variance. Group meanswere compared to control values using Dunnett's test (Steel and Torrie, 1960). Statistical outliers (a =0.02) were deleted from the calculation of mean values for food and water consumption (Grubbs, 1969). The nominal alpha level used for statistical analyses of all experimental parameters was a = 0.05.
Indices:
not applicable
Historical control data:
available for review

Results and discussion

Results: maternal animals

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
At the maternal LOAEL, effects including reduced body weight and food intake for the dams were observed at the 25 ppm exposure level. The NOEL for maternal toxicity was 2.5 ppm.


BODY AND LIVER WEIGHTS OF PREGNANT RATS EXPOSED TO EPICHLOROHYDRIN BY INHALATION
ppm Epichlorohydrin (a)
0 2.5 25

Number of Dams(b) 36 33 32

Maternal body weight (g)
on gestation day
6 277+/-26(c) 263+/-27(d) 262+/-20(d)
8 284+/-26 272+/-23 263+/-20(d)
10 294+/-27 280+/-24(d) 271+/-20(d)
12 305+/-28 294+/-23 283+/-21(d)
16 334+/-31 324+/-23 311+/-2l(d)
21 413+/-41 402+/-43 394+/-31

Maternal body weight gain (g)
on gestation days
6-7 6+/-3 8+/-6 1+/-7(d)
8-9 10+/-5 8+/-9 8+/-9
10-11 1+/-5 14+/-8 12+/-8
12-15 30+/-9 31+/-7 29+/-9
16-20 79+/-17 77+/-21 82+/-19
Total 6-21 136+/-27 138+/-34 131+/-28

Maternal Liver weight on day 21
Absolute(e) 15.97+/-1.77 16.12+/-1.72 15.83+/-1.76
Relative(f) 38.82+/-3.87 40.25+/-2.90 40.30+/-4.08

Gravid Uterus Weight on day 21
Absolute(d) 97+/-29 89+/-34 93+/-24
Adjusted body
weight (g) 316+/-25 313+/-22 297+/-22(d)


(a) Bred rats were exposed to 0, 2.5, or 25 ppm epichlorohydrin by inhalation for 7 hrs/day on days 6 through 15 of gestation.

(b) Data from non-pregnant rats were not included in these analyses.

(c) Mean +/- S.D.

(d) Significantly different frcm controls by Dunnett's test P<0.05.

(e) g, Mean +/- S.D.

(f) g, liver/kg body weight, mean +/- S.D.

(g) Maternal body weight minus the gravid uterus weight (g), mean +/- S.D.



FOOD AND WATER CONSUMPTION OF PREGNANT RATS EXPOSED TO
EPICHLOROHYDRIN BY INHALATION

ppm Epichlorohydrin(a)
0 2.5 25

Number of Dams(b) 36 33 32
Food consumption on gestation days(c)
6-8 23+/-3(d) 24+/-4 20+/-3(e)
9-11 24+/-3 24+/-3 22+/-3(e)
12-14 25+/-2 26+/-3 24+/-3(e)
15-17 31+/-4 30+/-4 30+/-3
18-20 30+/-3 29+/-3 31+/-3

Water consumption on gestation days(e)
6-8 47+/-8 47+/-8 49+/-11
9-11 52+/-9 55+/-10 58+/-11(e)
12-14 57+/-7 59+/-9 65+/-11(e)
15-17 68+/-9 70+/-10 74+/-9(e)
18-20 63+/-8 65+/-9 69+/-9(e)

(a) Bred rats were exposed to 0, 2.5, or 25 ppm epichlorohydrin by inhalation for 7 hrs/day on days 6 through 15 of gestation.

(b) Data from non-pregnant rats were not included in these analyses.

(c) Expressed as grams/rat/day.

(d) Mean +/- S.D.

(e) Significantly different from control values by Dunnett's test P<0.05.


OBSERVATIONS AT CESAREAN SECTION OF RATS EXPOSED TO EPICHLOROHYDRIN BY INHALATION
ppm Epichlorohydrin(a)
0 2.5 25

Number of bred females 46 44 43
Number of maternal
deaths/number bred 0/46 0/44 0/43
Apparent pregnancy rate, %(b) 85(39/46) 86(38/44) 77(33/43)
Pregnant with stain only, %(c) 29(2/7) 0(0/6) 10(1/10)
Percent pregnant, total(d) 89(41/46) 86(38/44) 79(34/43)
Number of Litters 36 33 32
Corpora lutea/dam(e) 14+/-2 14+/-3 15+/-2
Implantation sites/ dam(e) 13+/-3 12+/-4 13+/-3
Preimplantation loss(e) 2+/-2 3+/-0 2+/-0
Fetuses/Litter(e) 12+/-4 11+/-4 12+/-3
Resorptions/Litter (e,f) 1+/-1 1+/-1 1+/-2
Percent Implantations resorbed(f) 5(25/460) 4(17/386) 7(30/408)
Percent litters with resorptions(f) 44(16/36) 36(12/33) 44(14/32)
Litters totally resorbed(f) 0 0 0
Resorptions/litters
with resorptions(f) 1.6(25/16) 1.4(17/12) 2.1(30/14)
Percent dead fetuses 0.2(1/435) 0(0/369) 0(0/378)
Sex ratio, M:F, % 47:53 47:53 52:48
Fetal body weight, (grams)g 5.60+/-0.36 5.59+/-0.44 5.49+/-0.45
Fetal crown-rump length, (mm)g 42.7+/-1.8 42.3+/-1.4 42.4+/-1.5


(a) Bred rats were exposed to 0, 2.5 or 25 ppm epichlorohydrin by inhalation for 7 hrs/day on days 6 through 15 of gestation.

(b) Number of females with visible implantations at the time of C-section or necropsy/total bred. Among rats exposed to 2.5 ppm epichlorohydrin, 5 animals delivered a litter prior to day 21 of gestation. At the 25 ppm level, a single female delivered early, and 3 control rats delivered prior to gestation day 21.

(c) Number of females detected as being pregnant only after staining uterus with sodium sulfide stain/total stained.

(d) Number of females pregnant by visual inspection of the uterus or by sodium sulfide stain/total bred.

(e) Mean +/- S.D.

(f) Resorptions detected by sodium sulfide staining were not included in these calculations.

(g) Mean of litter means +/- S.D.

No values were significantly different from the control values by the appropriate statistical test, p<0.05.

Effect levels (maternal animals)

open allclose all
Dose descriptor:
NOAEL
Effect level:
2.5 ppm
Basis for effect level:
other: maternal toxicity
Dose descriptor:
LOAEL
Effect level:
25 ppm
Basis for effect level:
other: maternal toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
No adverse effects related to treatment were observed in the embryos or fetuses at any exposure level. Therefore, the fetal NOAEL was 25 ppm.

INCIDENCE OF FETAL ALTERATIONS AMONG LITTERS OF RATS
EXPOSED TO EPICHLOROHYDRIN BY INHALATION(a)

ppm Epichlorohydrin
0 2.5 25
Number Fetuses (Number Litters) 435(36) 369(33) 378(32)
SOFT TISSUE EXAMINATION 147(36) 127(33) 130(32
SKELETAL EXAMINATION 435(36) 369(33) 378(32)
BONES OF THE SKULL(b) 288(33) 242(29) 248(31)
Percent Affected (Number Affected)
Total Major F(e) 1(5) 1(3) 1(3)
Malformations L 11(4) 9(3) 9(3)
External Examination
Short Trunk, F 0.5(2) 0.3(l) 0(0)
Hypoplastic Tail* L 6(2) 3(l) 0(0)
Acaudia* F 0(0) 0(0) 0.3(1)
L 0(0) 0(0) 3(1)
Multiple Facial F 0.2(l) 0(0) 0(0)
Anomalies(d)* L 3(1) 0(0) 0(0)
Inward rotation of F 0(0) 0.3(1) 0(0)
right hind paw* L 0(0) 3(1) 0(0)
Inward rotation of F 0(0) 0(0) 0.3(1)
both hind limbs* L 0(0) 0(0) 3(1)
Subcutaneous Edema F 0.2(l) 0(0) 0(0)
L 3(l) 0(0) 0(0)
Soft Tissue Examination
Dilated Ureter F 3(4) 2(3) 1(1)
(Unilateral or Bilateral) L 6(2) 6(2) 3(1)
Dilated Renal pelvis F 1(1) 0(0) 2(2)
(Unilateral or Bilateral) L 3(1) 0(0) 6(2)
Encephalomeningocele* F 1(1) 0(0) 1(1)
L 3(1) 0(0) 3(1)
Unilateral External F 0(0) 1(1) 0(0)
Hydrocephalus* L 0(0) 3(1) 0(0)
Slightly Dilated F 0(0) 1(1) 0(0)
Ventricle (brain) L 0(0) 3(1) 0(0)
Shortened Left Lateral F 0(0) 0(0) 1(1)
Ventricle (brain)* L 0(0) 0(0) 3(1)
Subcutaneous Hemorrhage F 0(0) 0(0) 1(1)
(head-nasal area) L 0(0) 0(0) 3(1)
Skeletal Examination
Skull - delayed ossification F 15(44) 14(35) 12(30)
L 48(16) 41(12) 58(18)
- bone island F 1(2) 2(4) 1(2)
L 6(2) 14(4) 7(2)
- extra site of F 3(9) 2(6) 1(3)
ossification L 15(5) 14(4) 6(2)
- occipital unfused F 0(0) 0.4(1) 0(0)
and misshapen* L 0(0) 3(1) 0(0)
Sternebrae F 4(18) 5(18) 6(24)
- delayed ossification L 31(11) 39(13) 34(11)
- unfused F 0.2(1) 0.3(1) 0.3(1)
L 3(1) 3(1) 3(1)
Ribs- extra ribs F 1(4) 0.3(1) 0.5(2)
L 11(4) 3(1) 6(2)
- missing* F 0.2(1) 0(0) 0.3(1)
L 3(1) 0(0) 3(1)
- wavy ribs F 1(4) 0.3(1) 0.3(1)
L 11(4) 3(1) 3(1)
Vertebrae - missing* F 1(3) 0.3(1) 0.3(1)
L 8(3) 3(1) 3(1)
- misshapen* F 0.2(1) 0(0) 0(0)
L 3(1) 0(0) 0(0)
- delayed ossification F 25(107) 23(84) 28(105)
of cervical centra L 72(26) 76(25) 78(25)
- bilobed thoracic F 5(22) 7(25) 5(18)
centra L 39(14) 52(17) 28(9)
- unfused thoracic F 0.2(1) 1(3) 1(4)
centra L 3(1) 6(2) 13(4)
- spurs F 18(76) 14(50) 13(48)
L 75(27) 70(23) 59(19)
Other
- Asymmetric pelvis F 0(0) 0(0) 0.3(1)
L 0(0) 0(0) 3(1)
Multiple Skeletal Defects(e)* F 0.2(1) 0(0) 0(0)
L 3(1) 0(0) 0(0)

(a) Bred rats were exposed to 0, 2.5 or 25 ppm of epichlorohydrin by inhalation for 7 hrs/day on days 6 through 15 of gestation.
(b) In litters containing less than 4 fetuses, all fetuses were decapitated during the soft tissue examination and were therefore not available for examination of bones of the skull.
(c) F = Fetuses L = Litters
(d) One fetus in the control group exhibited multiple facial anomalies including the formation of a nasal proboscis, micrognathia of both upper and lower jaws, a small head and facial area, pinnae attached posterior to the normal position, and micropthalmia.
(e) One control fetus exhibited multiple skeletal defects, including misshapen tibias and fibulas; misshapen right femur; misshapen radii and ulnas; unossified centra and delayed ossification of all skull bones.
* Considered to be a major malformation.
No value differed significantly from the control value by a modified Wilcoxon Test, P<0.05.

Effect levels (fetuses)

Dose descriptor:
NOAEL
Effect level:
> 25 ppm
Basis for effect level:
other: teratogenicity

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

Pregnant rats exposed to ECH at 25 pprn had significantly decreased body weights and food consumption during gestation as compared to controls. No evidence of altered appearance or demeanor was observed among exposed rats. Liver weights (absolute and relative) were comparable to controls for rats exposed to ECH. Reproductive parameters including number of pregnant females, number of corpora lutea, number of implantations, litter size and incidence of resorbing fetuses were not adversely affected in rats exposed to ECH. Fetal body weights and measurements in rats were not affected by exposure to ECH. Low incidences of malformations were observed among rat fetuses from both the exposed and the control groups. Single occurrences of acaudia, rotated limb or paw, encephalomeningocele, shortened ventricles of the brain and missing ribs or vertebrae were observed in the 25 ppm group. At 2.5 pprn, single occurrences of shortened trunk, rotated limb or paw, external hydrocephalus, a misshapen occipital bone and missing vertebrae were observed. These malformations have been observed to occur sporadically among control litters of Sprague-Dawley rats in this laboratory. Several malformed fetuses were also observed among the concurrent control fetuses. In some cases 2 or more of the malformations occurred in the same fetus. When considered collectively, the total number of fetuses which had one or more major malformations was less in each of the 2 exposed groups (3 in 3 litters), than in controls where 5 fetuses in 4 litters were malformed.

Applicant's summary and conclusion

Conclusions:
Epichlorohydrin was not embryotoxic, fetotoxic or teratogenic in rats following inhalation exposure to concentrations up to 25 ppm during the period of major organogenesis and at concentrations which induced effects in the maternal animals.
Executive summary:

Pregnant Sprague-Dawley rats were exposed to vapors of epichlorohydrin (ECH) at concentrations of 0, 2.5 or 25 pprn. Exposures were for 7 hr/day on days 6 through 15 (rats) of gestation. Maternal effects including decreased body weight and food consumption were observed among rats inhaling 25 pprn of ECH. No evidence of an adverse effect to the embryo or fetus was observed among rats following exposure to ECH. Thus, ECH was not teratogenic or embryolethal in rats following inhalation exposure to concentrations which induced effects in the maternal animals.