Amides, tall-oil fatty, N,N'-(iminodi-2,1-ethanediyl)bis-, maleated

Endpoint summary

Description of key information

For the test substance negative in vitro studies on skin and eye irritation and skin corrosion are available. For the  analogue negative in vivo studies are available for skin and eye irritation (see rationale for read-across chapter 13).

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

Study period:

12-06-2013 to 17-06-2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: study performed according to the guideline and under GLP

Qualifier:

according to guideline

Guideline:

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

Qualifier:

according to guideline

Guideline:

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

Principles of method if other than guideline:

exposure period 15 minutes with post-exposure incubation period of 42 hours

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Control samples:

yes, concurrent negative control

Details on study design:

The EpiSKIN assay uses reconstructed human epidermis to assess skin irritation. For the viability test enzymatic conversion of MTT to formazan is used.

In a pre-test direct interaction of the test substance with the detection chemical MTT is assessed.
In the main study triplicate tissues were treated with 30 uL of the test substance during 15 minutes. In addition a negative control (DPBS) and a positive control (5% SDS) were included. After treatment tissues were rinsed, incubated in maintenance fluid for 42 hours at 37 °C and treated with MTT during 3 hours (at 37 °C). After MTT loading a total biopsy of each epidermis (epidermis and collagen) was made, that was suspended in acidified iso-propanol for 3 days and kept in the refrigerator at 1-10°C. Thereafter the optical density of the extracted MTT solutions was measured at 562 nm. Viability was expressed as percentage MTT conversion versus negative control.
Tissue viability > 50% means non-irritant
Tissue viability ≤ 50% means irritant

Irritation / corrosion parameter:

% tissue viability

Value:

81.8

Negative controls validity:

valid

Positive controls validity:

valid

Remarks:

rel viability 10.6%

Other effects:

The test substance did not interfere with the reduction of MTT to blue formazan salt.

	Mean OD562 ± SD	Relative mean viability ± SD
Negative control:	0.946 ± 0.119	100 ± 12.6%
Test substance:	0.774 ± 0.021	81.8 ± 2.2%
SDS:	0.100 ± 0.014	10.6 ± 1.5%

Interpretation of results:

GHS criteria not met

Conclusions:

The test substance is non-irritant in the EPISKIN reconstructed human epidermal model.
Criteria used for interpretation of results: other: tissue viability > 50%: non-irritant

Executive summary:

In this assay triplicate reconstructed human skin tissues were treated with 30 uL of the test substance during 15 minutes. After treatment tissues were rinsed, kept for 42 hours and treated with MTT. Optical density of extracted MTT solutions was measured at 562 nm. Viability was expressed as percentage MTT conversion versus negative control. The viability was 81.8%, which is indicative for non-irritant substances.

The test substance is non-irritant in the EPISKIN reconstructed human epidermal model.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

17-07-2013 to 18-07-2013

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: not formally validated study method

Principles of method if other than guideline:

The SkinEthic reconstructed human corneal epithelium assay is used

GLP compliance:

yes (incl. QA statement)

Details on study design:

The SkinEthic reconstructed human corneal epithelium assay uses corneal epithelial tissue to assess penetration and concomittant cytotoxicity of the test substance.

In a pre-test direct interaction of the test substance with the detection chemical MTT is assessed.
In the main study triplicate SkinEthic tissues were treated with 30 uL of the test substance during 10 minutes. In addition a negative control (Na2HPO4 0.142 g/L; glucose 1.802 g/L; HEPES 7,149 g/L; KCl 0.224 g/L; NaCl 7.597 g/L) and a positive control (2% SDS) were included. After treatment tissues (3 per treatment) were rinsed and treated with MTT. Optical density of extracted MTT solutions was measured at 562 nm. Viability was expressed as percentage MTT conversion versus negative control.
Tissue viability ≥ 60% means non-irritant
Tissue viability < 60% means irritant

Irritation parameter:

other: % tissue viability

Value:

89.4

Negative controls validity:

valid

Positive controls validity:

valid

Remarks:

6.7% rel viability

Other effects:

The test substance did not interfere with the reduction of MTT to blue formazan salt.

	Mean OD562	Relative mean viability
Negative control:	0.940	100%
Test substance:	0.840	89.4%
SDS:	0.063	6.7%

Interpretation of results:

GHS criteria not met

Conclusions:

The test substance is non-irritant in the SkinEthic reconstructed human corneal epithelium assay.
Criteria used for interpretation of results: other: < 60% viability is indicative for positive result

Executive summary:

In this assay triplicate SkinEthic tissues were treated with 30 uL of the test substance during 10 minutes. After treatment tissues were rinsed and treated with MTT. Optical density of extracted MTT solutions was measured at 562 nm. Viability was expressed as percentage MTT conversion versus negative control. The viability was 89.4%, which is indicative for non-irritant substances.

As the assay is not formally validated, the results need to be regarded as preliminary.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation/corrosion

In the EPISKIN assay triplicate reconstructed human skin tissues were treated with 30 uL of the test substance during 15 minutes. After treatment tissues were rinsed, kept for 42 hours and treated with MTT. Optical density of extracted MTT solutions was measured at 562 nm. Viability was expressed as percentage MTT conversion versus negative control. The viability was 81.8%, which is indicative for non-irritant substances (Harlan 2013g).

In the EpiSKIN test, duplicate reconstructed human skin tissues were treated with 50 uL of the test substance during 3, 60 and 240 minutes. After treatment tissues were rinsed with PBS and incubated with MTT for 3 hours (at 37°C, 5% CO2 in air). Optical density of homogene solutions of MTT treated tissues was measured at 540 nm. Viability was expressed as percentage MTT conversion versus negative control. The viability was 96.9 -107.4% for the different exposure times, which is indicative for non-corrosive substances (Harlan 2013h).

Eye irritation

In SkinEthic assay tissues (in triplicate) were treated with 30 uL of the test substance during 10 minutes. After treatment tissues were rinsed and treated with MTT. Optical density of extracted MTT solutions was measured at 562 nm. Viability was expressed as percentage MTT conversion versus negative control. The viability was 89.4%, which is indicative for non-irritant substances (Harlan 2013i).

Justification for classification or non-classification

The test substance does not need to be classified for skin and eye irritation.