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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
Additional information

A screening study for reproductive/developmental toxicity is not available. Reliable data has been included to cover toxicity to reproduction, as set out in Column 2 of Annex VIII, Section 8.7.1, Regulation (EC) No 1907/2006. A pre-natal developmental toxicity study performed with a source substance according to OECD 414 is available (Fabreguettes, 2000). No treatment-related effects were observed on any of the assessed reproductive and developmental parameters up to and including the highest dose level of 300 mg/kg bw/day. Thus, providing a screening study for reproductive/developmental toxicity is not necessary as the data requirements are met according to Column 2 of Annex VIII, Section 8.7.1, Regulation (EC) No 1907/2006.


Justification for selection of Effect on fertility via oral route:
No study required since a a pre-natal developmental toxicity study performed according to OECD guideline 414 is available. Referring to Column 2 of Annex VIII, Section 8.7.1, Regulation (EC) No 1907/2006, the required data is present and it is not necessary to provide a screening study for reproductive/developmental toxicity.

Effects on developmental toxicity

Description of key information
Oral: OECD 414, rat, NOAEL development ≥ 300 mg/kg bw/day
Oral: OECD 414, rat, NOAEL maternal systemic = 100 mg/kg bw/day
Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
23 Aug - 7 Sept 2000
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP-guideline study with acceptable restrictions. Dosing was performed on gestation day 6-17, 10 rats per dose group, no justification for the selected dose groups, analytical purity of the test substance was not specified.
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Deviations:
yes
Remarks:
dosing performed on gestation day 6-17, 10 rats per dose group, no justification for the selected dose groups
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
other: Sprague-Dawley Crl CD(SD) IGS BR
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River France, Saint Aubin-lès-Elbeuf, France
- Age at study initiation: approximately 11 weeks
- Weight at study initiation: 272 g (mean)
- Housing: the animals were housed individually in suspended wire-mesh cages (43.0 cm x 21.5 cm x 18.0 cm). A metal tray containing autoclaved sawdust (SICSA, Alfortville, France) was placed under each cage.
- Diet: A04 C pelleted diet, batch No. 90528 (U.A.R., Villemoisson-sur-Orge, France), ad libitum
- Water: tap water filtered using a 0.22 micron filter, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 ± 2
- Humidity (%): 50 ± 20
- Air changes (per hr): approximately 12
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 17 Aug 2000 To: 7 Sept 2000
Route of administration:
oral: gavage
Vehicle:
corn oil
Details on exposure:
PREPARATION OF DOSING SOLUTIONS: the solutions were prepared daily prior to dosing and homogenised using a magnetic stirrer. The solutions were stirred continously during the dosing procedure. The doses were adjusted according to the most recently recorded body weight.

VEHICLE
- Concentration in vehicle: 10, 33.33 and 100 mg/mL
- Amount of vehicle (if gavage): 3 mL/kg bw
- Lot/batch no. (if required): 107H1649 (Sigma, France)
Analytical verification of doses or concentrations:
no
Details on mating procedure:
- Impregnation procedure: purchased timed pregnant
- Proof of pregnancy: vaginal plug referred to as day 0 of pregnancy
Duration of treatment / exposure:
Day 6-17 of gestation
Frequency of treatment:
Daily, 7 days/week
Duration of test:
12 days, Day 6-20 of gestation
Remarks:
Doses / Concentrations:
30, 100 and 300 mg/kg bw/day
Basis:
other: nominal as gavage
No. of animals per sex per dose:
10 P females
Control animals:
yes, concurrent vehicle
Maternal examinations:
CAGE SIDE OBSERVATIONS: Yes
- Time schedule: twice daily for mortality and signs of morbidity, at least once daily for clinical signs
- Cage side observations included: evidence of absorption/resorption, signs of toxicity

DETAILED CLINICAL OBSERVATIONS: No

BODY WEIGHT: Yes
- Time schedule for examinations: gestation day 2 (prior to administration), 6, 9, 12, 15, 18 and 20 (prior to sacrifice). The net body weight change was calculated as (body weight on day 20 minus body weight on day 2) minus weight of gravid uterus.


FOOD CONSUMPTION: Yes
- Food consumption for each animal determined and mean daily diet consumption calculated as g food/kg body weight/day: Yes

WATER CONSUMPTION: No

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day 20
- Organs examined: ovaries, uterus, placenta, thoracic organs, abdominal organs
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: number and distribution of dead and live foetuses
Fetal examinations:
- External examinations: Yes: all per litter (sex, body weight, external anomalities and malformations)
- Soft tissue examinations: Yes: half per litter (stored in Bouin's fluid)
- Skeletal examinations: Yes: half per litter (eviscerated, skeletons stained with alizarin red S)
- Head examinations: No
Statistics:
Mean values were compared by one-way analysis of variance and Dunnett's test. Percentage values were compared by Fisher's exact probability test.
Indices:
Preimplantation loss:
((Number of corpora lutea - Number of implantation sites) / Number of corpora lutea) x 100

Post-implantation loss:

((Number of implantation sites - Number of live fetuses) / Number of implantation sites) x 100

Fetal findings:

(Number of fetuses with a particular finding/ total number of fetuses examined) x 100
Details on maternal toxic effects:
Maternal toxic effects:yes. Remark: 300 mg/kg bw/day: reduced body weight gain

Details on maternal toxic effects:
Systemic effects:
1/10 females in the control group died on gestation day 7. No clinical signs were observed prior to death, but during the histopathological examination the lungs were reddish and dilated. This may have been caused by mis-dosing, although the cause was not established.

In 1/10 rats in the high-dose group, piloerection, abdominal breathing and chromorhinorrea were observed on gestation day 8 to 11, indicating a poor health condition.

The female that showed clinical signs of toxicity also had a body weight loss of approximately 10% from gestation day 6 to 12. The body weight in females in the high-dose group was significantly reduced on day 15 and 18 (see Table 1 under 'Any other information on results incl. tables'). The body weight gain was significantly reduced during gestation day 6 to 18, indicating an effect during the entire dosing period (see Table 2 under 'Any other information on results incl. tables'). The total body weight gain was approximately 25% lower than in the control group. The food consumption was reduced in the high-dose group on gestation day 6-18, compared with the control group, although not statistically significant.

The female exhibiting clinical signs had brownish content of the uterus and dilatation of the cervix. The effects may be toxicologically relevant. No other treatment-related effects were noted.

Fertility effects:
There was no statistically significant effect on the fertility index, corpora lutea, resorptions, pre-implantation loss, post-implantation loss, fetal viability (in utero), gravid uterus weight and number of fetuses.

The slight, but not significant, increase in resorptions and pre-implantation losses observed in the high-dose group is attributable to a single female that was in poor health (14/37 pre-implantation losses). One female in the high-dose group exhibited clinical signs, had body weight loss and histopathological findings. The same female had several resorptions and dead fetuses. This is considered to be a consequence of the poor maternal health condition and is not a reproductive effect. All results were within the historical data range (see Table 6 under 'Any other information on results incl. tables').
Dose descriptor:
LOAEL
Effect level:
300 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Dose descriptor:
NOAEL
Effect level:
100 mg/kg bw/day (actual dose received)
Based on:
test mat.
Basis for effect level:
other: maternal toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
There were no statistically significant difference in post-implantation loss, embryo survival, sex ratio, body weight, malformations and abnormalities beween the control group and the treatment groups (see Table 4 and 5).

The slight, but not significant, increase in post-implantation losses observed in the high-dose group is attributable to a single female (7/12 post-implantation losses) that was in poor health. This is considered to be a consequence of the poor maternal health condition and is not a reproductive effect. All results were within the historical data range (see Table 6 under 'Any other information on results incl. tables').
Dose descriptor:
NOAEL
Effect level:
>= 300 mg/kg bw/day (nominal)
Based on:
test mat.
Sex:
male/female
Basis for effect level:
other: developmental toxicity
Abnormalities:
not specified
Developmental effects observed:
not specified

 

Table 1: Body weights, g (mean ± SD)

Dose group

Control

30 mg/kg bw/day

100 mg/kg bw/day

300 mg/kg bw/day

Gestation day 2

250 ± 14

248 ± 13

248 ± 12

247 ± 15

Gestation day 6

272 ± 15

272 ± 13

272 ± 12

271 ± 14

Gestation day 9

287 ± 18

284 ± 17

280 ± 17

273 ± 26

Gestation day 12

311 ± 21

305 ± 16

295 ± 28

285 ± 31

Gestation day 15

330 ± 24

324 ± 19

315 ± 22

303 ± 19*

Gestation day 18

370 ± 28

365 ± 22

348 ± 28

332 ± 25**

Gestation day 20

400 ± 31

400 ± 26

385 ± 26

370 ± 25

*d-ANOVA + Dunnett-test, p < 0.05

**d-ANOVA + Dunnett-test, p < 0.01

 

 

Table 2: Body weight gain, g (mean % change)

Dose group

Control

30 mg/kg bw/day

100 mg/kg bw/day

300 mg/kg bw/day

Day 2 to 6

9.1

9.6

9.8

9.9

Day 6 to 9

5.0

4.2

3.1

0.6

Day 9 to 12

8.3

7.5

5.3

4.7

Day 12 to 15

6.3

6.3

7.0

6.8

Day 15 to 18

12.0

12.5

10.5

9.6

Day 18 to 20

8.2

9.7

10.8

11.6

Day 2 to 20

60.2

61.0

55.7

50.2*

Day 6 to 18

35.4

34.0

28.1

22.7**

Day 6 to 12

13.7

12.0

8.6

5.2*

*d-ANOVA + Dunnett-test, p < 0.05 (based on absolute mean body weight)

**d-ANOVA + Dunnett-test, p < 0.01 (based on absolute mean body weight)

 

 

 

Table 3: Reproduction parameters for dams with live foetuses

Dose group

 

Control

30 mg/kg bw/day

100 mg/kg bw/day

300 mg/kg bw/day

Number of dams at term

9

10

10

10

Number of pregnant dams at term

9

10

10

10

Corpora lutea (total)

144

166

156

173

Corpora lutea per dam (mean ± SD)a

16.0 ± 1.6

16.6 ± 3.0

15.6 ± 3.4

17.3 ± 2.5

Implantation sites (total)

124

141

131

136

Implantation sitesa (mean ± SD)

13.8 ± 1.6

14.1 ± 1.9

13.1 ± 2.8

13.6 ± 2.1

Pre-implantation loss (total)b

20

25

25

37

Pre-Implantation loss (%)

13.9

15.1

16.0

21.4

Resorptions + scars (total)b

6

1

9

11

Resorptions + scars (% of implantation sites)b

4.5

0.7

6.9

8.1

Resorptions + scars (mean ± SD)a

0.7 ± 1.1

0.1 ± 0.3

0.9 ± 1.5

1.1 ± 2.1

Implant scars (total)

0

0

0

0

Early resorptions (total)b

3

1

8

11

Early resorptions as No. per dam (mean ± SD)a

0.3 ± 0.7

0.1 ± 0.3

0.8 ± 1.5

1.1 ± 2.1

Late resorptions (total)b

3

0

1

0

Late resorptions as No. per dam (mean ± SD)a

0.3 ± 1.0

0.0 ± 0.0

0.1 ± 0.3

0.0 ± 0.0

Fetuses (total)

118

140*

122

125

Fetuses per dam (mean ± SD)a

13.1 ± 2.3

14.0 ± 2.1

12.2 ± 4.2

12.5 ± 3.7

Live foetuses (%)

98.3

100.0

100.0

99.2

Dead foetuses (%)

1.7

0.0

0.0

0.8

Uterus weight(mean ± SD)a

79.3 ± 11.2

84.1 ± 12.8

72.1 ± 23.8

69.7 ± 21.2

ad-ANOVA +Dunnett-test based on pooled variance

bFishers exact test

*p < 0.05

 

Table 4: Developmental parameters for offspring

Dose group

 

Control

100 mg/kg bw/day

300 mg/kg bw/day

1000 mg/kg bw/day

Number of live foetusesb(m/f)

116 (62/56)

140*(74/66)

122 (58/64)

124 (63/62)

Sex ratio (m/f)

0.53/0.47

0.53/0.47

0.47/0.53

0.50/0.50

Weights of live foetuses, g

(mean ± SD)a

3.79 ± 0.32

4.01 ± 0.24

3.91 ± 0.47

3.46 ± 0.54

ad-ANOVA +Dunnett-test based on pooled variance

bFishers exact test

*p < 0.05

 

 

Table 5: Results of skeletal examination of offspring

Dose group

 

Control

100 mg/kg bw/day

300 mg/kg bw/day

1000 mg/kg bw/day

External malformations, number (% of total)b

1 (0.8%)

0 (0%)

0 (0%)

0 (0%)

External anomalies, number (% of total)b

0 (0%)

0 (0%)

0 (0%)

0 (0%)

ad-ANOVA +Dunnett-test based on pooled variance

bFishers exact test

*p < 0.05

 

 

Table 6: Historical data

 

Mean value (range)

Number of pregnant dams at term

829

Corpora lutea per dam

17.0 (14.6-21.5)

Implantation sites

14.0 (12.0-16.0)

Pre-implantation loss as

% of corpora lutea

17.1 (5.0-30.0)

Post-implantation loss as

% of corpora lutea

4.6 (0.0-16.6)

Resorptions

0.5 (0.0-2.3)

Live foetuses, % of implantation sites

95.4 (83.4-100)

Conclusions:
Hexanoic acid,-3,5,5-trimethyl-,-3,5,5-trimethylhexyl ester (CAS 59219-71-5) has no effect on intrauterine development.
Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Quality of whole database:
The available information comprises an adequate and reliable study, and is thus sufficient to fulfil the standard information requirements set out in Annex VIII-IX, 8.7, of Regulation (EC) No 1907/2006.
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

The potential of 3,5,5-trimethylhexyl 3,5,5-trimethylhexanoate to cause developmental toxicity was assessed in a prenatal developmental toxicity study performed according to OECD guideline 414 and under GLP conditions (Fabreguettes, 2000). 10 pregnant dams/dose were administered 30, 100 and 300 mg/kg bw/day in corn oil by gavage during gestation day 6 to 17. On day 20 of gestation the animals were euthanized and examined. No treatment-related systemic effects were observed in the P-females.

One female in the control group died, possibly due to a dosing error. In 1/10 dams in the high-dose group abdominal breathing and chromo rhinorrea were observed on gestation day 8 to 11, indicating poor health condition. The female that showed clinical signs of toxicity also had a body weight loss of approximately 10% from gestation day 6 to 12. For females in the high-dose group the body weight was significantly reduced on day 15 and 18. The body weight gain in the high-dose group was significantly reduced during gestation day 6 to 18, while total body weight gain was approximately 25% lower than in the control group. The food consumption was reduced in the high-dose group on gestation day 6-18, compared with the control group, although not statistically significant. In the female exhibiting clinical signs, a brownish content of the uterus and dilatation of the cervix was observed, but the toxicological relevance of this finding is unclear. No other treatment-related effects were noted. The NOAEL for systemic effects in the dams is considered to be 100 mg/kg bw/day.

There was no significant effect on the fertility index, corpora lutea, resorptions, pre-implantation loss, fetal viability (in utero), gravid uterus weight and number of fetuses. The slight, not significant, increase in resorptions and pre-implantation losses observed in the high-dose group is attributable to a single female that was in poor health (14/37 pre-implantation losses). All the results were within the historical data range. The slight changes observed are therefore not considered to be an adverse reproductive effect.

There were no significant differences between the control and treatment pups in the developmental parameters (post-implantation loss, embryo survival, body weight, sex ratio, viability). The slight, not significant, increase in post-implantation losses (7/12) observed in the high-dose group is attributable to a single female that was in poor health. This is considered to be a consequence of the poor maternal health condition and is not a reproductive effect. The incidence of malformations and variations in the offspring was comparable between the control and treatment groups. The NOAEL for developmental toxicity and teratogenicity was set at ≥ 300 mg/kg bw/day.

Based on the available data 3,5,5-trimethylhexyl 3,5,5-trimethylhexanoate is considered to be not toxic to development.


Justification for selection of Effect on developmental toxicity: via oral route:
The key GLP-compliant study performed similar to OECD guideline 414 was selected.

Justification for classification or non-classification

The available data on toxicity to reproduction does not meet the classification criteria according to Regulation (EC) 1272/2008 or Directive 67/548/EEC, and is therefore conclusive but not sufficient for classification.

An OECD Guideline 414 study will not provide evidence for definite claims of no fertility effects. However, since no adverse effects on reproduction and development was noted up to and including the highest dose level of 300 mg/kg bw/day, the conclusion of no classification (conclusive but not sufficient for classification) is considered to be justified.

Additional information

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