N-methyldidecylamine

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

from 2010-05-11 to 2010-05-13

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2010

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: OECD Guideline for the Testing of Chemicals No. 431 "In Vitro Corrosion: Human *skin Model Test" (adopted 13 April 2004).

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

N-methyldidecylamine

EC Number:

230-990-1

EC Name:

N-methyldidecylamine

Cas Number:

7396-58-9

Molecular formula:

C21H45N

IUPAC Name:

bis(decyl)(methyl)amine

Details on test material:

- Expiration date of the lot/batch: 11 February 2011
- Storage conditions of test material: room temperature in the dark under nitrogen.

Test animals

Species:

other: reconstituted human epidermis model

Details on test animals or test system and environmental conditions:

The EPISKINTM model is a three-dimensional reconstituted human epidermis model consisting of adult human-derived epidermal keratinocytes
seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. A highly differentiated and stratified epidermis
model is obtained after a 13-day culture period comprising the main basal, supra basal, spinous and granular layers and a functional stratum
corneum.

Time schedule of the test: from 18 May 2010 to 21 May 2010.

Test system

Vehicle:

unchanged (no vehicle)

Controls:

other: tissues were treated with the negative control material

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 50 µl of the test material was applied topically to the corresponding tissues ensuring uniform coverage of the tissues.

Duration of treatment / exposure:

Duplicate tissues were treated with the test material for exposure periods of 3, 60 and 240 minutes (duplicate tissues were treated with the positive
and negative control materials for an exposure period of 240 minutes).

Observation period:

Not applicable

Number of animals:

Not applicable.

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): At the end of each exposure period, each tissue was removed from the well using forceps and rinsed using a wash bottle
containing Phosphate Buffered Saline Dulbeccos (PBS) with Ca++ and Mg++.

SCORING SYSTEM:

Classification of corrosivity potential was based on relative viabilities for each exposure time: see table in the following section.

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

other: other: cell vialibility in percentage (based on optical density at 540 nm)

Value:

129.5

Remarks on result:

other:

Remarks:

Basis: mean. Time point: 3 minutes. Reversibility: other: not applicable. (migrated information)

Results 2

Irritation / corrosion parameter:

other: other: cell vialibility in percentage (based on optical density at 540 nm)

Value:

98.4

Remarks on result:

other:

Remarks:

Basis: mean. Time point: 60 minutes. Reversibility: other: not applicable. (migrated information)

Results 3

Irritation / corrosion parameter:

other: other: cell vialibility in percentage (based on optical density at 540 nm)

Value:

82.9

Remarks on result:

other:

Remarks:

Basis: mean. Time point: 240 minutes. Reversibility: other: not applicable. (migrated information)

Any other information on results incl. tables

Following the 3, 60 and 240‑Minute exposure periods the test material treated tissues appeared blue which was considered to be indicative of viable tissue. 

The relative mean tissue viability for the positive control treated tissues was 2.3% relative to the negative control treated tissues following the

240-Minute exposure period. The positive control acceptance criterion was therefore satisfied.

The mean OD value for the negative control was 0.168. The negative control acceptance criterion was therefore satisfied.

Mean OD₅₄₀Values and Viabilities for the Negative Control Material, Positive Control Items and Test Item

Material	Exposure Period	Mean OD540of duplicate tissues	Relative mean viability (%)
Negative Control Material	240 Minutes	0.129	100*
Positive Control Material	240 Minutes	0.003	2.3
Test Material	240 Minutes	0.107	82.9
	60 Minutes	0.127	98.4
	3 Minutes	0.167	129.5

*=     The mean viability of the negative control tissues is set at 100%

Applicant's summary and conclusion

Interpretation of results:

other: non-corrosive

Remarks:

Criteria used for interpretation of results: other: CLP (Reg. 1272/2008/EC) and Directive 67/548/EEC

Conclusions:

Under these experimental conditions, the test item was considered to be Non-Corrosive to the skin.

Executive summary:

The purpose of this test was to evaluate the corrosivity potential of the test material using the EPISKIN^TMin vitro Reconstituted Human Epidermis (RHE) Model after treatment periods of 3, 60 and 240 minutes. This method was designed to meet the requirements of the following guideline OECD Guideline for the Testing of Chemicals No. 431 “In Vitro Skin Corrosion: Human Skin Model Test” (adopted 13 April 2004)

The EPISKIN^TMmodel is able to distinguish between corrosive and non-corrosive chemicals for all of the chemical types studied. As stated in regulation 1272/2008/EEC a validated in vitro test such as in vitro skin corrosion test in the EPISKIN^TMmodel can be used for classification and labelling. The EPISKIN^TMmodel is also able to distinguish between known R35 (UN packing group I) and R34 (UN packing group II & III) chemicals.

Duplicate tissues were treated with the test material for exposure periods of 3, 60 and 240 minutes. At the end of the exposure period the test material was rinsed from each tissue before each tissue was taken for MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) loading. After MTT loading a total biopsy of each epidermis was made and placed into micro tubes containing acidified isopropanol for extraction of formazan crystals out of the MTT-loaded tissues. 

At the end of the formazan extraction period each tube was mixed thoroughly and duplicate 200 µl samples were transferred to the appropriate wells of a pre-labelled 96‑well plate. The optical density was measured at 540 nm.

Data are presented in the form of percentage viability (MTT reduction in the test material treated tissues relative to negative control tissues).

The relative mean viability of the test material treated tissues was:

240 minutes exposure	:	82.9%
60 minutes exposure	:	98.4%
3 minutes exposure	:	129.5%

The quality criteria required for acceptance of results in the test were satisfied.The test item was considered to be Non-Corrosive to the skin.