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

Endpoint summary

Description of key information

The LD50 of 766 mg/kg bw was established in acute oral toxicity study with rats. As the substance is corrosive to skin, the performance of studies by other routes of exposure is not indicated in accordance with Column 2 of REACH Annex VII.

Key value for chemical safety assessment

Acute toxicity: via oral route

Link to relevant study records

Reference

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 January 1985 - 2 July 1985

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was performed similar to OECD 425 with some additional investigations. The study was not performed under GLP but in-house quality assurance was in place.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 425 (Acute Oral Toxicity: Up-and-Down Procedure)

GLP compliance:

no

Remarks:

in-house QA was in place

Test type:

up-and-down procedure

Limit test:

no

Species:

rat

Strain:

other: CFY (Sprague-Dawley Origin)

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Interfauna UK Ltd., (formerly Hacking and Churchill Limited,) England
- Age at study initiation: 4-6 weeks
- Weight at study initiation: 91-150g
- Fasting period before study: overnight
- Housing: They were housed in groups by sex or single in metal cages with wire mesh floors.
- Diet (e.g. ad libitum): ad libitum, standard laboratory rodent diet (Labsure LAD 1)
libitum. The batch
- Water (e.g. ad libitum): ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-22
- Humidity (%): 56
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12


IN-LIFE DATES: From: 22 January 1985 To: 2 July 1985

Route of administration:

oral: gavage

Vehicle:

other: unchanged or in distilled water

Details on oral exposure:

Administered as supplied or as 25% w/v in distilled water.
VEHICLE
- Concentration in vehicle: 25% w/v
- Amount of vehicle (if gavage): 0.1 ml/kg
- Justification for choice of vehicle: no data
- Lot/batch no. (if required): no data
- Purity: no data

MAXIMUM DOSE VOLUME APPLIED: unchanged: 2.3 ml/kg in distilled water: 0.1 ml/kg

Doses:

Stage I: 2000, 200, 25 mg/kg bw.
Stage II: males, 640, 800, 1000, 1260 mg/kg bw
females, 320, 400, 500, 640, 800, 1000 mg/kg bw

Stage III:
320, 200, 80 mg/kg bw

No. of animals per sex per dose:

Stage I: 2
Stage II: One animal per dosing occasion.
Stage III: 5

Control animals:

other: Not in stage I and II but at stage III a control group was included.

Details on study design:

Stage I, II and III
- Duration of observation period following administration: 14 days
- Frequency of observations and weighing: Animals were observed soon after dosing; then at frequent intervals for the remainder of Day 1. On
subsequent days the animals were observed at least twice per day. Clinical signs were recorded at each observation. Individual bodyweights of rats on Days 1 (day of dosing), 4, 8 and 15 and at death.
- Necropsy of survivors performed: yes
- Other examinations performed: other: only in stage III
On Days 2 and 7 samples of blood were withdrawn, under light ether anaesthesia, from the orbital sinus of all rats. The blood samples were placed in EDTA anti-coagulant tubes for subsequent; haernatological investigations. The following estimations were performed:
Total white cell count (WBC Total), Differential WBC counts - standard, microscopy of blood smear, stained with modified Wright's stain counting 100 cells, Neutrophils (N), Lymphocytes (L), Eosinophils (E), Basophils (B), Monocytes (M).

Statistics:

The LD50 and its 95% confidence limits were estimated for both males and females separately, based on the method of Brownlee, Hodges and
Rosenblatt. For haematological data statistical analysis was carried out using the individual animal as the basic experimental unit. The following sequence of statistical tests was used for haematological parameters in which the relative frequency of the mode was less than 75% (i.e. the data did not consist predominant of one particular value):

(i) Bartlett's test was applied to test for heterogeneity of variance between treatments. Where significant (at the 1% level) heterogeneity was found, a log transformation was tried to see if the heterogeneity was removed.
(ii) If no satisfactory transformation was found the Kruskal-Wallis analysis of ranks was used.
(iii) Analyses of variance were followed by Student's 't' test for a-dose-related response. The Kruskal-Wallis analyses were followed by the non-parametric equivalent of the, ' t' test.

For those parameters in which the relative frequency of the mode is 75% or more, the number of animals in each group with values different
from the mode will be analysed using Fisher's exact test and Mantel's test. Fisher's test will be used to detect general differences between treatments, whereas Mantel's test will be used specifically for detecting dose-related trends in the numbers of such animals.

Sex:

male/female

Dose descriptor:

LD50

Effect level:

> 200 - < 2 000 mg/kg bw

Remarks on result:

other: Stage I

Sex:

male

Dose descriptor:

LD50

Effect level:

895 mg/kg bw

95% CL:

657 - 1 219

Remarks on result:

other: Stage II

Sex:

female

Dose descriptor:

LD50

Effect level:

637 mg/kg bw

95% CL:

467 - 869

Remarks on result:

other: Stage II

Sex:

male/female

Dose descriptor:

other: Haematological changes

Remarks on result:

other: No effects observed after a single dose of 80 mg/kg bw on day 2 and day 7 after treatment.

Mortality:

Stage I:
All rats dosed at 2000 mg/kg were found dead within one or two days of treatment.

Stage II:
Death occurred amongst male rats at 800 mg/kg and above and amongst female rats at 500 mg/kg and above within one to four days of dosing.

Stage III:
Based on the results from stage I and II rats were dosed at the expected maximum sub-lethal dose.
The treatmnet started with 320 mg/kg bw but one female rat was found dead within one day. A further group of ten rats was therefore dosed at a lower level of 200 mg/kg. Two male rats were found dead within. one day of dosing. A third group of ten rats was then dosed at 80 mg/kg and no mortalities occured.

Clinical signs:

other: Stage I: Signs of reaction to treatment observed shortly after dosing in all rats were pilo-erection, hunched posture and abnormal gait (waddling). These signs were accompanied by: increased salivation in all rats at 200 and 2000 mg/kg, pallor of extremit

Gross pathology:

Stage I:
In the animals that died at 2000 mg/kg bw autopsy revealed congestion or haemorrhage of the lungs and pallor o f the liver, spleen and kidneys. In the surviving animals no abnormalities were observed.

Stage II:
Autopsy of the animals that died revealed congestion of the lungs and pallor of the liver, spleen and kidneys. Pallor of the kidney cortex was also observed in one male rat at 1000 mg/kg. In the surviving animals no abnormalities were observed.

Stage III:
No abnormalities were observed.

Other findings:

Haematology:
No statistically significant effects were noted.

Stage I:

Time and number of deaths:

sex	Dose mg/kg bw	No. of deaths	Day 1	2 a	2b	3a	3b	4-15
m	25	0
m	200	0
m	2000	2		1		1
f	25	0
f	200	0
f	2000	2		1	1

a = first observation, b = second observation, m = male, f = female

Stage II:

Mortality data:

		mortality data on each dosing occasion
sex	dose (mg/kg)	1	2	3	4	5	6	7	8	9
m	640								0
m	800	0				0		1*
m	1000		0		1*		1*
m	1260			1*
f	320				0
f	400					0
f	500			1*			0
f	640		1*					0
f	800	1							0
f	1000									1*

1 = death, 0 = survival, * = late death

Time and number of deaths:

Day

Sex

Dose (mg/kg)

No. death/No. dosed

1

2a

b

3a

b

4 a

b

5a

b

6 -15

m

640

0/1

m

800

1/3

1

m

1000

2/3

1

1

m

1260

1/1

1

f

320

0/1

f

400

0/1

f

500

1/2

1

f

640

1/2

1

f

800

1/2

1

f

1000

1/1

1

a = first observation, b = second observation, m = male, f = female

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The LD50 in rats was calculated to be 766 mg/kg bw for males and females combined.

Executive summary:

An acute oral toxicity study was performed with "Lilamin LS 33". Animals were administered the test substance by oral gavage. The study was divided in three stages. Stage on was a range finding study were rats (2/sex dose) were given a single dose of 2000, 200 or 25 mg/kg bw. Based on these results an up-and-down procedure was followed dosing 1 animal at a time to determine a more exact LD50. In phase III, based on the LD50 findings, 10 animals (5/sex) were given a dose of 80 mg/kg bw and haematological parameters were measured 2 and 7 days after dosing. The LD50 in rats is between 200 and 2000 mg/kg bw, 895 (CI 657-1219) mg/kg bw for males and 637 (CI 467-869) mg/kg bw for females. Based on the performed acute oral toxicity studies the test substance should be classified as harmful (Xn and R22). No effects on haematology were observed after a single oral dose of 80 mg/kg bw.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed

Dose descriptor:

LD50

Value:

766 mg/kg bw

Acute toxicity: via inhalation route

Link to relevant study records

Reference

Endpoint:

acute toxicity: inhalation

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because the substance is classified as corrosive to the skin

other:

Endpoint conclusion

Endpoint conclusion:

no study available

Acute toxicity: via dermal route

Link to relevant study records

Reference

Endpoint:

acute toxicity: dermal

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because the substance is classified as corrosive to the skin

other:

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Oral

Branched triamine C16 -18 (Y-tallow) was tested for acute oral toxicity by Acute Up-and-Down Procedure (OECD 425) with some additional investigations. Following an initialrange finding study in which rats (2/sex dose) were dosed at 2000, 200 or 25 mg/kg bw, an up-and-down procedure was followed dosing 1 animal at a time to determine a more exact LD50. Then, based on the LD50 findings, 10 animals (5/sex) were given a dose of 80 mg/kg bw (maximum non-lethal dose, as at 320 and 200 mg 1 or 2 of the 10 dosed animals died) and haematological parameters were measured 2 and 7 days after dosing. The LD50 in rats is between 200 and 2000 mg/kg bw, 895 (CI 657-1219) mg/kg bw for males and 637 (CI 467-869) mg/kg bw for females, and766 mg/kg bw for males and females combined. No effects on haematology were observed after a single oral dose of 80 mg/kg bw.

 

Comparable other triamines (alkyl dipropylenetriamine) and tetramines (alkyl tripropylenetatramine) show similar results, where those with on average shorter alkyl chains show a somewhat higher toxicity compared to those with longer alkyl chain lengths:

 

LD50 in mg/kg bw

Acute toxicity Triamines:
Coco	50-300	Cat.3; cut off 200
Tallow	300-2000	Cat.4; cut of 500
Oleyl	300-2000	Cat.4; cut of 500
Y12	261	Cat.3
YTallow	766	Cat.4
Acute toxicity Tetramines:
Tallow	300-2000	Cat.4; cut of 500
Oleyl	300-2000	Cat.4; cut of 1000

 

In conclusion, there is little effect of the actual number propylene amine groups between linear triamine, branched triamine or linear tetramine. With increasing length of the alkyl chain, the acute toxicity decreases.

 

Dermal

Acute dermal toxicity: Polyamines are corrosive to the skin. Testing for acute dermal toxicity is therefore not justified. Toxicity following dermal exposure is characterised by local tissue damage, rather than the result of percutaneously absorbed material.

The mode of action of for polyamines follows from its structure, consisting of an apolar fatty acid chain and a polar end of a primary amine linked to a secondary amine. The structure can disrupt the cytoplasmatic membrane, leading to lyses of the cell contentand consequently the death of the cell.

The polyamines are completely protonated under environmental conditions which causes them to strongly adsorb to organic matter. This all leads to a low dermal absorption as is shown by a dermal absorption study performed on astructurally related branched triamine N-(3-aminopropyl)-N-dodecylpropane-1,3-diamine (branched triamine C12)for 24 hours, resulted in a dermal penetration of less than 0.01% whereas 0.92% of the applied dose did pass the stratum corneum but remained further fixed in the skin. For thebranched triamine C16 -18,a similar or even lower (because of higher alkyl chain lengths) dermal penetration can be expected.

 

Inhalation:

Acute inhalation toxicity: Physical-chemical properties of polyamines indicate a low likelihood for exposure via inhalation having a boiling point > 300 °C and a low vapour pressure (1.4 x 10-4 Pa at 25°C for thebranched triamine C12, with the shortest average alkyl chain length representing the highest vapour pressure for the group of branched polyamines).

Furthermore, the substance is classified as severe corrosive and no acute toxicity testing should normally be conducted.

Justification for classification or non-classification

According to the criteria in Directive 67/548/EEC, based on the LD50 of 766 mg/kg bw,Branched triamine C16 -18needs to be classified as Xn; R22 (Harmful if swallowed). According to EU Classification, Labeling and Packaging of Substances and Mixtures (CLP) Regulation (EC) No. 1272/2008 it needs to be classified as Category IV, H301 (Harmful if swallowed).

 

Acute dermal testing with these very corrosive materials is not justified. As a consequence no classification can be made for acute dermal toxicity. Effects will be characterised by local tissue damage. Systemic uptake via skin is very limited and absorption studies indicated that dermal absorption is at least a factor three lower than oral absorption. The very corrosive effects on skin in combination with the relatively low acute oral toxicity, make systemic effects from acute dermal exposures are unlikely.

 

Also for acute inhalation toxicity information for classification is lacking, and is testing not justified. Due to very low vapour pressure is the likelihood of exposure low. Condidering the relatively low systemic toxicity after acute oral dosing, suggest that inhalation of this very corrosive substance at levels at levels causing systemic effects is very unlikely.

 

Polyamines do not contain containing aliphatic, alicyclic and aromatic hydrocarbons and so do not indicate an immediate concern for aspiration hazard.