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

Acute Toxicity: oral

Administrative data

Endpoint:

acute toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Study period:

26 April 2012 - 21 June 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 423 (Acute Oral toxicity - Acute Toxic Class Method)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.1 tris (Acute Oral Toxicity - Acute Toxic Class Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test type:

acute toxic class method

Limit test:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Amines,N-(C16-18 and C18 unsaturated alkyl) trimethylenedi-,diacetates.

EC Number:

800-153-0

Cas Number:

1313206-64-2

Molecular formula:

R-NH2+ -(CH2)3-NH3+, (CH3 COO-)2 where R = C16-18 and C18-unsat alkyl

IUPAC Name:

Amines,N-(C16-18 and C18 unsaturated alkyl) trimethylenedi-,diacetates.

Test material form:

liquid

Remarks:

yellow liquid

Details on test material:

Chemical registery number : 1313206-64-2
Chemical name : Amines, N-(C16-18 and C18-unsatd. alkyl)trimethylenedi-, diacetates

Based on the qualitative and quantitative information on the composition, the sample used are representative of the boundary composition shared and agreed by each registrant.

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: breeder: Janvier, Le Genest-Saint-Isle, France
- Age at study initiation: approximately 8 weeks old on the day of treatment
- Mean body weight at study initiation: 210 g (range: 202 g to 219 g)
- Fasting period before study: yes, during the night before treatment
- Housing: polycarbonate cages with stainless steel lids
- Diet: SSNIFF R/M-H pelleted diet (free access)
- Water: tap water filtered with a 0.22 µm filter (free access)
- Acclimation period: at least 6 days before treatment.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2°C
- Humidity (%): 50 ± 20%
- Air changes (per hr): approximately 12 cycles/hour of filtered, non-recycled air
- Photoperiod (hrs dark / hrs light): 12 h/12 h.

IN-LIFE DATES: 16 May 2012 to 14 June 2012.

Administration / exposure

Route of administration:

oral: gavage

Details on oral exposure:

VEHICLE
- Concentration in vehicle: 30 and 200 mg/mL
- Justification for choice of vehicle:
The vehicle used in this study was selected from the results of solubility assays performed by the CiToxLAB France Pharmacy.
In the absence of recommendation from the Sponsor, the solubility assay first started at the concentration of 200 mg/mL, and the first choice vehicle was drinking water treated by reverse osmosis using an Elix 5 plus apparatus (Millipore SA, Saint-Quentin-en-Yvelines, France).
After 10 minutes under magnetic stirring, a yellow solution was obtained at the concentration of 200 mg/mL in this vehicle.
- Maximum dose-volume applied: 20 mL/kg

DOSAGE PREPARATION (if unusual):
The test item was administered as a solution in the vehicle. The test item was mixed with the required quantity of vehicle and then homogenized using a magnetic stirrer.
Only glass was used during preparation.
Dose formulations preparations were prepared by the CiToxLAB France Pharmacy extemporaneously on the day of each administration.
The dose formulations were stored at room temperature and delivered to the study room in brown glass flasks.

CLASS METHOD (if applicable):
- Rationale for the selection of the starting dose:
Since some discrepancies were observed when the estimation of a lethal dose-level had been performed with the test item with solvent, it was decided to administer the test item without solvent at the starting dose level of 300 mg/kg for animal welfare reasons.
After treatment at the starting dose-level, the next dose-levels were administered in a sequential manner, under the same conditions to different animals, based to the flow charts of OECD Guideline No. 423, 17th December 2001, which are equivalent to those of Commission Regulation (EC) No. 440/2008, B.1tris (30 May 2008). The selected dose-levels corresponded to the current limit values of classification of chemical substances. As no mortality was observed after treatment of the first group at the dose-level of 300 mg/kg, the second group was treated at the dose-level of 2000 mg/kg.
As mortality was observed after treatment of the second group at the dose-level of 2000 mg/kg, the third group was treated at the dose-level of 300 mg/kg.

Doses:

300 and 2000 mg/kg.

No. of animals per sex per dose:

3 females per treatment step

Control animals:

no

Details on study design:

- Duration of observation period following administration: 14 days
- Clinical observations: frequently during the hours following treatment; then, at least once a day.
- Body weight: just before treatment on day 1; then on days 8 and 15.
- Necropsy of survivors performed: yes (macroscopic).

Results and discussion

Effect levels

Sex:

female

Dose descriptor:

LD50

Effect level:

> 300 - < 2 000 mg/kg bw

Based on:

test mat.

Mortality:

At 2000 mg/kg, 1/3 females was found dead on day 2. Prior to death, piloerection was observed on day 1 (0h30 minutes after treatment).
One other female from this group was prematurely sacrificed on day 7. Prior to sacrifice, piloerection, thin appearance, soiled urogenital region, hypothermia (cold to the touch) and hypoactivity were observed.

Clinical signs:

other: At 300 mg/kg (first assay), all animals presented piloerection during the first 4 hours after treatment. At 2000 mg/kg, piloerection, soiled urogenital region, thin appearance, hypoactivity and hunched posture were recorded in the surviving female. At 30

Gross pathology:

No gross findings were attributed to treatment with the test item.

Any other information on results incl. tables

see Executive summary

Applicant's summary and conclusion

Interpretation of results:

Category 4 based on GHS criteria

Conclusions:

The oral LD50 of the test item was comprised between 300 and 2000 mg/kg in rats.
According to the criteria of Regulation (EC) No 1272/2008, the test item should be classified category 4 and assigned the signal word "warning" and the hazard statement "H302: Harmful if swallowed".

Executive summary:

The objective of this study was to evaluate the potential acute toxicity of the test item following a single oral administration (gavage) to rats.

 

Methods

The test item was administered once by oral route (gavage) to three groups of three fasted female Sprague-Dawley rats under a dosage-volume of 10 mL/kg. The test item was prepared in drinking water treated by reverse osmosis.

Since some discrepancies were observed when the estimation of a lethal dose-level had been performed with the test item with solvent, it was decided to administer the test item without solvent at the starting dose-level of 300 mg/kg for animal welfare reasons.

After the first assay, the next higher dose-level of 2000 mg/kg was tested. Then, as mortality was observed after treatment of the second group at the dose-level of 2000 mg/kg, the third group was treated at the dose-level of 300 mg/kg.

Each animal was observed once a day for mortality and clinical signs for 15 days. Body weight was recorded on day 1 and then on days 8 and 15.

On completion of the observation period, the animals were sacrificed and then submitted for a macroscopic post-mortem examination.Macroscopic lesions were preserved in buffered formalin, then destroyed at the finalization of the study report as no microscopic examination was performed.

 

Results

At 2000 mg/kg, 1/3 females was found dead on day 2. Piloerection was observed prior to death. Piloerection, soiled urogenital region, thin appearance, hypoactivity and hunched posture were recorded in the surviving female.

One other female from this group was prematurely sacrificed on day 7. Prior to sacrifice, piloerection, thin appearance, soiled urogenital region, hypothermia (cold to the touch) and hypoactivity were observed.

At 300 mg/kg (first assay), all animals presented piloerection during the first four hours after treatment.

At 300 mg/kg (confirmatory assay), piloerection was noted in all females. Hypoactivity, hunched posture, loud and abdominal breathing, half-closed eyes, thin appearance, increase in size of abdomen and pallor of extremities were recorded in 1/3 females.

When compared to CiToxLAB historical control data, a lower body weight gain was noted at 300 mg/kg (first assay) in two female between day 1 and day 15.

A body weight loss and lower body weight gain was noted at 2000 mg/kg in the surviving female between day 1 and day 8 and between day 8 and day 15, respectively.

At 300 mg/kg (confirmatory assay), a body weight loss and lower body weight gain were noted in 1/3 females between day 1 and day 8 and between day 8 and day 15, respectively. A lower body weight gain was also noted in another female of this group between day 1 and day 8.

No gross findings were attributed to treatment with the test item.

 

Conclusion

The oral LD₅₀ of the test item was comprised between 300 and 2000 mg/kg in rats.

According to the criteria of Regulation (EC) No 1272/2008, the test item should be classified category 4 and assigned the signal word "warning" and the hazard statement "H302: Harmful if swallowed".