N-methoxy-1-(2,4,6-trichlorophenyl)propan-2-amine

Endpoint summary

Description of key information

Two GLP in-vitro studies for Skin irritation/corrosion were conducted with the substance according to OECD 431 and 439.

In the in vitro EPISKIN (SM) model test according to OECD 431, the results indicate that the test item is not corrosive. In the in vitro EPISKIN (SM) model test according to OECD 439 the mean cell viability was 42% compared to the negative control and therefore irritant.

One GLP in-vitro Eye irritation test in Isolated Chicken Eyes was conducted according to OECD 438 and one in-vivo acute eye irritation study in rabbits accordimg to OECD 405 was conducted with the test item. Mild irritation were noted in all animals in the in vivo study but all symptoms had fully reversed in all animals by the 72 -hour observation.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline study, available as unpublished report, no restrictions, fully adequate for assessment.

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: UN (2009), United Nations Globally Harmonized System of Classification and Labelling of Chemicals (GHS), third revised edition, UN New York and Geneva.

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: EpiSkin™ SOP, Version 1.8 (February 2009), ECVAM Skin Irritation Validation Study: Validation of the EpiSkin™ test method 15 min - 42 hours for the prediction of acute skin irritation of chemicals.

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Species:

other: Reconstructed human epidermis model EPISKIN (SM)

Details on test animals or test system and environmental conditions:

HUMAN SKIN: EPISKIN (SM) (Manufacturer: SkinEthic, France, Batch No.:13-EKIN-045, Expiry Date: 23 December 2013) is a three-dimensional human epidermis model.

EXPERIMENTAL DATES: From: 18 December 2013 To: 20 December 2013

Duration of treatment / exposure:

15 mins

Irritation / corrosion parameter:

other: other: optical density

Value:

0.311

Remarks on result:

other:

Remarks:

Basis: mean. (migrated information)

Irritation / corrosion parameter:

other: other: viability

Value:

42

Remarks on result:

other:

Remarks:

Basis: mean. (migrated information)

Irritant / corrosive response data:

The optical density for the test item treated skin was 0.311 and had a viability of 42%.
The optical densities for the negative and positive controls were 0.740 and 0.088, respectively and had viabilities of 100% and 12% respectively.

The test item is, therefore, considered to be irritant to skin, as the mean relative viability after 15 minutes exposure and 42 hours post incubation, is less or equal to 50% of the negative control. The positive control was irritant.

Table 1: Optical density and calculated % viability of cells

Substance	Optical density		Viability (%)
Negative control PBS	1	0.770	104
	2	0.728	98
	3	0.721	97
	mean	0.740	100
Positive control SDS 5%	1	0.100	14
	2	0.095	13
	3	0.070	9.5
	mean	0.088	12
Test item	1	0.259	35
	2	0.415	56
	3	0.259	35
	mean	0.311	42

 

Interpretation of results:

irritating

Remarks:

Migrated information Criteria used for interpretation of results: other: UN GHS

Conclusions:

In an in vitro EPISKIN (SM) model test, the results indicate that the test item is Irritant [UN GHS: Category 2].

Executive summary:

The reconstructed human epidermis model EPISKIN (SM) is designed to predict and classify the skin irritant potential of chemicals, by measuring its cytotoxic effect, as reflected in the MTT assay, on the EPISKIN reconstituted human epidermis. This method is approved by international regulatory agencies as a replacement for the identification of irritants / corrosives in the in vivo rabbit skin assay (OECD 404).

Disks of EPISKIN (three units / chemical) were treated with the test item and incubated for 15 minutes at room temperature. Exposure of test material was terminated by rinsing with phosphate buffered saline (PBS). Epidermis units were then incubated at 37°C for 42 hours in an incubator with 5% CO2. The viability of the epidermis on each disk was assessed by incubating the tissues for 3 hours with MTT solution at 37°C in 5% CO2 protected from light. The formazan extract in acidified isopropanol was then spectrophotometrically evaluated for optical density (OD) and quantified. SDS 5% and PBS treated epidermis were used as positive and negative controls, respectively. An additional disk was used to provide an estimate of colour contribution from the test item. For each treated tissue viability was expressed as a % relative to negative control. If the mean relative viability after 15 minutes exposure and 42 hours post incubation is less or equal (≤) to 50% of the negative control, the test substance is considered to be irritant to skin.

Following exposure with the test item, the mean cell viability was 42% compared to the negative control and, therefore, irritant. All validity criteria were within acceptable limits and therefore the study can be considered as valid.

In the in vitro EPISKIN (SM) model test, the results indicate that the test item is Irritant [UN GHS: Category 2].

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Remarks:

The in vivo eye irritation study was conducted solely to comply with a non-EU national registration requirement, and has been provided here in accordance with REACH, Article 22(1)e.

Type of information:

experimental study

Adequacy of study:

key study

Study period:

25 February 2014 to 06 March 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP compliant, guideline study, available as unpublished report, no restrictions, fully adequate for assessment.

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2400 (Acute Eye Irritation)

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Age at study initiation: 12-13 weeks
- Weight at study initiation: 3378-3688 g
- Housing: Individually housed in AAALAC approved open-wire metal cages.
- Diet: UNI diet for rabbits ad libitum
- Water: Tap water ad libitum
- Acclimation period: At least 20 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18.1-20.4
- Humidity (%): 28-68
- Air changes (per hr): 15-20
- Photoperiod (hrs dark / hrs light): 12 / 12

IN-LIFE DATES: From: 25 February 2014 To: 06 March 2014

Vehicle:

unchanged (no vehicle)

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL

Duration of treatment / exposure:

Single application (treated eye rinsed with physiological saline solution 1 hour after application).

Observation period (in vivo):

72 hours.

Number of animals or in vitro replicates:

3 males.

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Yes
- Time after start of exposure: 1 hour.

SCORING SYSTEM: Draize (1977) and OECD 405 (02 October 2012)

TOOL USED TO ASSESS SCORE: Fluorescein

Irritation parameter:

cornea opacity score

Basis:

animal #1

Remarks:

mean

Time point:

24/48/72 h

Score:

0.3

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritation parameter:

cornea opacity score

Basis:

animal #2

Remarks:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

cornea opacity score

Basis:

animal #3

Remarks:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

4

Irritation parameter:

iris score

Basis:

animal #1

Remarks:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

iris score

Basis:

animal #2

Remarks:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Irritation parameter:

iris score

Basis:

animal #3

Remarks:

mean

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: not applicable

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #1

Remarks:

mean

Time point:

other: 24-72 h

Score:

1.3

Max. score:

3

Reversibility:

fully reversible within: 72 h

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #2

Remarks:

mean

Time point:

24/48/72 h

Score:

1

Max. score:

3

Reversibility:

fully reversible within: 72 h

Irritation parameter:

conjunctivae score

Remarks:

redness

Basis:

animal #3

Remarks:

mean

Time point:

24/48/72 h

Score:

0.7

Max. score:

3

Reversibility:

fully reversible within: 72 h

Irritation parameter:

chemosis score

Basis:

animal #1

Remarks:

mean

Time point:

24/48/72 h

Score:

0.3

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritation parameter:

chemosis score

Basis:

animal #2

Remarks:

mean

Time point:

24/48/72 h

Score:

0.3

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritation parameter:

chemosis score

Basis:

animal #3

Remarks:

mean

Time point:

24/48/72 h

Score:

0.3

Max. score:

4

Reversibility:

fully reversible within: 48 h

Irritant / corrosive response data:

Conjunctival redness (score 2), chemosis (score 1) and discharge (score 3) were seen in all rabbits 1 hour after treatment. Conjunctival redness (score 1 or 2), chemosis (score 1) and discharge (score 1 or 2 or 3) were seen in all rabbits and corneal opacity (score 1, area 3) was noted in one rabbit 24 hours after treatment. Conjunctival redness (score 1 or 2) and discharge (score 1) were seen in all rabbits 48 hours after treatment. All symptoms had fully reversed in all animals by the 72 hours observations. Fluorescein staining was positive in one rabbit at 24 hours after instillation. Fluorescein staining was negative in two animals at 24 and in all animals at 48 and 72 hours after instillation. Control eyes were symptom-free during the study.

Other effects:

None.

Table 1: Eye irritation scores according to the Draize scheme

Time		Cornea			Iris			Conjunctiva
								Redness			Chemosis
	Animal number	719	718	889	719	718	889	719	718	889	719	718	889
after 1 hour		0	0	0	0	0	0	2	2	2	1	1	1
after 24 hours		1	0	0	0	0	0	2	2	1	1	1	1
after 48 hours		0	0	0	0	0	0	2	1	1	0	0	0
after 72 hours		0	0	0	0	0	0	0	0	0	0	0	0
mean scores 24-72 hours		0.3	0	0	0	0	0	1.3	1.0	0.7	0.3	0.3	0.3

 

Interpretation of results:

not classified

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The test item was graded as a mild irritant (Class 4 on a 1 to 8 scale) to the rabbit eye according to the modified Kay and Calandra classification system.

Executive summary:

The test item was administered as a single dose of 0.1 mL into the conjunctival sac of the left eye of each of 3 young adult male New Zealand White rabbits, the untreated right eye served as a control. Irritation was scored approximately 1, 24, 48 and 72 hours after administration. Observations with fluorescein staining were made approximately 24 hours before and 24, 48 and 72 hours after treatment. Rabbits were treated with analgesic and anaesthetic as per the regulatory guideline.

Conjunctival redness (score 2), chemosis (score 1) and discharge (score 3) were seen in all rabbits at 1 hour after treatment. Conjunctival redness (score 1 or 2), chemosis (score 1) and discharge (score 1 or 2 or 3) were seen in all rabbits and corneal opacity (score 1, area 3) was noted in one rabbit at 24 hours after treatment. Conjunctival redness (score 1 or 2) and discharge (score 1) were seen in all rabbits at 48 hours after treatment. All symptoms had fully reversed in all animals by the 72 hours observations. Fluorescein staining was positive in one rabbit at 24 hours after instillation. Fluorescein staining was negative in two animals at 24 and in all animals at 48 and 72 hours after instillation. The control eye of animals was symptom-free during the study. No mortality occurred during the study. The bodyweights of all rabbits were considered to be within the normal range of variability.

The test item was graded as a mild irritant (Class 4 on a 1 to 8 scale) to the rabbit eye according to the modified Kay and Calandra classification system.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the above stated assessment of the skin irritation/corrosion potential, N-methoxy-1 -(2,4,6 -trichlorophenyl)propan-2 -amine does have to be classified for Skin Irritation Category 2 (H315 Causes skin irritation) according to CLP (Regulation (EC) No 1272/2008 Of the European parliament and of the Council.

N-methoxy-1 -(2,4,6 -trichlorophenyl)propan-2 -amine does not have to be classified for Eye irritation according to CLP (Regulation (EC) No 1272/2008 Of the European parliament and of the Council.