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Toxicological information

Developmental toxicity / teratogenicity

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Administrative data

Endpoint:
developmental toxicity
Remarks:
also includes potential effect on fertility
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Comparable to guideline study with acceptable restrictions.
Justification for type of information:
Organ weights were determined and full histopathology was performed on male (epididymides, testes, prostate, seminal vesicles) and female (ovaries, non-gravid uterus) gonads. These data can be used to conclude on potential effects on fertility (Assessment on Fertility of Dioxolane based on a Developmental Toxicity Study in Rats conducted by Hoberman (1991) by Dr. Jochen Bushmenn (2018), see attachment in Section 13.2..

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report Date:
1991

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
The scheme for dose administration (day 6 through 15 of presumed gestation) agreed with OECD 414 (1981), but not with OECD 414 (2001), which states day 5 post mating to the day prior to scheduled caesarian section.
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): 1,3-dioxolane
- Physical state: liquid
- Analytical purity: 99.9%
- Lot/batch No.: UN 1166

Test animals

Species:
rat
Strain:
other: Crl:CD®BR VAF/Plus®
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Breeding Laboratories, Inc., Raleigh, North Carolina, USA
- Age at study initiation: approximately 12 weeks
- Weight at study initiation: 240-279 g
- Fasting period before study: none
- Housing: individually housed in wire-bottomed stainless steel cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: two weeks
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 70-78°F (equivalent to 21-26°C).
- Humidity (%):targeted 40-70% (minor deviations occurred)
- Air changes (per hr): protocol stated: at least 10 changes per hour
- Photoperiod (hrs dark / hrs light): fluorescent light, 12 hours light, 12 hours dark
IN-LIFE DATES: From: July 10-13, 1990 To: July 30-August 02, 1990

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
Dose levels were 0, 125, 250, 500 and 1000 mg/kg bw/day, administered from day 6 through 15 of presumed gestation (female rats with spermatozoa in vaginal lavage or a copulatory plug in situ were considered to be at day 0 of presumed gestation).
VEHICLE
- Justification for use and choice of vehicle (if other than water): not provided
- Concentration in vehicle: 0, 25, 50, 100 and 200 mg/mL
- Amount of vehicle (if gavage): 5 mL/kg bw
- Lot/batch no. (if required): no data
- Purity: pure
Details on analytical verification of doses or concentrations:
Dosing solutions were prepared weekly. Samples from each test solution were taken for analytical verification of the accuracy and stability of the 1,3 dioxolane concentrations. The samples were dissolved in hexane, diluted and analysed by GC/FID. Validation for this method was submitted. Mean recovery (accuracy) at 25 and 500 mg/g, respectively, was 97.7% and 97.5% (RSD 0.8% and 6.0%, n = 10 and n=9). Precision (% RSD) at 25 and 200 mg/mL, respectively, was 1.0% and 1.1% (n = 5 at both levels). A 7 point calibration gave a coefficient of correlation of 0.99996.
Details on mating procedure:
- Impregnation procedure: cohoused. On July 9, 1990, 160 virgin females were cohoused with 160 males. On July 10, 1990, the remaining 20 virgin females were cohoused with male rats that had mated during the first night of cohabitation. In all cases one male was cohoused with one female. Day 0 of presumed gestation occurred on July 10 through July 13, 1990, for the 125 female rats that were assigned to the study.
Duration of treatment / exposure:
Dose levels were 0, 125, 250, 500 and 1000 mg/kg bw/day, administered from day 6 through 15 of presumed gestation (female rats with spermatozoa in vaginal lavage or a copulatory plug in situ were considered to be at day 0 of presumed gestation).
Frequency of treatment:
See above.
Duration of test:
21 days (Caesarian section was performed on day 20 of presumed gestation)
Doses / concentrationsopen allclose all
Dose / conc.:
0 mg/kg bw/day
Dose / conc.:
125 mg/kg bw/day
Dose / conc.:
250 mg/kg bw/day
Dose / conc.:
500 mg/kg bw/day
Dose / conc.:
1 000 mg/kg bw/day
No. of animals per sex per dose:
25 presumed pregnant females per dose
Control animals:
yes, concurrent vehicle

Examinations

Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily on day 0 through 5 of presumed gestation; immediately prior to intubation and about 1 hour postdosage on day 6 through 15 of presumed gestation; once daily from day 16 through 20 of presumed gestation.
- Cage side observations checked in table were included.
DETAILED CLINICAL OBSERVATIONS: No
BODY WEIGHT: Yes
- Time schedule for examinations: on day 0 and day 6 through 20 of presumed gestation
FOOD CONSUMPTION AND COMPOUND INTAKE (if feeding study): Yes
- Time schedule for examinations: on day 0 and day 6 through 20 of presumed gestation
WATER CONSUMPTION AND COMPOUND INTAKE (if drinking water study): No
POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: uterus was excised and weighed; thoracic and abdominal cavities were examined for gross lesions.
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: numbers of live and dead fetuses were recorded.
Fetal examinations:
- External examinations: Yes: [all per litter]; fetuses were also weighed and sex was determined
- Soft tissue examinations: Yes: [half per litter]
- Skeletal examinations: Yes: [half per litter]
- Head examinations: No
Statistics:
Maternal and fetal incidence data: Variance test for homogeneity of the binomial distribution.
Maternal body weights, body weight changes, gravid uterine weights, feed consumption values, litter averages for fetal body weights, % male fetuses, % resorbed conceptuses per litter, fetal ossification sites, % fetal alterations: Bartlett’s test of homogeneity; if homogeneous, analysis of variance, followed by a Dunnett’s test; if not homogeneous, the Kruskal-Wallis test followed by Dunn’s test.
All other Caesarion section data: Kruskal-Wallis test followed by Dunn’s test.
All tests at 5% or 1% significance level.
Historical control data:
Historical control data were provided in the report.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:
not specified
Dermal irritation (if dermal study):
no effects observed
Mortality:
no mortality observed

Maternal developmental toxicity

Details on maternal toxic effects:
Maternal toxic effects:yes

Details on maternal toxic effects:
Maternal toxicity was observed at 500 and 1000 mg/kg bw and consisted of reduced food consumption and reduced body weight gain.

Effect levels (maternal animals)

Dose descriptor:
NOAEL
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Basis for effect level:
other: developmental toxicity

Results (fetuses)

Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Adverse treatment related changes on litters and foetuses were only observed at maternally toxic doses: reduced foetal weight at 1000 mg/kg; increased incidence of tail malformations and ventricular septal defects at 1000 mg/kg; increased incidence of vertebral malformations including bifid centra in thoracic vertebrae at 500 and 1000 mg/kg, and reduced ossification of centra in the thoracic and lumbar vertebrae, absent sacral and caudal vertebrae and rib-vertebral malformations, all at 1000 mg/kg; reduced number of ossified metacarpals per foetus at 500 and 1000 mg/kg.

Effect levels (fetuses)

Key result
Dose descriptor:
NOAEL
Effect level:
250 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: Developmental NOAEL: reduced ossification (increased incidence of bifid centra in the thoracic vertebrae and reduced number of ossified metacarpals per foetus) is 250 mg/kg bw/d. Maternal NOAEL (250 mg/kg): reduced food consumption and body weight gain.

Fetal abnormalities

Abnormalities:
not specified

Overall developmental toxicity

Developmental effects observed:
not specified

Any other information on results incl. tables

Concentrations of 1,3 -dioxolane in the dosing samples were 99-114% of the target concentration. For the samples used for the first week of dosing, at 25, 50, 100 and 200 mg/mL the concentrations after seven days of storage were 77%, 92%, 94% and 96% of the initial concentrations. For the samples used for the second week of dosing, at 25, 50, 100 and 200 mg/mL the concentrations after seven days of storage were 96%, 100%, 98% and 98% of the initial concentrations.

The number of pregnant rats ranged between 23 and 25 out of 25 rats tested. The remaining biological results are summarised in the Table below. The changes are represented either as percentage change relative to the control (e..g -35 = 35% reduction, +10 = 10% increase) or as incidence (e.g. 3/10 represents 3 out of 10 rats).  Where a dose relationship was observed, this is indicated in the last column.

Under the table with findings, a discussion of the study results is provided and the NOAEL is derived.

Table 1. Results from oral developmental toxicity study with 1,3-dioxolane

dose (mg/kg bw/d)

0

125

250

500

1000

DR

Mortality

none

 

Clinical signs

no treatment related findings

 

Gravid uterine weight

no treatment related findings

 

Body weight gain

 

 

 

 

 

 

   day 6-7(A)

 

 

 

-147

-260*

DR

   day 6-16

 

 

 

 

-11

 

Food consumption

 

 

 

 

 

 

   day 6-7

 

 

-14

-16*

-18*

DR

   day 12-16

 

 

+9*

 

 

 

   day 6-16

 

 

 

 

-8*

 

Necropsy

no treatment related findings

 

Litter observations

 

 

 

 

 

 

   foetal body weight

 

 

 

 

-7*

 

Foetal examinations(B)

 

 

 

 

 

 

   Gross external alterations

 

 

 

 

 

 

       Tail malformations(C)    

0/25 (0/405)

 

 

 

7/24* (8/376)*

 

   Soft tissue alterations

 

 

 

 

 

 

       Heart, ventricular septal defect

 

 

 

 

3/24* (3/180)*

 

   Skeletal alterations

 

 

 

 

 

 

       Vertebral scrambling

3/25 (4/208)

 

 

7/23 (11/187)

16/24* (36/196)*

DR

              Thoracic, centra, bifid(D)

2/25 (3/208)

 

 

7/23 (11/187)

12/24* (25/196)*

DR

       Pelvis, pubes incompletely ossified

7/25 (12/208)

4/24 (4/199)*

2/25 (3/197)*

3/23 (4/187)*

2/24 (2/196)*

 

       Ossification sites per foetus per litter

 

 

 

 

 

 

              Sternal centers

 

 

+8*

 

 

 

              Metacarpals

 

 

 

-5.2*

-5.8*

 

DR          dose related

*              Statistically significant difference from control at 5% level or below

 (A)         At 500 and 1000 mg/kg, respectively, a weight loss 0.7 and 2.4 g was observed, compared to a weight gain of 1.5 g in the control.

(B)          Data format a/b (c/d), where a = the number of litters affected; b = the number of litters evaluated; c = the number of foetuses affected; d = the number of foetuses evaluated. Empty cells indicate that the finding was not observed.

(C)          The malformations included constrictions of the tail and tail absent (each in 1 foetus and 1 litter), tail short (2 foetuses, 2 litters) and tail threadlike (4 foetuses and 4 litters, both statistically significant at 1% level).

(D)          Bifid centra in thoracic vertebrae was the only finding at 500 mg/kg. Apart from bifid centra in thoracic vertebrae, at 1000 mg/kg the scrambling included reduced ossification of centra in the thoracic vertebrae (asymmetric, bifid (statistically significant at 1% level), unilateral or absent ossification (both statistically significant at 1% level), fused centrum and arch), the lumber vertebrae (fused centrum and arch, centra that were bifid (statistically significant at 1% level) or not ossified, and arches that were small or not ossified)), absent sacral and caudal (statistically significant at 1% level) vertebrae and rib-vertebral malformations.

(E)          Data format: % change relative to control.

Evaluation

Increased food consumption of dams at 250 mg/kg bw is considered to be a fortuitous finding in the absence of similar effects at higher dose levels. The reduction of the number of foetuses with incompletely ossified pubes at 125-1000 mg/kg is considered to be unrelated to the treatment (no dose relationship, no adverse effect). The accelerated sternal ossification at 250 mg/kg bw is also considered to be unrelated to the treatment (no dose relationship, no adverse effect). 

 

Maternal toxicity was observed at 500 and 1000 mg/kg bw (reduced food consumption and body weight gain). The slightly reduced food consumption (14%) at 250 mg/kg is not considered to be toxicologically relevant in the absence of an effect on body weight gain at that dose. The maternal NOAEL is 250 mg/kg based reduced food consumption and body weight gain.

 

Adverse treatment related changes on litters and foetuses were only observed at maternally toxic doses: reduced foetal weight at 1000 mg/kg; increased incidence of tail malformations and ventricular septal defects at 1000 mg/kg; increased incidence of vertebral malformations including bifid centra in thoracic vertebrae at 500 and 1000 mg/kg, and reduced ossification of centra in the thoracic and lumbar vertebrae, absent sacral and caudal vertebrae and rib-vertebral malformations, all at 1000 mg/kg; reduced number of ossified metacarpals per foetus at 500 and 1000 mg/kg.

 

The only treatment related developmental effects at 500 mg/kg were increased incidence of bifid centra in thoracic vertebrae and reduced number of ossified metacarpals per foetus. The author of the report claimed that the NOAEL for developmental toxicity was 500 mg/kg. The author of the report did not mention or discuss the increased incidence of bifid centra in thoracic vertebrae at 500 mg/kg. The incidence of this finding at 500 mg/kg, however, although not statistically significant, is clearly increased relative to the concurrent control, and also relative to the reported historical control data. Moreover, a dose-relationship was observed in the range 500-1000 mg/kg. Therefore this finding is considered to be an effect of the treatment. The author of the report stated that the reduced number of ossified metacarpals per foetus at 500 mg/kg was not biologically important and unrelated to the treatment for the following reasons: (1) no other statistically significant delays in ossification occurred at 500 mg/kg; (2) the values for the 500 and 1000 mg/kg did not reflect the difference in the dosage. (3) The statistical significance at 500 mg/kg reflected the relatively small standard deviation for this group, compared to the other groups. However, the increased incidence of bifid centra in thoracic vertebrae represented other evidence of reduced ossification at 500 mg/kg.

 

The developmental NOAEL based on reduced ossification (increased incidence of bifid centra in thoracic vertebrae and reduced number of ossified metacarpals per foetus) is 250 mg/kg bw/d.

Applicant's summary and conclusion

Conclusions:
The maternal NOAEL is 250 mg/kg based reduced food consumption and body weight gain. The developmental NOAEL based on reduced ossification (increased incidence of bifid centra in thoracic vertebrae and reduced number of ossified metacarpals per foetus) is 250 mg/kg bw/d.
Executive summary:

An oral developmental toxicity study on 1,3-dioxolane was performed in presumed pregnant Crl:CD®BR rats (25 rats/dose) according to OECD 414 (1981) at dose levels of 125, 250, 500 and 1000 mg/kg bw/day, administered from day 6 through 15 of presumed gestation (female rats with spermatozoa in vaginal lavage or a copulatory plug in situ were considered to be at day 0 of presumed gestation). The test material was administered in corn oil (dose volume 5 mL/kg). Corn oil controls were included. Observations on toxicological signs of dams were made daily, and body weights and food consumption of dams were measured on on day 0 and day 6 through 20 of presumed gestation. Dams were sacrificed on gestation day 20, the uterus was excised and weighed, and thoracic and abdominal cavities were examined for gross lesions. Intra-uterine and fetal examinations were conducted according to OECD 414. Maternal toxicity was observed at 500 and 1000 mg/kg bw (reduced food consumption and body weight gain). The maternal NOAEL is 250 mg/kg. Adverse treatment related changes on litters and foetuses were only observed at maternally toxic doses: reduced foetal weight at 1000 mg/kg; increased incidence of tail malformations and ventricular septal defects at 1000 mg/kg; increased incidence of vertebral malformations including bifid centra in thoracic vertebrae at 500 and 1000 mg/kg, and reduced ossification of centra in the thoracic and lumbar vertebrae, absent sacral and caudal vertebrae and rib-vertebral malformations, all at 1000 mg/kg; reduced number of ossified metacarpals per foetus at 500 and 1000 mg/kg. The developmental NOAEL based on reduced ossification (increased incidence of bifid centra in thoracic vertebrae and reduced number of ossified metacarpals per foetus) is 250 mg/kg bw/d.