N,N'-bis(2,2,6,6-tetramethylpiperidin-4-yl)hexane-1,6-diamine

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

275 mg/kg bw/day

Study duration:

subchronic

Species:

rat

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

The potential toxicity of the substance was evaluated in a study compliant with OECD guideline 408 and GLP, following daily oral administration to rats for 13 weeks and after a 6-week recovery period (Study director 2008a) which is described in more detail under the endpoint Repeated dose toxicity.

The test substance, prepared in maize oil, was administered daily by oral gavage at dose levels of 44, 110, 275 mg/kg bw/day to three groups of 10 males and 10 female Sprague-Dawley rats, daily for 90 days. A fourth group was exposed to the vehicle only and served as control. Two additional groups of 5 male and 5 female rats per group concurrently treated with the vehicle and high dose level were retained following the final exposure for a further 6 weeks to assess the reversibility of any adverse findings. The selection of the dose-levels was based on the results of a previously conducted 28-day dose-range-finding study (Study director 2006).

No test substance-related influences were noted on fertility and reproduction parameters and organ weights. The histomorphological examination of the spermatogenic staging did not reveal any test substance-related morphological lesions in the testis and epididymis at any dose level.

In conclusion, for the substance the NOAEL for toxicity to reproductive organs was above 275 mg/kg bw/day derived from the subchronic toxicity study in the rat.

Short description of key information:
In the subchronic toxicity study in the rat no signs of toxicity to reproductive organs were observed up to the highest dose level of 275 mg/kg bw/day.

Justification for selection of Effect on fertility via oral route:
In a study compliant with OECD guideline 408 and GLP, the effects on certain parameters indicative for toxicity to reproduction were assessed following daily oral administration to rats for 13 weeks and after a 6-week recovery period (Study director 2008a). The study is described in more detail under the endpoint Repeated dose toxicity. Please refer to the discussion.

Effects on developmental toxicity

Description of key information

Oral: NOAEL (rat) - teratogenicity: 275 mg/kg bw/day
Oral: NOAEL (rat) - embryotoxicity: 110 mg/kg bw/day (slight increase in incidence of soft tissue variations at 275 mg/kg bw/day, probably due to maternal toxicity) 

Link to relevant study records

Reference

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 Oct 2005 - 07 Mar 2006

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP - Guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 414 (Prenatal Developmental Toxicity Study)

Version / remarks:

adopted January 22, 2001

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Freie und Hansestadt Hamburg, Behörde für Soziales, Familie, Gesundheit und Verbraucherschutz, Hamburg, Germany

Limit test:

no

Species:

rat

Strain:

other: CD/Crl: CD (SD)

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Deutschland GmbH, Sulzfeld, Germany
- Age at study initiation: 8-9 weeks
- Weight at study initiation: 193-243 g
- Housing: singly in Makrolon cages
- Diet (ad libitum): commercial Ssniff R-Z V1324
- Water (ad libitum): tap water
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 55 ± 15
- Photoperiod (hrs dark / hrs light): 12 / 12

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

The test substance is instable in water. Hence, corn oil was employed as vehicle. Test substance was suspended in the vehicle to the appropriate concentrations and was administered orally at a constant volume of 5 mL/kg bw. The dose of the test substance was adapted to the animal´s body weight daily.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

At start of the administration period and at termination (12 samples). The samples were analysed according to a gas chromatographic method.

Details on mating procedure:

Sexually mature male rats (8-9 weeks) served as partners. 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 and the stage of oestrus cycle. 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).

Duration of treatment / exposure:

GD 6-19

Frequency of treatment:

once daily

Duration of test:

until gestation day 20; 14 day treatment period

Remarks:

Doses / Concentrations:
44, 110, 275 mg/kg bw/d
Basis:
nominal conc.

No. of animals per sex per dose:

25

Control animals:

yes, concurrent vehicle

Details on study design:

Dose levels were selected based on the results of the dose-range-finding study in rats (LPT study no. 19520/05, REG-No. 2005-0298-DGT). Test substance was administered to two dams/group once daily from the 6th to 19th day of pregnancy. Dose levels of 40, 120 and 360 mg/kg bw/d were administered at a constant administration volume of 5 mL/kg bw.

Maternal examinations:

CAGE SIDE OBSERVATIONS: Yes
- Time schedule: daily
behavior, external appearance and nature of faeces, viability

BODY WEIGHT: Yes
- Time schedule for examinations: body weight was recorded on day 0 of gestation followed by daily weighings. Body weight gain was also calculated in intervals.

FOOD CONSUMPTION: Yes
Quantity of food consumed by each rat was recorded daily with the exception of gestation day 20. Food intake per rat (g/rat/day) was calculated using the total amount of food given to and left by each rat in each group on completion of a treatment day. The relative food consumption was calculated.

WATER CONSUMPTION: Yes
- Time schedule for examinations: Daily monitoring by visual appraisal of the drinking water bottles was maintained throughout the study.

POST-MORTEM EXAMINATIONS: Yes
- Sacrifice on gestation day #20
- Organs examined: The ovaries and the uteri of the dams were removed; the uteri (in total) were weighed. In order to check for possible drug effects, a dissection with macroscopic examination of the internal organs and placentae of the dams was carried out on the day of scheduled laparatomy or on the day on which the animals were found dead. In case of macroscopical findings, the affected maternal tissues were preserved in 7% buffered formalin for possible future histopathological examinations.

Fetal examinations:

Fetuses were removed and following examinations performed: macroscopic inspection of the placentae (focal indurations), number of fetuses (alive and dead) and placentae, sex and viability, number and size of resorptions, corpora lutea in the ovaries, implantations and location of fetuses in the uterus, gravid uterus weight, weights of fetuses and weights of the placentae, fetuses were inspected externally for damages, especially for malformations

Statistics:

For all numerical values, homogeneity of variances was tested using the Bartlett chi-square test. When the variances were homogenous, the Dunnett test was used to compare the experimental groups with the control group. In case of heterogeneity of variances, the Student´s t-test was carried out.

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
Mortality: 275 mg/kg bw/d: 3 of 21 pregnant dams died prematurely on GD 12 or 17
Clinical signs: 110 mg/kg bw/d: one dam showed piloerection on some test days; 275 mg/kg bw/d: pilo-erection, laboured breathing or clonic convulsions were noted in individual high-dosed dams on one to several test days
Body weight: 275 mg/kg bw/d: significant reductions were determined for the mean maternal body weight change on GD 6 to 9 and 9 to 12 and for the net weight change from day 6 onwards
Food and water consumption: no effects
Necropsy: 275 mg/kg bw/d: necropsy of the three prematurely decreased dams revealed reddened lungs in two dams and a reddened stomach in one dam.
Uterus and carcass weight: no effects

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:yes

Details on embryotoxic / teratogenic effects:
Corpora lutea, implantation sites, resorptions, weight and number of live fetuses, placental weights: no effects
Mortality: no
Malformations or Variations: no
Soft tissue evaluations: 275 mg/kg bw/d: slightly but significantly increased fetal and litter incidences for hemaorrhagic foci in the liver and for total soft tissue variations.
Retardations: no effects

Dose descriptor:

NOAEL

Effect level:

275 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: teratogenicity

Dose descriptor:

NOAEL

Effect level:

110 mg/kg bw/day (nominal)

Based on:

test mat.

Basis for effect level:

other: embryotoxicity

Abnormalities:

not specified

Developmental effects observed:

not specified

Analysis of the test substance-carrier mixtures revealed that formulations were correctly prepared and the results of 94.1% to 109.4% of the nominal value were within the admissible limits of 90% to 110%.

The maternotoxic effects reported in this study correlate with those observed in a subchronic study carried out with the test substance (refer to Section 7.5.1). In that study, all changes observed were considered to be due to the local corrosive changes noted, causing behavioural, biochemical and haematological changes and in some cases mortality. The changes observed were not considered to be due to systemic toxicity of the test substance given by gavage but due to its corrosive properties.

Conclusions:

The test substance possessed no teratogenic properties up to a dose level of 275 mg/kg bw/d. No test substance-related increase was noted in the incidence of fetal malformations, external or skeletal variations. At the maternotoxic dose (275 mg/kg bw/d) marginal embryotoxic properties were noted in form of slightly increased incidences of soft tissue variations.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

275 mg/kg bw/day

Species:

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

The developmental toxicity potential of the test substance by the oral route was evaluated in rats in a study performed according to the OECD guideline 414 and in compliance with GLP (Study director 2008b).The test substance was suspended in corn oil and administered by gavage to 3 groups of 25 pregnant rats from gestation days (GD) 6 to 19 at daily doses of 44, 110, and 275 mg/kg bw/day. A control group of 25 animals was treated with the vehicle only. A constant application volume of 5 mL/kg bw/day was used for all concentrations. The selection of the dose-levels was based on the results of a dose-range-finding study previously conducted.

Maternal toxicity was observed at 110 mg/kg bw/day in form of piloerection in one dam. At 275 mg/kg bw/day, piloerection, laboured breathing and clonic convulsions were noted in individual dams on one to several test days. A reduced net weight change was determined. Furthermore, 3/21 pregnant dams died prematurely, revealing reddened lungs and/or reddened stomach at necropsy.

At 275 mg/kg bw/day, soft tissue evaluation of the foetuses (Wilson, 1965) revealed slightly increased incidences for haemorrhagic foci in the foetal liver and for total soft tissue variations. There was no test substance-related increase in the incidence of foetal malformations, external or skeletal variations or skeletal retardations.

No effects were observed in the other parameters assessed. The NOAEL for the foetuses was 110 mg/kg bw/day based on the marginal embryotoxic properties noted in form of slightly increased incidences of soft tissue variations at 275 mg/kg bw/day, which were probably due to maternal toxicity. No test substance-related increase in the incidence of foetal malformations, external or skeletal variations were observed at the highest dose level. Therefore, the NOAEL for teratogenicity was 275 mg/kg bw/day.

Wilson, J.G. Methods for administration agents and detecting malformations in experimental animals; in: Wilson, J.G. and Warkany, J. (1965). Teratology - Principles and Techniques. The University of Chicago Press, Chicago and London, pp. 262 -277.

Justification for selection of Effect on developmental toxicity: via oral route:
There is only one study available for this endpoint.

Justification for classification or non-classification

The available data on the reproductive toxicity of the test substance is conclusive but not sufficient for classification according to DSD (67/548/EEC) and GHS (CLP, 1272/2008/EC).