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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin irritation / corrosion
Remarks:
other: In Vitro (Reconstructed Human Epidermis)
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study was conducted by a GLP accredited laboratory using OECD Testing Guideline 439. The study was conducted on the substance 6-[(p-tosyl)amino]hexanoic acid, compound with 2,2’,2’’- nitrilotriethanol (1:1) (tosyl salt) which is structurally similar to the registered substance.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013
Report Date:
2013

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD 439 In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method
Deviations:
no
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: Waxy solid
Details on test material:
Identity: Triethanolamine salt
Alternative Name: 6-[(p-tosyl)amino]hexanoic acid, compound with 2,2',2''-nitrilotriethanol (1:1)
CAS Number: 93981-14-7
Intended Use: Offshore drilling (anti-corrosion agent)
Appearance: Brown-orange waxy solid
Storage Conditions: Room temperature (ca. 20°C)

Test animals

Species:
other: Reconstructed human epidermis
Strain:
other: Reconstructed human epidermis
Details on test animals and environmental conditions:
Receipt of Tissues

On receipt, the kit contents were checked and the inserts with tissues on agarose were stored at room temperature until use. The kit was used within the expiry date indicated by the supplier (expiry date: 17th June 2013). The maintenance medium was pre-warmed to 37°C. The tissues were removed from the agar and placed into wells of 12 well plates containing 2 ml pre-warmed maintenance medium per well. The tissues were incubated for a minimum of 24 hours at 37°C ± 2°C in a humidified atmosphere of 5% CO2 in air.

Test system

Type of coverage:
other: The test substance was ground using a pestle and mortar and dispensed over each tissue using glass weighing boats.
Preparation of test site:
not specified
Vehicle:
unchanged (no vehicle)
Controls:
yes
Amount / concentration applied:
The test substance was ground using a pestle and mortar. A weight of 10 mg ± 2 mg was dispensed over each tissue using glass weighing boats.
Duration of treatment / exposure:
15 minutes (triplicate tissues)
Observation period:
After exposure the tissues were incubated for 42 hours. After 42 hours the tissues were transferred to a well containing MTT and incubated for 3 hours.
Number of animals:
Not applicable
Details on study design:
The negative control was sterile Dulbecco's Phosphate Buffered Saline (DPBS) with magnesium and calcium. The positive control was 5% Sodium Dodecyl Sulphate (SDS) in purified water. The controls and their results were shared with another study performed in the same assay.

The test involves the application of the test substance for 15 minutes to the EPISKIN™ three-dimensional human skin model. The model consists of normal, human-derived epidermal keratinocytes which have been seeded on a dermal substitute consisting of a collagen type 1 matrix coated with type IV collagen. After 13 days in culture a multilayered, highly differentiated model of the human epidermis with a functional multi-layered stratum corneum has formed. The epidermis surface area supplied is 0.38 cm2. The EPISKIN™ kits include assay medium, maintenance medium, 12 well plates and the tissues which are shipped on nutritive agar.

The principle of the assay is that irritant substances are sufficiently cytotoxic to cause cell death in the cell layers. The cell viability was determined by mitochondrial dehydrogenase activity, assessed by the reduction of MTT (3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide) to a soluble, coloured, formazan product. The prediction model (OECD 439) uses the percentage viability values (compared to negative control viability) to identify irritant and non-irritant substances. The test includes acceptance criteria for both negative and positive controls.

Results and discussion

In vivo

Results
Irritation parameter:
other: Tissue viability
Basis:
mean
Score:
97.8
Max. score:
113.5
Reversibility:
no data
Remarks on result:
other: Mean of three replicate tissues
Other effects:
Negative control: The mean absorbance of the triplicate negative control values was 0.862 which was between the minimum and maximum values of 0.6 and 1.5. The standard deviation (SD) of the % viability was 9.8 which was below the maximum value of 18.
Positive control: The percentage mean viability of the positive control was 25.7 ± 2.4 of the negative control. These were below the maximum acceptance values of 40% viability and SD of 18.

Any other information on results incl. tables

The mean Optical Density (OD) for the 6 replicate blanks was subtracted from the individual test substance and control tissues OD.

The viability of each tissue was expressed as a percentage of the mean negative control value.

Episkin Results

Sample

Tissue Replicate

Optical Density (OD)

OD – Blank

% Negative Control

Negative Control

a

1.033

0.994

0.891

0.852

101.1

b

0.916

0.913

0.775

0.771

89.7

c

1.117

1.049

0.975

0.907

109.2

Replicates a, b, c

Average sd

0.862

0.080

100.0

9.8

Positive Control

a

0.388

0.371

0.246

0.230

27.6

b

0.344

0.335

0.202

0.193

22.9

c

0.379

0.361

0.238

0.219

26.5

Replicates a, b, c

Average sd

0.221

0.021

25.7

2.4

Test Substance

a

1.139

1.100

0.998

0.958

113.5

b

0.944

0.907

0.803

0.765

91.0

c

0.931

0.886

0.789

0.744

88.9

Replicates a, b, c

Average sd

0.843

0.107

97.8

13.6

Blank

 

0.135

0.145

0.144

0.140

0.143

0.143

 

 

Average sd

0.142

0.004

 

sd: Standard Deviation

Note: Rounded values only are displayed; unrounded numbers are used for calculations by Excel spreadsheet

Applicant's summary and conclusion

Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
It was concluded that the test substance had a mean tissue viability of 97.8% ± 13.6% and was predicted as non-irritant to the skin. The test substance is not classified as a skin irritant, in accordance with EU CLP criteria.
Executive summary:

The skin irritation potential of the test substance was determined in accordance with the OECD Guideline for Testing of Chemicals 439. It was concluded that the test substance had a mean tissue viability of 97.8% ± 13.6% and was predicted as non-irritant to the skin. The test substance is not classified as a skin irritant, in accordance with EU CLP criteria. Read-across between the registered substance and the test substance (tosyl salt) is considered justified owing to the structural similarity of the two substances. Both are salts of a carboxylic acid and triethanolamine. The acid components are structural isomers and differ only in the position of a single methyl group in the N-methyl salt. This is not expected to change the behavior or properties of the substances, and both substances have shown similarity in the results of other physical endpoints such as water solubility.