Registration Dossier

Administrative data

Key value for chemical safety assessment

Effects on fertility

Description of key information
No other studies available.
Link to relevant study records
Reference
Endpoint:
toxicity to reproduction
Data waiving:
other justification
Justification for data waiving:
other:
Reproductive effects observed:
not specified
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

According to the ECHA Factsheet (ECHA, 2009) regarding the information requirements for reproductive toxicity, ECHA considers registration dossiers for substances >= 100 t/y (and 1000 t/y) as technically complete if the dossier contains a testing proposal for a pre-natal developmental toxicity study in accordance with Section 8.7.2 of column 1, Annex IX. In this dossier, a pre-natal developmental toxicity gavage study (rat, gavage, OECD 414) is proposed. Therefore, a two generation reproductive toxicity study (OECD 416) does not need to be conducted.


Short description of key information:
No studies have been performed to explicitly address the question of reproductive effects in animals caused by decamethylenediamine. A data waiver is claimed.

Effects on developmental toxicity

Description of key information
A prenatal development toxicity study was performed in rats according to OECD 414 (oral exposure). The test item did not possess any teratogenic effect. The no-observed adverse effect level (NOAEL) for teratogenicity was above 30 mg test item/kg b.w./day.
Link to relevant study records
Reference
Endpoint:
developmental toxicity
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2015-04-13 to 2015-07-20
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study
Qualifier:
according to
Guideline:
OECD Guideline 414 (Prenatal Developmental Toxicity Study)
Version / remarks:
adopted January 22, 2001
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.31 (Prenatal Developmental Toxicity Study)
Version / remarks:
Regulation (EC) No. 440/2008 (May 30, 2008)
GLP compliance:
yes
Limit test:
no
Species:
rat
Strain:
Sprague-Dawley
Details on test animals and environmental conditions:
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH Sandhofer Weg 7, 97633 Sulzfeld, Germany
- Age: 59 days
- Weight at study initiation: 181.3 - 236.8 g.
- Housing: single
- Diet: Pelleted maintenance diet, ad libitum, Commercial ssniff® R/Z V1324 (ssniff Spezialdiäten GmbH, 59494 Soest, Germany,
- Water: tap water, ad libitum
- Acclimation period: 11 days
ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 +/- 3 °C
- Humidity (%): 55 +/- 15 %
- Illumination: 12 hours artifical fluorescent light and 12 hours dark
Route of administration:
oral: gavage
Vehicle:
other: 0.5% aqueous hydroxyl propyl methylcellulose gel
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:

VEHICLE
- Vehicle: 0.5% aqueous hydroxyl propyl methylcellulose gel
- Concentration in vehicle: 3 mg / 10 mg /30 mg/kg b. w. /day
- Total volume applied: 5 ml/kg bw/treatment
- Test item preparation: The test item was suspended in the vehicle to the appropriate concentrations and the test item formulations were
administered orally at a constant volume (5 mL/kg b.w./day) once daily from day 6th to 19th day of gestation.
- Control: Control animals received the vehicle at a constant volume of 5 mL/kg b.w. orally once daily in the same way
- Male rats: Male rats for mating remained untreated
Analytical verification of doses or concentrations:
yes
Details on analytical verification of doses or concentrations:
The results confirmed that the test item formulations were correctly prepared and the concentrations were in good agreement to those expected. The homogeneity of the samples was warranted during the procedure of treatment.
In detail, the concentrations of test item in the low dose samples were between 93.7% and 96.9%, those of the intermediate samples between 95.7%
and 97.8% and those of the high dose samples between 96.0% and 99.2% of the nominal concentrations.
No test item could be detected in any of the control samples.
Details on mating procedure:
- Sexually mature ('proved') male rats of the same breed served as partners.
- The female breeding partners were randomly chosen.
- Mating was monogamous: 1 male and 1 female animal were placed together in one cage during the dark period. Each morning a vaginal smear was
taken to check for the presence of sperm. If findings were negative, mating was repeated with the same partner. The day on which sperm was found
was considered as the day of conception (day 0 of pregnancy).
- This procedure was repeated until enough pregnant dams were available for all groups.
- Rats which did not become pregnant were excluded from the analysis of the results and replaced by other animals.
- A post-mortem negative staining according to SALEWSKI was carried out in the replaced animals in order to confirm the non-pregnancy status.
Duration of treatment / exposure:
- 1 mating day
- 14 administration days from gestation day 6 to 19
Frequency of treatment:
daily
Duration of test:
until day 20 post-coitum
Remarks:
Doses / Concentrations:
3, 10, 30 mg/kg bw/day
Basis:
other: nominal in vehicle
No. of animals per sex per dose:
Treated animals: Groups 1 - 4: 25 animals per group
Evaluated litters: Groups 1 - 4: 20 litters per group
Control animals:
yes, concurrent vehicle
Details on study design:
- The dose levels were selected in agreement with the Sponsor based on the results of a dose-range-finding study for a prenatal developmental
toxicity study
- In the dose-range finding study, test item was administered to female rats at dose levels of 30, 100, or 300 mg/kg b.w./day orally, by gavage,
once daily from gestation day 6 to 19.
- Under the present test conditions, the no-observed-adverse-effect level (NOAEL) was below 30 mg test item/kg b.w./day for the dams
- Signs of clinical toxicity (e.g. breathing sounds, piloerection) were noted at 30 mg test item/kg b.w./day and with an increased incidence at
100 mg test item/kg b.w./day. Reductions in body weight and food consumption were noted for the dams treated with 100 mg test item/kg b.w./day.
- At 300 mg test item/kg b.w./day 2 of 3 dams died prematurely 4 days after dosing and the dose level group was terminated due to this high
mortality. No premature death was noted for the dams treated with 30 and 100 mg test item/kg b.w./day. Necropsy revealed no changes for the
dams treated with 30 or 100 mg test item/kg b.w./day.
- The no-observed-adverse effect level (NOAEL) for the development of the fetal organism was below 30 mg test item/kg b.w./day.
- A slightly increased number of resorptions was noted at 30 mg test item/kg b.w./day, followed by a higher incidence of resorptions at
100 mg test item/kg b.w./day. A reduced fetal body weight was noted at 100 mg test item/kg b.w./day.
- The external examination of the fetuses revealed no malformations or variations at 30 or 100 mg test item/kg b.w./day
- For the present rat embryotoxicity study, dose levels of 3, 10, or 30 mg test item/kg b.w./day, administered orally, by gavage once daily
(at a constant volume of 5 mL/kg b.w./day) from the 6th to the 19th day of pregnancy, were selected in agreement with the Sponsor.
Maternal examinations:
PARAMETERS ASSESSED DURING STUDY:
- Clinical signs: Individual animals were observed daily for behavioural changes, reaction to treatment, or illness
- Viability:Checks were made early in the morning and again in the afternoon of each working day to look for dead or moribund animals
- Body weight: day 0 of gestation (the day of detection of a positive mating sign), followed by daily weighing - always at the same time of the day
- Body weight change: 3-6, 6-9, 9-12, 12-15, 15-18 and 18-20, furthermore the net weight change from day 6 is given
- Food consumption: Quantity of food consumed by each rat was recorded daily
- Water consumption: Daily monitoring by visual appraisal of the drinking water bottles was maintained throughout the study

ORGANS EXAMINED AT NECROPSY (MACROSCOPIC AND MICROSCOPIC):
- Macroscopic examination of the internal organs and placentae of the dams was carried out on the day of sacrifice. The following target organs of
the dams were weighed: Heart, kidney (2), and liver.
Ovaries and uterine content:
Examination of uterine content: Ovaries and the uteri of the dams were removed; the gravid uteri (in toto) were weighed.
See below: Fetal examinations

Fetal examinations:
Examination of the fetuses: The fetuses were removed and the following examinations performed:
- Macroscopic inspection (gross evaluation) of the placentae for example for focal indurations.
- The number of fetuses (alive and dead) and placentae (location in the uterus and the assignment of the fetuses) was determined.
- Sex and viability of fetuses were determined. Animals are said to be viable when they are found alive (spontaneous breathing, spontaneous
movement).
- Number and size of resorptions were determined.
- Corpora lutea in the ovaries, implantations and location of fetuses in the uterus were determined.
- Weights of fetuses and weights of the placentae were determined (fetuses were considered as runts if their weight was less than 70% of the
mean litter weight).
- All fetuses (dead or alive) were inspected externally for damages, especially for malformations .
- The fetuses were sacrificed by an ether atmosphere.
Examination of fetuses and determination of number and kind of retardations, variations or malformations:
- 50% of the number of fetuses in each litter were examined for skeletal anomalies. The thorax and peritoneal cavity (without damage to ribs and
sternum) were opened and the location, size and condition of the internal organs were determined.
The skeleton was double-stained with Alcian blue for the examination of cartilage and with Alizarin red to reveal ossifications (according to
DAWSON). The skeletal system was examined (determination of the number and type of retardations, variations as well as malformations).
- The remaining 50% of the number of fetuses in each litter were examined for soft tissue anomalies. Body sections were made and examined
according to WILSON.
The fetuses were allocated to the evaluation of DAWSON or WILSON on an alternating basis.
Statistics:
Statistical analyses of the parametrical values captured by Provantis software were calculated using the following settings:
- Analysis of normal distribution and homogeneity of variances was performed by using the SHAPIRO-WILKS test and the BARTLETT test. Data not
normally distributed or with heterogeneous variances between the groups were stepwise log- or rank-transformed.
- One-way analysis of variance (ANOVA) was performed with non-transformed or log-transformed data. The KRUSKAL-WALLIS test was used for
rank-transformed data.
- In case of significant differences (found by ANOVA or KRUSKAL-WALLIS test), intergroup comparisons with the control group were made by
parametric or non-parametric DUNNETT multiple comparison tests (p ≤ 0.05 and p ≤ 0.01).
- Parametrical values not captured by Provantis (e.g. number and weight of the fetuses) were analysed by the DUNNETT test (p ≤ 0.05 and p ≤ 0.01).
Prior to the DUNNETT test homogeneity of variances was tested using the BARTLETT test. In case of heterogeneity of variances, the STUDENT's t-test was carried out (p ≤ 0.05 and p ≤ 0.01).
- Statistical analyses of non-parametrical data (e.g. resorption-, malformation-, variation and retardation rates) were performed using the
following settings:
FISHERs exact test, n < 100; (p ≤ 0.05 and p ≤ 0.01) or
chi2-test with Yates' correction for continuity, n ≤ 0.01 (p ≤ 0.05 and p ≤ 0.01)
- Significantly different data are indicated in the summary tables of the result sections of the report.
Historical control data:
No test item-related differences between the ratio of live male fetuses to live female fetuses were noted between the control group and the treatment
groups (3, 10 or 30 mg test item/kg b.w./day, p.o.). All values, with the following exception, were within the range of LPT background data.
Details on maternal toxic effects:
Maternal toxic effects:no effects

Details on maternal toxic effects:
Under the present test conditions, the no-observed adverse effect level (NOAEL) was 30 mg test item/kg b.w./day for the dams.
Dose descriptor:
NOAEL
Effect level:
30 mg/kg bw/day
Basis for effect level:
other: maternal toxicity
Details on embryotoxic / teratogenic effects:
Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
No malformation was noted at any of the test item treated groups. There was no test item-related increase in the incidence of fetal external / internal, skeletal or soft tissue variations or skeletal retardations at any tested dose level
Dose descriptor:
NOAEL
Effect level:
> 30 mg/kg bw/day
Basis for effect level:
other: teratogenicity
Abnormalities:
not specified
Developmental effects observed:
not specified

Examination of the dams

Test item

Group 1

Control

Group 2

3 mg/kg

Group 3

10 mg/kg

Group 4

30 mg/kg

Treated dams

25

25

25

25

Not pregnant dams

0

2

1

0

Dams without viable fetuses

0

0

0

0

Not examined dams

(spare animals)

5

3

4

5

Evaluated litters

20

20

20

20

 

Summary of animals evaluated

Test item

dose in mg/kg

b.w./day, p.o.

Animal nos.

of mated

rats

Animal nos.

of rats with evaluable litters

at laparotomy

Dams not pregnant

(animal nos.)

Dams

with total

implantation loss

(animal nos.)

Reserve

animals, not

examined

(animal nos.)

Control

1 - 25

1 - 20

none

none

21 - 25

3

26 - 50

26, 27, 29,

31 - 47

28, 30

none

48 - 50

10

51 - 75

52 - 71

51

none

72 - 75

30

76 - 100

76 - 95

none

none

96 -100

Behaviour, external appearance, faeces, mortality

No test item-related changes of behaviour or external appearance were noted in the dams treated orally with 3 or

10 mg test item/kg b.w./day.

At 30 mg test item/kg b.w./day, salivation was noted for three high dose dams on one treatment day each, starting immediately to

5 minutes after administration and lasting for 20 to 60 minutes:

-   Extreme salivation was observed for dam no. 78 on gestation day 9 and for dam no. 83 on gestation day 7.

-  Moderate salivation was observed for dam no. 76 on gestation day 18.

This finding is considered to be a marginal not adverse effect. No further changes were noted in behaviour or external appearance of the high dose dams.

None of the dams died prematurely. The faeces were of normal consistency throughout the experimental period.

 

Body weight and body weight gain

No test item-related changes in body weight or body weight change (examined in 3-day intervals) were noted in comparison to the control group for the low and intermediate dose dams (3 or 10 mg test item/kg b.w./day). The mean body weight gain at the time point of laparotomy on gestation day 20 was similar to the body weight gain of the control dams when calculated from the start value (day 0 of pregnancy) as well as when calculated from gestation day 6 (start of treatment). For details, see table.

The mean body weight change of the high dose dams (treated with 30 mg test item/kg b.w./day) was markedly (statistically not significant) below the control values between gestation days 12 and 15 (by 32.6% in comparison to the control group) due to a moderate reduction in food consumption during this period. This finding is considered to be a marginal effect of the test item.

In the following 3-day intervals (15 to 18; 18 to 20) until laparotomy, the body weight change of the high dose dams was again in the range of the control group. This finding is considered to be a marginal not adverse effect. No influence was seen on the body weight gain calculated from the start value (day 0 of pregnancy) or calculated from gestation day 6 (start of treatment) until laparotomy on gestation day 20. For details, see table

 

Test item - Mean body weight gain

Time interval

Group 1:

Control

Group 2:

3 mg/kg

Group 3:

10 mg/kg

Group 4:

30 mg/kg

GD 0 to GD 20

66%

68%

69%

65%

GD 6 to GD 20

41%

44%

44%

40%

Food and drinking water consumption

No test item-related changes in food consumption were noted between the dams of the control group and the dams treated orally with 3 or 10 mg test item/kg b.w./day.

At 30 mg test item/kg b.w./day, moderate reductions in food consumption of the dams (up to 10.3% below the control, statistically not significant) were noted between gestation days 12 and 16. These transient reductions in food intake are considered to be marginal not adverse effects.

Drinking water consumption showed no test item-related changes at any tested dose level as observed during daily visual appraisal.

Examination of the dams at termination

Necropsy findings

Macroscopic inspection of the internal organs of the dams at laparotomy revealed no changes in the control and the test item-treated

groups(3, 10, or 30 mg test item/kg b.w./day).

 

Uterus weight and net body weight change

No test item-related changes in the gravid uterus weight, the carcass weight (terminal body weight minus uterine weight) and the net

weight change from day 6 onwards (carcass weight minus day 6 body weight) in comparison to the control group were noted for the low and intermediate dose dams(3 or 10 mg test item/kg b.w./day).

At 30 mg test item/kg b.w./day, the net body weight change from day 6 onwards (24.72 g) was below that of the control dams

(32.82 g). This difference (statistically not significant) is considered to be a marginal not adverse effect.

 

Organ weights

No test item-related changes in the absolute or relative organ weights of heart, liver or kidneys were noted in comparison to the control group for the dams treated orally with 3, 10, or 30 mg test item/kg b.w./day.

 

Reproduction data of the dams

No test item-related influence was detected on the prenatal fetal development at 3, 10, or 30 mg test item/kg b.w./day with respect to the implantation sites, resorptions, number of live fetuses, the values calculated for the post-implantation loss and the sex distribution of fetuses when compared to the control.

The statistical analyses of the parameters of reproduction were performed by comparison of the ratios of:

-

implantation sites/corpora lutea (described as pre-implantation loss)

-

resorptions/implantation sites

-

live fetuses/implantation sites (described as post-implantation loss)

 

Pre-treatment period

All parameter determined before start of test item-treatment on gestation day 6were well within the range of LPT background data. This was true forthe number ofcorpora lutea or implantation sites and the values calculated for the pre-implantation loss.

As a spontaneous finding, theratio of corpora lutea vs. implantation sites was slightly but statistically decreased (at p </= 0.01) in the intermediate dose dams compared to the control dams leading to a reduced pre-implantation loss. This finding is not test item-related but incidental as the pre-implantation loss occurred before start of test item-treatment on gestation day 6.

Treatment period (during organogenesis)

No test item-related influence on the prenatal fetal development was detected after treatment with 3, 10, or 30 mg test item/kg b.w./day with respect to the number of resorptions and live fetuses or the values calculated for the post-implantation loss.

Slight but statistically significant decreases (at p </= 0.01 or p >/= 0.05) were noted for the ratio of early, late and total resorptions vs. implantation sites. These differences compared to the control were due to the relatively high incidence of 21 total resorptions (14 early and 7 late resorptions) in the control group while low incidences of only 6, 4, or 9 total resorptions (including early and late resorptions) were noted in the low, intermediate and high dose groups.

Significant differences in reproduction data which arenotconsidered to be test item-related are given in the Table below:

 Parameter

Ref. Table No.

Increase

Decrease

Group

Statistical significance

Reason

Ratio implantation sites vs. corpora lutea

(pre-implantation loss)

6

Decrease

3

p </= 0.01

A, C, D

Ratio total resorptions vs. implantation sites

6

Decrease

2, 3

4

p </=0.01 

p >/= 0.05

A, C

Ratio early resorptions vs. implantation sites

6

Decrease

2, 3

p </= 0.05

A, B, C

Ratio late resorptions vs. implantation sites

6

Decrease

, 4

p </= 0.01

A, C

Ratio live fetuses vs. implantation sites

(post-implantation loss)

6

Increase

2, 3

4

p </= 0.01

p </= 0.05c

A, C

A:        the slight alteration in comparison to control animals is without biological relevance

B:        lacking dose dependence

C:        effect is due to the relative low or high value observed for the control group

D:       effect observed before start of treatment

Table of reproduction data:

Parameter

Group 1

Control

(n=20)

Group 2

3 mg/kg

(n=20)

Group 3

10 mg/kg

(n=20)

Group 4

30 mg/kg

(n=20)

 

Corpora lutea

total

per dam

 

286

14.3

279

14.0

277

13.9

288

14.4

 

Implantation sites ##1

total

per dam

 

273

13.7

271 #1

13.6

274** #2

13.7

279

14.0

 

Resorptions ##2

total

per dam

 

21

1.1

6** #2

0.3

4** #2

0.2

9* #2

0.5

 

Early resorptions ##2

total

per dam

 

14

0.7

4 * #2

0.2

4* #2

0.2

9

0.5

 

Late resorptions ##2

total

per dam

 

7

0.4

2

0.1

0** #2

0.0

0** #2

0.0

 

Live fetuses ##3

total

per dam

 

252

12.6

266** #1,2

13.3

270** #2

13.5

270 * #2

13.5

 

Dead fetuses at laparotomy

total

 

0

0

0

0

 

Pre-implantation loss

mean %

 

4.0

2.8

1.1

2.5

 

Post-implantation loss

mean %

 

7.6

2.3

1.5

3.2

 

 

 

 

 

*

Significantly different from the controls at p ≤ 0.05, Chi2test.

 

**

Significantly different from the controls at p ≤ 0.01, Chi2test.

 

 

 

 

#1

Including a set of twins

 

#2

Statistical difference is considered as incidental finding,nottest item-related.

 

 

 

 

 

 

 

 

 

 

For the statistical analyses of the above mentioned parameters of reproduction the following comparisons were performed:

 

##1

comparing the values of implantation sites/corpora lutea of the test group with the ratio of implantation sites/corpora lutea of the control group using the Chi2test. This comparison is described as pre-implantation loss (seeSection 5.3'Evaluation parameters').

 

##2

comparing the values of resorptions/implantation sites of the test group with the ratio of resorptions/implantation sites of the control group using the Chi2test.

 

##3

comparing the values of live fetuses/implantation sites of the test group with the ratio of fetuses/implantation sites of the control group using the Chi2test. This comparison is described as post-implantation loss (seeSection 5.3'Evaluation parameters').

Examination of the fetuses

Mortality

No dead fetuses were noted in the litters of the dams of the control group and in the litters of the dams treated orally with 3, 10, or 30 mg test item/kg b.w./day.

 

Sex distribution of the fetuses

No test item-related differences between the ratio of live male fetuses to live female fetuses were noted between the control group and the treatment groups (3, 10, or 30 mg test item/kg b.w./day). All values were within the range of LPT background data.

Parameter

Sex ratio

observed in this study

(LPTReport 31734)

 

(male / female)

LPT Background Data#

range of mean values per group

(n=56 control or

n= 143 test item groups;

Data taken from 2000 to July 2014)

Sex ratio of fetuses

(male/female)

Group 1:

0.92

0.82 - 1.38

0.64 - 1.38

(Control)

(test item groups)

Group 2:

1.04

Group 3:

0.79

Group 4:

0.85

#: not audited by QAU

Weight of placentae

No test item-related differences of the placental weights were noted between the control group and the treatment groups (3, 10, or 30 mg test item/kg b.w./day).

 

 

Weight of the fetuses

No test item-related differences of the fetal weights were noted between the control group and the treatment groups (3, 10, or 30 mg test item/kg b.w./day).

No test item-related effect was noted for the number of runts. In total, four runts were noted at laparotomy: one runt each was found at 3 and 10 mg test item/kg b.w./day and two runts (in one litter) at 30 mg test item/kg b.w./day. The afore-mentioned incidences are within the spontaneous range of variability.

For details on runts refer to the following Table:

 

 

Group 1: Control

Group 2: 3 mg/kg

Group 3: 10 mg/kg

Group 4: 30 mg/kg

Dam no.

Fetus no.

Dam no.

Fetus no.

Dam no.

Fetus no.

Dam no.

Fetus no.

-

no runts

35

15 f

62

7 f

81

3 m

 

 

 

 

 

 

7 f

 

External and internal macroscopic examination of the fetuses

External examination at laparotomy

Twins

As a spontaneous finding a set of twins (implant no. 1: one female fetus and one male fetus) was noted in the litter of the low dose dam no. 32 (treated with 3 mg test item/kg b.w./day).

Malformations / Variations

No macroscopically visible malformations or variations were noted for the fetuses of the control group and the treatment groups (3, 10, or 30 mg test item/kg b.w./day) during the external examination of the fetuses at laparotomy.

  

Internal examination at laparotomy

Malformations / Variations

A macroscopic internal examination was performed to detect gross changes of the internal organs. No malformations or variations were noted during the internal examination of the fetuses of the control group and the treatment groups (3, 10, or 30 mg test item/kg b.w./day).

Skeletal examination of the fetuses

Malformations

No malformations were noted during skeletal examination according to DAWSON at any tested dose level (3, 10, or 30 mg test item/kg b.w./day).

Variations

The skeletal variations observed were related to the sternum (sternebra(e) bipartite, dumbbell-shaped, fused or misaligned to a slight degree) or the rib(s) (rib(s) short or wavy).

No test item-related increase in the incidence of skeletal variations in comparison to the control group was noted in any of the test item treated groups (3, 10, or 30 mg test item/kg b.w./day). All findings are regarded to be spontaneous.

The statistically significant difference in the fetal incidence of the following skeletal sternal variation represents a decrease and is not considered to be test item-related as given in the Table below.

 

Skeletal variation

Mean value observed

in this study

(LPTReport 31734)

 

[fetal incidence in%]

LPT Background Data#

range of mean values per group

[fetal incidence in mean %]

(n=55 control or

n= 141 test item groups;

Data taken from 2000 to

July 2014)

Sternebra(e)bipartite

Control:    3.2

Group 2:   0.0 *

Group 3:   0.7

Group 4:   1.5

0.0 - 10.4

0.0 - 10.9

(control)

(test item groups)

#:

not audited by QAU

*/**:

(p ≤ 0.05 / p ≤ 0.01) Fisher or Chi2- test

Retardations

The observed skeletal retardations were related to the skull (incomplete ossification of frontal, parietal, interparietal, and/or supraoccipital areas), the hyoid (unossified), the sternum (sternebra(e) incompletely ossified, reduced in size or unossified), thethoracic vertebral bodies (bipartite, dumbbell-shaped or unossified), the lumbar or sacral vertebral bodies (unossified), the caudal vertebral bodies (only one body ossified or all bodies unossified), the os ischii (incompletely ossified or unossified), the os pubis (reduced in size or unossified), and the metacarpalia and metatarsalia (absence of ossification in metacarpalia 2 to 5 and metatarsalia 2 to 5).

No test item-related influence was noted on the incidence of skeletal retardations at 3, 10, or 30 mg test item/kg b.w./day.All incidences were in the range of LPT background data

The following statistically significant differences in fetal incidences of skeletal retardations were noted in test groups 2, 3, or 4 (dams treated with 3, 10, or 30 mg test item/kg b.w./day) which are not considered to be test item-related as given in the Table below. They are well within or only slightly above the range of LPT background data, appeared without any dose response relationship and/or represent a decrease compared to the control.


Skeletal retardations

Mean value observed

in this study

(LPTReport 31734)

 

[fetal incidence in %]

LPT Background Data#

range of mean values per group

[fetal incidence in mean %]

(n=55 control or

n= 141 test item groups;

Data taken from 2000 to

July 2014)

Skull incomplete

ossification

Control:  15.9

Group 2: 26.3 *

Group 3: 25.2 *

Group 4: 18.5

1.4 - 71.3 (control)

1.4 - 79.5 (test item groups)

Hyoid unossified

Control:  57.9

Group 2: 63.9

Group 3: 46.7 *

Group 4: 54.8

0.0 - 89.4 (control)

0.0 - 93.7 (test item groups)

Sternebra(e) unossified

Control:  85.7

Group 2: 88.0

Group 3: 85.9

Group 4: 64.4**

5.8 - 91.9 (control)

1.2 - 88.5 (test item groups)

Os ischii incompletely ossified

Control:    2.4

Group 2:   1.5

Group 3:   7.4*

Group 4:   2.2

0.0 -   2.5 (control)

0.0 -   5.3 (test item groups)

Absence of ossification in metacarpalia 2 to 5

Control:  82.5

Group 2: 97.7**

Group 3: 81.5

Group 4: 69.6**

0.0 - 94.9 (control)

0.0 - 97.6 (test item groups)

#:

not audited by QAU

*/**:

(p ≤ 0.05 / p ≤ 0.01) Fisher or Chi2- test

Soft tissue examination of the fetuses

Malformations

No malformations were noted during soft tissue examinations of the fetuses according to WILSON in the control group and in any of the treatment groups (3, 10, or 30 mg test item/kg b.w./day).

 

Variations

Soft tissue variations were noted in the cerebral ventricle (dilatation of the 4th ventricle, rarely dilatation of the 3rd ventricle), the kidneys (uni- or bilateral dilatation of the renal pelvis), and the liver (haemorrhagic focus/foci).

No test item-related differences in the incidences of the observed soft tissue variations were noted between the control group and the test item treated groups (3, 10, or 30 mg test item/kg b.w./day). All incidences were in the range of LPT background data.

 

 

Conclusions:
In conclusion, no malformation was noted at any of the test item treated groups. There was no test item-related increase in the incidence of fetal
external / internal, skeletal or soft tissue variations or skeletal retardations at any tested dose level.
The test item did not possess any teratogenic effect. The no-observed-adverse effect level (NOAEL) for teratogenicity was above 30 mg test item/kg
b.w./day.
Executive summary:

Aim of the study is the examination of the influence of the test item administered orally during the critical period of organogenesis and the fetal development (6th to 19th day of gestation) on the pregnant rat and the fetus.

In this rat embryotoxicity study, the test item was administered to female rats at dose levels of 3, 10 or 30 mg/kg b.w./day (administration volume: 5 mL/kg b.w./day), orally by gavage from the 6th to 19th day of pregnancy.

Under the present test conditions, the no-observed adverse effect level (NOAEL) was 30 mg test item/kg b.w./day for the dams.

Oral treatment with 30 mgtest item/kg b.w./day caused transient marginal reductions in body weight change, net body weight change and food consumption of the high dose dams as well as salivation in few of them. However, these changes were not considered to be adverse.

The no-observed-effect level (NOEL) for the fetal organism was above 30 mgtest item/kg b.w./day.

No malformation was noted at any of the test item treated groups. There was no test item-related increase in the incidence of fetal external / internal, skeletal or soft tissue variations or skeletal retardations at any tested dose level.

The test item did not possess any teratogenic effect. The no-observed-adverse effect level (NOAEL) for teratogenicity was above 30 mg test item/kg b.w./day.

Effect on developmental toxicity: via oral route
Endpoint conclusion:
no adverse effect observed
Dose descriptor:
NOAEL
30 mg/kg bw/day
Study duration:
subacute
Species:
rat
Quality of whole database:
The study is valid without restriction (Klimisch score 1).
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

In the dose-range-finding study for a prenatal developmental toxicity study test item was administered to female rats at dose levels of 30, 100 or 300 mg test item/kg b.w./day orally, by gavage from the 6th to 19th day of pregnancy (LPT, 2015b). Signs of clinical toxicity (e.g. breathing sounds, piloerection) were noted at 30 mg test item/kg b.w./day and with an increased incidence at 100 mg test item/kg b.w./day. Reductions in body weight and food consumption were noted for the dams treated with 100 mg test item/kg b.w./day.

 At 300 mg test item/kg b.w./day 2 of 3 dams died prematurely 4 days after dosing (on gestation day 9) and the dose level group was terminated due to this high mortality.

No premature death was noted for the dams treated with 30 and 100 mg test item/kg b.w./day.

Necropsy revealed no changes for the dams treated with 30 or 100 mg test item/kg b.w./day.

A slightly increased number of resorptions was noted at 30 mg test item/kg b.w./day, followed by a higher incidenceof resorptions at 100 mg test item/kg b.w./day.

A reduced fetal body weight was noted at 100 mg test item/kg b.w./day.

The external examination of the fetuses revealed no malformations or variations at 30 or 100 mg test item/kg b.w./day.

Based on the data obtained in this dose-range-finding study, the following dose levels are suggested for the main study:

Group 1: Control, Group 2:  3 mg test item/kg b.w./day, p.o, Group 3: 10 mg test item/kg b.w./day, p.o, Group 4: 30 mg test item/kg b.w./day, p.o

In the prenatal developmental study according to OECD 414 (LPT, 2015a), the test item was administered to female rats at dose levels of 3, 10 or 30 mg/kg b.w./day (administration volume: 5 mL/kg b.w./day), orally by gavage from the 6th to 19th day of pregnancy. Under the present test conditions, the no-observed adverse effect level (NOAEL) was 30 mg test item/kg b.w./day for the dams. Oral treatment with 30 mg test item/kg b.w./day caused transient marginal reductions in body weight change, net body weight change and food consumption of the high dose dams as well as salivation in few of them. However, these changes were not considered to be adverse.

The no-observed-effect level (NOEL) for the fetal organism was above 30 mg test item/kg b.w./day. No malformation was noted at any of the test item treated groups. There was no test item-related increase in the incidence of fetal external / internal, skeletal or soft tissue variations or skeletal retardations at any tested dose level.The test item did not possess any teratogenic effect. The no-observed-adverse effect level (NOAEL) for teratogenicity was above 30 mg test item/kg b.w./day.


Justification for selection of Effect on developmental toxicity: via oral route:
Relevant key study for this toxicological endpoint

Justification for classification or non-classification

Regarding toxicity to reproduction, decamethylenediamine is not classified according to the criteria of EC Directive 67/548/EEC and EC Regulation 1272/2008.