Amines, N-(3-aminopropyl)-N-(C16-18 and C18-unsatd. alkyl)trimethylenedi-

Endpoint summary

Description of key information

Data is based on read across from a structurally similar branched triamine C12. A NOAEL of 7 mg/kg bw/day was established in a 90-day repeated dose toxicity study with rats, based on tubular nephropathy in the kidney and the appearance of foamy macrophages in mesenteric lymph nodes at the next dose level. In a 90-day study with dogs, a NOAEL of 20 mg/kg bw/day was established, based on decreased body weight, increased absolute and relative gall bladder weight and increased potassium and ALAT and ASAT activities in plasma. In a combined chronic repeated dose toxicity/carcinogenicity diet study with rats a NOAEL of 4 mg/kg bw/day was established, based on increased ASAT activity, increase of lymphohistiocytic inflammatory reactions in the kidney, skeletal muscle and heart, and foamy macrophages in the mesenteric lymph nodes and alveoli of the lungs at the next dose level.

Key value for chemical safety assessment

Toxic effect type:

dose-dependent

Repeated dose toxicity: via oral route - systemic effects

Link to relevant study records

Reference

Endpoint:

chronic toxicity: oral

Remarks:

combined repeated dose and carcinogenicity

Type of information:

read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Justification for type of information:

For details and justification of read-across please refer to the report attached in section 0.2 of IUCLID.

Dose descriptor:

NOAEL

Effect level:

4 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no adverse effects observed

Dose descriptor:

LOAEL

Effect level:

8 mg/kg bw/day (actual dose received)

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: see 'Remark'

Critical effects observed:

not specified

Conclusions:

Based on the present results the no-observed-effect-level (NOEL) was 4 mg N-(3-aminopropyl)-N-dodecylpropane-1,3-diamine/kg b.w. via the diet.

Executive summary:

Materials & methods:

Study has been performed according to GLP and following the OECD Guideline for the Testing of Chemicals No. 453: Combined chronic toxicity/carcinogenicity studies, with some additions to comply also to OPPTS 870.4300 Combined chronic toxicity/ carcinogenicity, August 1998.

In total 500 animals (250 males and 250 females) were included:

- Carcinogenicity study (main study): 400 (200 male and 200 female);

    50 animals/sex/dose group

- Chronic toxicity study (satellite study): 100 (50 male and 50 female);

    10 animals/sex for groups 1, 2, 3 and 20 animals/sex for group 4 (high dose)

Dosing was done via dietary admixture. Concentrations in food were freshly prepared weekly and adjusted to reach test substance intake of 4, 8, 20 mg/kgBW during the whole course of the study.

Chronic part:

Evaluations included in the chronic study part consisted of twice daily mortality, daily individual observations for any signs of behavioural changes, reaction to treatment or illness, and weekly individual observations in standard arena, including changes in skin, fur, eyes, mucous membranes, occurrence of secretions and excretions and autonomic activity (e.g. lacrimation, pilo-erection, pupil size, unusual respiratory pattern). Changes in gait, posture and response to handling as well as the presence of clonic or tonic movements, stereotypies (e.g. excessive grooming, repetitive circling) or bizarre behaviour (e.g. self-mutilation, walking backwards) were also recorded.

Neurological screening was performed at week 52 consisting of an extensive observational screen and functional tests.

Body weights were recorded weekly, and at two week interval after week 14. The quantity of food left by individual animals was recorded on a weekly basis throughout the first 13 test weeks and from test week 14 onwards throughout the experimental period every second week.

Haematology, biochemistry and urinalysis were performed during week 13, 26 and 52. After week 4 additional biochemistry was performed in control and high dose group.

After week 52 all satellite animals were sacrificed at which point all gross pathological observation were recorded, all organs weighted and processed for histopathological examination.

 

Results and discussion

Mortality:

Eight of 20 male animals treated with 20 mg via the diet died during the course of the study.

Clinical signs:

No clinical signs of systemic toxicity were noted at any of the tested dose levels.

Functional observations (neuropharmacological screening):

The functional observation battery did not reveal any test item-related influence on the rats treated with 4, 8 or 20 mg N-(3-aminopropyl)- N-dodecylpropane-1,3-diamine/kg b.w./day via the diet. No influence was noted on fore- and hindlimb strength or spontaneous motility in any animal treated with the test item at any dose level.

Body weight and body weight gain:

The body weight of the rats treated with 20 mg N-(3-aminopropyl)-N-dodecylpropane-1,3-diamine/kg b.w. via the diet was reduced by up to 34 % from test week 15 onwards for the male animals an by up to 22% from test week 33 onwards for the female animals. The body weight gain changed accordingly. The body weight at autopsy was reduced by 35% for the males and by 19% for the females treated with 20 mg N-(3-aminopropyl)-Ndodecylpropane- 1,3-diamine/kg b.w. via the diet.

Food consumption:

The food consumption of the animals treated with 20 mg N-(3-aminopropyl)-N-dodecylpropane- 1,3-diamine/kg b.w. via the diet was increased by up to 18% for the male rats in test weeks 39 to 45 and by up to 20% for the females in test weeks 31 to 51.

The visual appraisal of the drinking water consumption did not reveal any test item-related influence.

Test item intake

The results of the samples taken immediately after preparation were within 80-120% of the nominal concentrations.

Haematology

The animals treated with 20 mg via the diet revealed changes in the haemoglobin content, the numbers of leucocytes, reticulocytes, platelets, neutrophilic granulocytes, lymphocytes, monocytes, eosinophilic granulocytes, large unstained cells, basophilic granulocytes, the haematocrit value, the MCV, the MCH and the MCHC.

Clinical biochemistry

The animals treated with 20 mg via the diet revealed increases in the plasma level of urea and in the plasma activities of ALAT, ASAT, LDH and Gamma-GT as well as decreases in the plasma levels of albumin, cholesterol, glucose, protein (total) and chloride.

For the male and female animals treated with 8 mg via the diet an increase of the plasma activity of ASAT by 122% for the males (test week 52) and by 91% for the females (test week 26) was noted.

 

Urinalysis

No test item-related influence was noted at any tested dose level.

Macroscopic post mortem findings

At necropsy, discoloured or reddened lungs were noted for the males of the high dose and enlargement of the pituitary gland was noted for the females of the intermediate (8 mg/kg) and high dosed (20 mg/kg) group. No histopathological correlate could be established for the pituitary.

Organ weights

Increased absolute kidney weights were noted for the females and decreased absolute liver weight were noted for the males treated with 20 mg via the diet.

Bone marrow evaluation

The myeloid:erythroid ratio of the male animals treated with 20 mg via the diet was significantly increased (Control: 0.993 : 1, Group 4: 1.847 : 1).

Histopathology

The male and female animals treated with 8 or 20 mg via the diet showed a dose dependent increase of lymphohistiocytic inflammatory reactions in the kidney, skeletal muscle and heart. The inflammation was associated with degenerative changes of the tubular epithelial cells of the kidneys and of the myofibers of heart and skeletal muscles.

The suppurative inflammation was more pronounced in the prostate and mesenteric lymph nodes of the animals treated with 20 mg via the diet compared to the control animals and the other test item-treated groups.

Granulomas with central neutrophilic granulocytes were only noted in the mesenteric lymph node of male and female rats of the high dose group.

A dose dependent increase of large macrophages with cytoplasmatic vacuoles was noted in the mesenteric lymph nodes of the animals treated with 8 or 20 mg via the diet.

Furthermore, the lungs of the male and female animals treated with 8 or 20 mg via the diet showed an increased incidence of foci of foamy macrophages (alveolar histiocytosis) in the alveoli of the lungs.

 

Conclusion:

LOAEL: The first effects as appeared in the mid-dose group (8 mg/kg) were:

-  an enlarged pituitary gland for 3 of 10 females without histopathological correlate.

-  increase ASAT by 122% for the males (test week 52).

-  large macrophages with cytoplasmatic vacuoles was noted in the mesenteric lymph nodes

-  increased incidence of foci of foamy macrophages (alveolar histiocytosis) in the alveoli of the lungs

-  increase of lymphohistiocytic inflammatory reactions in the kidney, skeletal muscle and heart.

Based on the present results the no-observed-effect-level (NOEL) was 4 mg N-(3-aminopropyl)-N-dodecylpropane-1,3-diamine/kg b.w. via the diet.

This information is used in a read-across approach in the assessment of the target substance. For details and justification of read-across please refer to the read-across report attached to IUCLID section 0.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

NOAEL

4 mg/kg bw/day

Study duration:

chronic

Species:

rat

Quality of whole database:

Guideline study

Repeated dose toxicity: inhalation - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: inhalation - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - systemic effects

Endpoint conclusion

Endpoint conclusion:

no study available

Repeated dose toxicity: dermal - local effects

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Data is based on read across from astructurally similar branched triamine C12.Branched triamine C16-18 only differs in the length of the alkyl chain, which issuspected to influence bioavailability, but not chemical reactivity and route of metabolisation, aspects that influence specific mechanisms of toxicity. For these reasons, many of the studies can best be performed on the substance with the shortest chain length within a group of structurally similar substances, as this is considered to result to the lowest NOAEL or most likely able to show specific effects..

Oral: The 90-day feeding study in rats, performed in accordance with EU Method B.26 and under GLP (CIT, 2003) resulted in a NOAEL of 100 ppm of the test substance in diet, which corresponded to ca. 7 mg/kg bw/day ingested substance (7 mg/kg bw/day for male and 8 mg/kg bw/day for female rats). Groups of 10 male and 10 female rats received 0, 100, 300 and 900 ppm of the test substance in diet for 90 days (corresponding to actual ingested doses of 7, 20 and 57 mg/kg bw/day for males and 8, 22 and 65 mg/kg bw/day for females). Animals were observed for clinical signs of toxicity, body weight changes and food consumption. In addition functional battery observations and haematological, clinical chemistry and

ophthalmological examinations were performed. All animals were sacrificed and subjected to gross necropsy and histopathological examinations.

The critical effects were tubular nephropathy of the kidney, associated with increased relative kidney weight, and the appearance of foamy macrophages in the mesenteric lymph nodes.

The 90-day dog study (LPT Laboratory of Pharmacology and Toxicology KG, 2004), in which animals received the test substance at dose levels of 0, 200, 500 and 1500 ppm in diet (corresponding to actual ingested doses of 0, 8.0, 20.0 and 55.1 (males) and 52.6 (females) mg/kg bw/day , resulted in a NOAEL of 500 ppm (20 mg/kg bw/day of ingested substance) in both males and females. At this dose level increased plasma ALAT and ASAT activities were noted; however, in the absence of pathological changes or signs of systemic toxicity they were considered to be non-adverse. At the next higher level of 1000 ppm (corresponding to 55.1 mg/kg bw/day and 52.6 mg/kg bw/day for female dogs) reduction in food consumption with a decrease in body weight was observed, as well as increased absolute and relative weight of the gall bladder, increased potassium and increased ALAT and ASAT activity in plasma.

Results from the rat and dog studies indicate species differences. In the rat study effects on the kidney were clearly present, while such effects were not observed in dogs. This is possibly related to higher metabolic rate in smaller animals, leading to higher uptake and renal excretion rates, and a subsequent higher concentration of the substance in the renal tubuli.

A combined chronic/carcinogenicity study in rats by dietary administration is currently running (LPT Laboratory of Pharmacology and Toxicology KG, 2008-2009). The chronic 1-year toxicity results from this study indicate a NOAEL of 4 mg/kg bw/day, reached from dosing mostly at a dietary concentration of 100 ppm. Based on dietary concentrations, the NOAEL from the chronic study is comparable to the NOAEL of the 90-day study. The next highest level in this study is spaced a factor 2 higher at 8 mg/kg bw/day and represents the LOAEL, at which signs of toxicity were a higher ASAT activity, increase of lymphohistiocytic inflammatory reactions in the kidney, skeletal muscle and heart, and foamy macrophages in the mesenteric lymph nodes and alveoli of the lungs. Additionally, an enlarged pituitary gland was observed in a few females without a histopathological correlate.

Dermal: In accordance with Column 2 of REACH Annex VIII, a repeated dose toxicity study using the most appropriate route of administration with regard to possible human exposure needs to be conducted. Data on repeated dose toxicity by oral route are available. As the substance is corrosive, and its dermal uptake is low (< 1% in 24 hr), dermal exposure is considered to be not relevant and the performance of a dermal repeated dose toxicity study is not indicated.

Inhalation: In accordance with Column 2 of REACH Annex VIII, a repeated dose toxicity study using the most appropriate route of administration with regard to possible human exposure needs to be conducted. Data on repeated dose toxicity by oral route are available. As the substance is not volatile (at 25 ºC vapour pressure is 0.00014 Pa), the inhalation route of exposure is considered not likely and the conductance of an inhalation repeated dose toxicity study is not indicated.

Justification for classification or non-classification

Data is based on read across from astructurally similar branched triamine C12 which is considered to be predictive, and even worst case for Branched triamine C16-18.

Based on the results of the 90 -day oral repeated dose toxicity studies with rats showing kidney pathology (Tubular nephropathy of the kidneys) at levels between 10 - 100 mg/kg bw/d (21 mg/kg bw), the substance needs to be classified as R48/22: danger of serious damage to health by prolonged exposure according to EU Directive 67/548/EEC. According to EU Classification, Labelling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008 the substance needs to be classified as Category 2, STOT, H373.