Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Effect on fertility: via oral route
Endpoint conclusion:
no study available
Effect on fertility: via inhalation route
Endpoint conclusion:
no study available
Effect on fertility: via dermal route
Endpoint conclusion:
no study available

Effects on developmental toxicity

Description of key information
A developmental toxicity study performed in the rat with CTF reports maternal and developmental NOAELs of 300 mg/kg bw/d.   
Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
22nd January 2001
Deviations:
no
GLP compliance:
yes
Limit test:
no
Specific details on test material used for the study:
Identity: CTF
Alternative name: Cyclic Trimethylolpropane Formal
Batch no.: 150803652
EC number: 225-967-8
EC name: 5-ethyl-1,3-dioxane-5-methanol
CAS number: 5187-23-5
IUPAC name: (5-ethyl-1,3-dioxan-5-yl)methanol
Expiry date: 12 August 2016
Appearance: Colourless undefined liquid
Purity: 99.8%
Storage conditions: Room temperature, under nitrogen
Species:
rat
Strain:
Sprague-Dawley
Remarks:
Hsd: SpragueDawley SD
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Envigo RMS srl, San Pietro al Natisone (UD), Italy
- Age at study initiation: at least 11 weeks
- Weight at study initiation: 213-221 g
- Fasting period before study: None
- Housing: group
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 18 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 40-70
- Air changes (per hr): 15-20
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 18th January 2016 To: 16th February 2016
Route of administration:
oral: gavage
Vehicle:
water
Details on exposure:
CTF was administered orally by gavage (in water) at a dose volume of 10mL/kg bw; control animals received the vehicle alone at the same dose volume. The dose was administered to each animal on the basis of the most recently recorded body weight and the volume administered was recorded for each animal. All animals were dosed once a day on Gestation Days 6-19.
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
28-hour stability at room temperature and 8-day stability at +4°C were verified for CTF in water at 100 mg/mL; 28- hour stability at room temperature and 8-day were also verified at 10 mg/mL. Concentrations following storage were within acceptable limits (90-110%). The proposed formulation procedure was checked in the range from 10-100 mg/mL by chemical analysis (concentration) during the pre-treatment period to confirm that the method was suitable. Final results for all levels were within acceptable limits (90-110%). Samples of the formulations prepared during the first and final weeks of the were analysed to check the concentration. Results of the analyses were within acceptable limits (90-110%). The analytical method was previously validated (RTC Study A1846) in the range 5-100 mg/mL. Linearity, accuracy and precision were within acceptable limits (r > 0.98; accuracy 90-110%; precision CV < 5%).
Details on mating procedure:
- Impregnation procedure: cohoused
- If cohoused:
- M/F ratio per cage: 1:1
- Length of cohabitation: not reported
- Verification of same strain and source of both sexes: yes
- Proof of pregnancy: presence of sperm in the vaginal smear or by the presence of a copulation plug, was considered as Day 0 of gestation
Duration of treatment / exposure:
All animals were dosed once a day on Gestation Days (GD) 6-19.
Frequency of treatment:
Daily
Duration of test:
All animals were dosed once a day on Gestation Days (GD) 6-19 and terminated on GD20.
Dose / conc.:
100 mg/kg bw/day
Remarks:
Low dose group
Dose / conc.:
300 mg/kg bw/day
Remarks:
Intermediate dose group
Dose / conc.:
1 000 mg/kg bw/day
Remarks:
High dose group
No. of animals per sex per dose:
24 mated females per group
Control animals:
yes, concurrent vehicle
Details on study design:
- Dose selection rationale: dose levels were based on range-finding data
- Rationale for animal assignment: random
Maternal examinations:
Mortality
All animals were checked twice daily.

Clinical signs
Each animal was observed daily from allocation to termination and clinical signs recorded.

Body weight
All animals were weighed on Gestation Days 0, 6, 9, 12, 15 and 20.

Food consumption
Food consumption was measured on Gestation Days 0, 6, 9, 12, 15 and 20. Values were recorded per cage and the amount was divided for
the animals in the cage and for the days of the interval of measurement, in order to obtain the value of food consumed by animal per day.

Animals were euthanised with carbon dioxide on Gestation Day 20 and subjected to gross necropsy.
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: gross evaluation of placentae
Fetal examinations:
All live foetuses were examined externally. Approximately one-half of the foetuses in each litter was preserved in Bouin’s solution for subsequent fixed-visceral examination. The remaining foetuses were eviscerated after which the carcasses were fixed in 95% (v/v) ethanol for subsequent skeletal examination. Skeletal and fixed-visceral examinations were performed in all groups.
Statistics:
For continuous variables the significance of the differences amongst group means was assessed by Dunnett’s test or a modified t test, depending on the homogeneity of data. Statistical analysis of non-continuous variables was carried out by means of the Kruskal-Wallis test and intergroup differences between the control and treated groups assessed by a non-parametric version of the Williams test.
Indices:
Pre-implantation loss, pos-implantation loss and total implantation loss were calculated.
Historical control data:
Not referenced.
Clinical signs:
no effects observed
Mortality:
no mortality observed
Body weight and weight changes:
effects observed, treatment-related
Description (incidence and severity):
A slight (but statistically significant) decrease in bodyweight was seen in mid-dose females on GD15 (approximately -3%) and in high dose females from GD9-20 (up to approximately -5%), compared to controls. These changes were minimal and not considered to be of toxicological relevance. Weight gain was significantly reduced in the high dose group on GD 9 and 12, compared to controls.
Food consumption and compound intake (if feeding study):
effects observed, treatment-related
Description (incidence and severity):
Statistically significant decreases were detected in food consumption of mid-dose (up to approximately -12%) and high dose females (up to approximately -25%) from GD9-15, compared to controls.
Food efficiency:
not examined
Water consumption and compound intake (if drinking water study):
not examined
Ophthalmological findings:
not examined
Haematological findings:
not examined
Clinical biochemistry findings:
not examined
Urinalysis findings:
not examined
Behaviour (functional findings):
not examined
Immunological findings:
not examined
Organ weight findings including organ / body weight ratios:
effects observed, treatment-related
Description (incidence and severity):
A statistically significant decrease in uterus weight (-12%) was observed in females of the high dose group, compared to controls.
Gross pathological findings:
no effects observed
Neuropathological findings:
not examined
Histopathological findings: non-neoplastic:
not examined
Histopathological findings: neoplastic:
not examined
Other effects:
no effects observed
Details on results:
No deaths occurred and no signs of maternal toxicity were observed. Marginal (but statistically significant) and transient effects on mean bodyweight were observed at 300 and 1000 mg/kg bw/d; a significant reduction in weight gain was seen at 1000 mg/kg bw/d. Food consumption was also reduced at 300 and 1000 mg/kg bw/d.
Number of abortions:
no effects observed
Pre- and post-implantation loss:
no effects observed
Total litter losses by resorption:
effects observed, non-treatment-related
Description (incidence and severity):
In the high dose group one female showed unilateral implantation with total resorption. This single incidence is not considered to be related to treatment.
Early or late resorptions:
no effects observed
Dead fetuses:
no effects observed
Changes in pregnancy duration:
not examined
Description (incidence and severity):
Migrated Data from removed field(s)
Field "Effects on pregnancy duration" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsMaternalAnimals.MaternalDevelopmentalToxicity.EffectsOnPregnancyDuration): not examined
Changes in number of pregnant:
effects observed, non-treatment-related
Description (incidence and severity):
A total of two females were found not pregnant at necropsy: one in the low dose group and one in the mid-dose group. FIndings are not related to treatment.
Other effects:
no effects observed
Details on maternal toxic effects:
A total of two females were found not pregnant at necropsy: one in the low dose group and one in the mid-dose group. FIndings are not related to treatment. In the high dose group one female showed unilateral implantation with total resorption. This single incidence is not considered to be related to treatment. A statistically significant decrease in terminal body weight (-6%), uterus weight (-12%), and absolute weight gain (-19%) was observed in females of the high dose group, compared to controls. A slight statistically significant decrease was also observed in the absolute weight gain of the mid-dose group (-10%), with respect to controls. The decrease in absolute weight gain in mid- and high dose females was an indication of a slight treatment and dose-related maternal toxicity; the slight decrease in uterus weight in high dose females was also associated with the lower mean foetal weight.
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day
Based on:
test mat.
Basis for effect level:
body weight and weight gain
food consumption and compound intake
Remarks on result:
other:
Remarks:
No toxicologically significant effects were seen at the mid-dose level
Key result
Abnormalities:
no effects observed
Fetal body weight changes:
effects observed, treatment-related
Description (incidence and severity):
A slight (but statistically significant) decrease was seen for mean foetal weight (approximately -6%) at the high dose level.
Migrated Data from removed field(s)
Field "Fetal/pup body weight changes" (Path: ENDPOINT_STUDY_RECORD.DevelopmentalToxicityTeratogenicity.ResultsAndDiscussion.ResultsFetuses.FetalPupBodyWeightChanges): not examined
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):
A slight (but statistically significant) decrease was seen for mean foetal weight (approximately -6%) and mean litter weight (approximately -13%) at the high dose level.
Changes in postnatal survival:
not examined
External malformations:
no effects observed
Skeletal malformations:
no effects observed
Visceral malformations:
effects observed, non-treatment-related
Description (incidence and severity):
The following malformations were detected:

- hepatic lobe agenesis seen in 3 foetuses in the control group and 1 foetus in the low dose group
- extreme renal pelvic dilatation in 3 foetuses in the control group and 1 foetus in the high dose group
- extreme enlargement of the ureters in 1 foetus in the control group and 1 foetus in the high dose group
- kinked ureters in 1 foetus in high dose group

Since these findings were seen only in few foetuses, both in the control and high dose groups with similar incidence, they were to be considered incidental.
No toxicological significance was attributed to the other findings observed, since they were seen in treated as well as in control groups with similar incidence.
Other effects:
effects observed, treatment-related
Description (incidence and severity):
Incomplete ossification or no ossification of sternal elements (5th and 6th), as well as incomplete ossification of the centrum of the thoracic vertebrae, unossified metacarpals (4th), were seen in all groups with a higher incidence in the high dose group. This higher incidence could be associated with the decreases in mean foetal weight and litter weight observed in the high dose group.
Details on embryotoxic / teratogenic effects:
No evidence for embryotoxicity or teratogenicity.
Key result
Dose descriptor:
NOAEL
Effect level:
300 mg/kg bw/day
Based on:
test mat.
Sex:
male/female
Basis for effect level:
fetal/pup body weight changes
other: Reduced foetal skeletal ossification
Key result
Abnormalities:
no effects observed
Key result
Developmental effects observed:
yes
Lowest effective dose / conc.:
1 000 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:
yes

Summary of maternal and litter findings:

Dose level (mg/kg bw/d)

0

100

300

1000

Mated (#)

24

24

24

24

Not pregnant (#)

-

1

1

-

Total resorption (#)

-

-

-

1

Litters (#)

24

23

23

23

Weight (g)

Day 9

273.97

271.92

269.03

265.58*

Day 12

289.07

286.59

282.57

277.45**

Day 15

308.98

306.41

300.15*

296.33**

Day 20

382.17

379.90

375.64

361.97**

Weight gain (g/d)

Day 6-9

3.903

3.006

2.777

1.483**

Day 9-12

5.035

4.992

4.514

3.958*

Weight gain (g)

Day 6-9

11.71

9.02

8.33

4.45

Day 0-20

145.77

143.69

140.94

127.31

Day 6-20

119.91

117.00

114.94

100.84

Food consumption (g/d)

Day 9

21.24

20.92

18.74*

15.98**

Day 12

22.54

22.08

20.29**

17.91**

Day 15

22.96

21.97

20.49**

19.53**

Terminal bodyweight (g)

378.88

375.16

370.35

356.08*

Absolute weight gain (g)

64.01

59.95

57.87*

52.12*

Gravid uterus weight (g)

78.45

79.00

77.77

69.30*

Corpora lutea (#)

14.79

14.39

14.26

13.43

Pre-implantation loss (%)

1.90

1.72

1.71

0.64

Implantations (#)

14.50

14.13

14.00

13.35

Post-implantation loss (%)

5.74

2.67

3.30

7.02

Foetuses (#)

13.67

13.74

13.52

12.52

Foetal weight (g)

3.83

3.79

3.75

3.60*

Litter weight (g)

52.24

52.07

50.68

45.19*

*significantly different to control (p<0.05); **p<0.01)

Summary of foetal findings:

Dose level (mg/kg bw/d)

0

100

300

1000

Hepatic lobe agenesis

3(3)

1(1)

-

-

Renal pelvis dilation (extreme)

1(1)

-

-

1(1)

Ureter enlarged (extreme)

1(1)

-

-

1(1)

Kinked ureter (extreme)

-

-

-

1(1)

5thsternebra: no ossification

1(1)

2(2)

3(3)

4(4)

5thsternebra: incomplete ossification

21(9)

15(11)

11(10)

21(12)

6thsternebra: no ossification

-

1(1)

1(1)

1(1)

6thsternebra: incomplete ossification

15(8)

11(7)

15(10)

26(13)

Centrum: no ossification

2(2)

1(1)

-

-

Centrum: incomplete ossification

10(5)

11(8)

9(6)

23(14)

Conclusions:
No evidence of teratogenicity was seen in this study. Findings were limited to slight maternal toxicity (reduced weight gain and food consumption) and slight foetal effects (reduced foetal weight, reduced skeletal ossification at the high dose level). The study report concludes that none of the effects seen were of toxicological significance; however maternal and developmental NOAELs of 300 mg/kg bw/d are determined for this study.
Executive summary:

Groups of 24 mated female SD rats were gavaged with CTF (in water) at dose levels of 0 (vehicle control), 100, 300 or 1000 mg/kg bw/d on Gestation Days 6-19. No deaths occurred and no signs of toxicity were observed during the study period. Mean bodyweights of high dose females were slightly (but statistically significantly) lower than controls at Day 9 (-3.1%), Day 12 (-4.0%), Day 15 (-4.1%) and Day 20 (-5.3%); the mean bodyweight of mid-dose females was significantly lower (-2.9%) at Day 15 only. Maternal weight gain at the high dose level was lower over the treatment period (-16%) and over the study period (-13%); weight gain over the initial part of the treatment period was also lower in this group. Reduced food consumption was seen at the high dose level and, to a lesser extent, at the mid-dose level. Gravid uterus weight was significantly lower at the high dose level (-12%); however this may be due, in part, to the slightly smaller litter size in this group. Single females in the low- and mid-dose groups were found not to be pregnant. Total resorption of implantations was seen in one female at the high dose level; this finding is not considered to be treatment-related due to the low incidence. Mean litter size at the high dose level was slightly lower than the other groups; however this was a consequence of the lower numbers of corpora lutea and is not related to treatment. Mean foetal weight was slightly (-6%) but significantly lower than controls; mean litter weight in this group was significantly lower (-13.5%) due to a combination of the smaller litter size and lower foetal weight. There was no effect of treatment on the incidence or pattern of foetal malformations. A slight increase in the incidence of foetal skeletal variations (reduced or absent ossification of sternebra(e), vertebral centra and/or metacarpals) was seen in the high dose group. Findings are likely to be secondary to reduced foetal weight and represent a slight developmental delay, secondary to maternal toxicity. A maternal NOAEL of 300 mg/kg bw/d is determined for this study based on reduced weight gain at the highest dose level. Although effects on weight gain and food consumption were seen at the mid-dose level, the magnitude of change was not great and findings are not considered to be of toxicological significance. A developmental NOAEL of 300 mg/kg bw/d is determined for this study, based on the reduced foetal weight and skeletal ossification seen at the highest dose level.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
adverse effect observed
Dose descriptor:
NOAEL
300 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
A high quality modern GLP- and guideline-compliant study is available for CTF in the rat.
Effect on developmental toxicity: via inhalation route
Endpoint conclusion:
no study available
Effect on developmental toxicity: via dermal route
Endpoint conclusion:
no study available
Additional information

No evidence of teratogenicity was seen in an oral developmental toxicity study with CTF. Findings were limited to slight maternal toxicity (reduced weight gain and food consumption) and slight foetal effects (reduced foetal weight, reduced skeletal ossification at the high dose level). Maternal and developmental NOAELs of 300 mg/kg bw/d are therefore determined for this study.

Justification for classification or non-classification

Evidence for the developmental toxicity of CTF was seen in the rat study, but is limited to slightly reduced foetal skeletal ossification in the presence of maternal toxicity. Findings are not sufficient to classify CTF for reproductive (pre-natal developmental) toxicity according to the CLP Regulation.