Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Toxicological information

Skin irritation / corrosion

Currently viewing:

Administrative data

Endpoint:
skin irritation / corrosion
Remarks:
other: in vitro
Type of information:
experimental study
Adequacy of study:
key study
Study period:
The study was performed between 30 March 2010 and 05 April 2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP conditions.
Justification for type of information:
A discussion and report on the read across strategy is given as an attachment in Section 13
Cross-reference
Reason / purpose for cross-reference:
read-across: supporting information
Reference
Endpoint:
skin irritation / corrosion, other
Remarks:
In Vitro
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
The study was performed between 30 March 2010 and 05 April 201
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see "Remark"
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP condition
Justification for type of information:
A discussion and report on the read across strategy is given as an attachment in Section 13
Reason / purpose for cross-reference:
read-across source
Qualifier:
according to guideline
Guideline:
other: EU Guideline Testing of Chemicals B46
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD Draft Test Guideline
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Test system:
human skin model
Remarks:
human skin model reconstituted human epidermis model
Source species:
human
Cell type:
other: reconsituted human epidermal cells
Justification for test system used:
ECVAM recommended in vitro method. Also EU Guideline B46
Vehicle:
unchanged (no vehicle)
Duration of treatment / exposure:
15 minutes
Duration of post-treatment incubation (if applicable):
42 hours
Number of replicates:
Three
Type of coverage:
other: Applied topically to reconstiyuted human epidermal cells
Vehicle:
unchanged (no vehicle)
Irritation parameter:
other: Viability of cells
Basis:
other: Mean viability of cells
Time point:
other: Day 6
Reversibility:
other: Not applicable
Remarks on result:
other: See relative mean viability below.
Irritant / corrosive response data:
The relative mean viability of the test material treated tissues was 93.1% after a 15 minute exposure.
Other effects:
no
Interpretation of results:
other: Not irritating
Remarks:
expert judgment
Conclusions:
The test material was considered to be Non-Irritant (NI)
Executive summary:

Introduction:

The purpose of this test was to evaluate the skin irritation potential of the test material using the EPISKINTM reconstituted human epidermis model after a treatment period of 15 minutes followed by a post-exposure incubation period of 42 hours. The principle of the assay was based on the measurement of cytotoxicity in reconstituted human epidermal cultures following topical exposure to the test material by means of the colourimetric MTT reduction assay. Cell viability is measured by enzymatic reduction of the yellow MTT tetrazolium salt (3 -[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to a blue formazan salt (within the mitochondria of viable cells) in the test material treated tissues relative to the negative controls. The concentration of the inflammatory mediator IL-1α in the culture medium retained following the 42 hour post-exposure incubation period is also determined for test materials which are found to be borderline non-irritant based upon the MTT reduction endpoint. This complimentary end-point will be used to either confirm a non-irritant result or will be used to override the non-irritant result.

Methods:

Triplicate tissues were treated with the test material for an exposure period of 15 minutes. At the end of the exposure period each tissue was rinsed before incubating for approximately 42 hours. At the end of the postexposure incubation period each tissue was taken for MTT-loading. The maintenance medium from beneath each tissue was transferred to pre-labelled micro tubes and stored in a freezer for possible inflammatory mediator determination. 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).

Results:

The relative mean viability of the test material treated tissues was 93.1% after a 15-minute exposure.

Quality criteria:

The quality criteria required for acceptance of results in the test were satisfied. Conclusion:  The test material was considered to be Non-Irritant (NI).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report date:
2010

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
other: EU Guideline Testing of Chemicals B46
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: OECD Draft Test Guideline
Deviations:
no
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Reference substance name:
Fatty acids C18-(unsaturated) lithium salts
EC Number:
700-426-3
Cas Number:
502962-81-4
Molecular formula:
Since this is a complex mixture of constituents no single formula can be assigned
IUPAC Name:
Fatty acids C18-(unsaturated) lithium salts
Details on test material:
Sponsor's identification : Fatty acids C18-(unsaturated) lithium salts
Description : off white solid
Batch number : SF 173
Date received : 21 January 2010
Expiry date : 21 January 2012
Storage conditions : room temperature in the dark

In vitro test system

Test system:
human skin model
Remarks:
reconstituted human epidermis model
Source species:
human
Cell type:
other: reconsituted human epidermal cells
Justification for test system used:
ECVAM recommended in vitro method. Also EU Guideline B46
Vehicle:
unchanged (no vehicle)
Duration of treatment / exposure:
15 MINUTES
Duration of post-treatment incubation (if applicable):
42 hrs
Number of replicates:
Three

Test system

Type of coverage:
other: Applied topically to reconstiyuted human epidermal cells
Vehicle:
unchanged (no vehicle)
Controls:
yes
yes, concurrent positive control
yes, concurrent negative control
no
Amount / concentration applied:
TEST MATERIAL

- The test Material was applied neat.

- Amount(s) applied (volume or weight with unit):
Approximately 10 mg of the test material was applied to the epidermis surface. The epidermis surface had previously been moistened with 5 µl of sterile distilled water to improve contact between the solid test material and the epidermis.

- Concentration (if solution):
The test material was used as supplied.

VEHICLE
No vehicle used
Duration of treatment / exposure:
15 Minutes & 42 hour post exposure incubation
Observation period:
Not applicable
Number of animals:
Not applicable
Details on study design:
TEST SITE
- Area of exposure:
Approximately 10 mg of the test material was applied to the epidermis surface. The epidermis surface had previously been moistened with 5 µl of sterile distilled water to improve contact between the solid test material and the epidermis.

- % coverage:
The test material was applied topically to the corresponding tissues ensuring uniform covering.

- Type of wrap if used:
None used

REMOVAL OF TEST SUBSTANCE
- Washing (if done):
At the end of the exposure period, each tissue was removed from the well using forceps and rinsed using a wash bottle containing PBS with Ca++ and Mg++. Rinsing was achieved by filling and emptying each tissue insert for approximately 40 seconds using a constant soft stream of PBS to gently remove any residual test material.

- Time after start of exposure:
15 Minutes post exposure

SCORING SYSTEM:
Quantitative MTT Assessment (percentage tissue viability)
For the test material the relative mean tissue viabilities obtained after the 15 minute treatment followed by the 42 hour post-exposure incubation period were compared to the mean of the negative control treated tissues (n=3). The relative mean viabilities were calculated in the following way:

mean OD540 of test material / mean OD540 of negative control x 100 = Relative mean tissue viability (percentage of negative control)

Classification of irritation potential is based upon relative tissue viability following the 15 minute exposure period followed by the 42 hour post-exposure incubation period according to the following:

Mean tissue viability is ≤50% : Irritant (I) R38

Mean tissue viability is >50% : Non-Irritant (NI)

Results and discussion

In vivo

Results
Irritation parameter:
other: Viability of cells
Basis:
mean
Remarks:
Viability of cells
Time point:
other: Day 6
Reversibility:
other: Not Applicable
Remarks on result:
other: See relative mean viability below.
Irritant / corrosive response data:
The relative mean viability of the test material treated tissues was 93.1% after a 15 minute exposure.
Other effects:
No

Any other information on results incl. tables

RESULTS

Direct MTT Reduction

An assessment found the test material was able to directly reduce MTT. Therefore, an additional procedure using water-killed tissues was performed during the determination of skin irritation potential. The test material was adequately rinsed from the tissues, however the results obtained showed that during rinsing no degree of interference due to direct reduction of MTT occurred. It was therefore considered unnecessary to use the results of the water-killed tissues for quantitative correction of results or for reporting purposes.

Test Material, Positive Control Material and Negative Control Material

The individual and mean OD540 values, standard deviations and tissue viabilities for the test material, negative control material and positive control material are given in Table 1. The mean viabilities and standard deviations of the test material and positive control, relative to the negative control are also given in Table 1.

The relative mean viability of the test material treated tissues was 93.1% after a 15 -minute exposure.

The qualitative evaluation of tissue viability is given in Table 2.

Following the 15-minute exposure the test material treated tissues appeared blue/white which was considered indicative of viable tissue.

Quality Criteria

The relative mean tissue viability for the positive control treated tissues was ≤40% relative to the negative control treated tissues and the standard deviation value of the percentage viability was ≤20%. The positive control acceptance criterion was therefore satisfied.

The mean OD540 for the negative control treated tissues was ≥0.6 and the SD value of the percentage viability was ≤20%. The negative control acceptance criterion was therefore satisfied.

Table 1 : Mean OD540 Values and Percentage Viabilities for the Negative Control Material, Positive Control Material and Test Material

Material

OD540of tissues

Mean OD540of triplicate tissues

±SDof OD540

Relative individual tissue viability (%)

Relative mean viability (%)

± SD of Relative mean viability (%)

Negative Control Material

1.016

0.962

0.002

105.6

100*

5.4

0.958

99.6

0.913

94.9

Positive Control Material

0.068

0.064

0.009

7.1

6.7

0.9

0.054

5.6

0.070

7.3

Test Material

0.877

0.896

0.021

91.2

93.1

2.2

0.919

95.5

0.892

92.7

SD=    Standard deviation

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


Table 2 : Qualitative Evaluation of Tissue Viability (MTT uptake visual evaluation)

Material

Tissue 1

Tissue 2

Tissue 3

Negative Control Material

-

-

-

Positive Control Material

++

++

++

Test Material

-

-

-

MTT visual scoring scheme
-          =         blue tissue (viable)
+         =         blue/white tissue (semi-viable)
++       =         tissue is completely white (dead

Applicant's summary and conclusion

Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: expert judgment
Conclusions:
The test material was considered to be Non-Irritant (NI)
Executive summary:
Introduction: The purpose of this test was to evaluate the skin irritation potential of the test material using the EPISKINTM reconstituted human epidermis model after a treatment period of 15 minutes followed by a post-exposure incubation period of 42 hours. The principle of the assay was based on the measurement of cytotoxicity in reconstituted human epidermal cultures following topical exposure to the test material by means of the colourimetric MTT reduction assay. Cell viability is measured by enzymatic reduction of the yellow MTT tetrazolium salt (3 -[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) to a blue formazan salt (within the mitochondria of viable cells) in the test material treated tissues relative to the negative controls. The concentration of the inflammatory mediator IL-1α in the culture medium retained following the 42 hour post-exposure incubation period is also determined for test materials which are found to be borderline non-irritant based upon the MTT reduction endpoint. This complimentary end-point will be used to either confirm a non-irritant result or will be used to override the non-irritant result.

Methods:

Triplicate tissues were treated with the test material for an exposure period of 15 minutes. At the end of the exposure period each tissue was rinsed before incubating for approximately 42 hours. At the end of the post-exposure incubation period each tissue was taken for MTT-loading. The maintenance medium from beneath each tissue was transferred to pre-labelled micro tubes and stored in a freezer for possible inflammatory mediator determination. 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).

Results: 

The relative mean viability of the test material treated tissues was 93.1% after a 15-minute exposure.

Quality criteria: 

The quality criteria required for acceptance of results in the test were satisfied.

Conclusion:  The test material was considered to be Non-Irritant (NI).