Sodium hydrogen-5-sulphoisophthalate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

22 July 2010

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

24 August 2009

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Sodium hydrogen-5-sulphoisophthalate

EC Number:

228-845-2

EC Name:

Sodium hydrogen-5-sulphoisophthalate

Cas Number:

6362-79-4

Molecular formula:

C8H6O7S.Na

IUPAC Name:

Sodium 3,5-dicarboxybenzenesulfonate

Constituent 2

Reference substance name:

288-845-2

IUPAC Name:

288-845-2

Constituent 3

Reference substance name:

Sodium-3,5-dicarboxybenzenesulfonate

IUPAC Name:

Sodium-3,5-dicarboxybenzenesulfonate

Constituent 4

Reference substance name:

5-(Sodiosulfo)isophthalic acid

IUPAC Name:

5-(Sodiosulfo)isophthalic acid

Test material form:

solid: particulate/powder

Details on test material:

- Name of test material (as cited in study report): 5-(sodiosulfo)isophthalic acid
- Physical state: white powder
- Analytical purity: 99.4%
- Lot/batch No.: 6F21174000
- Expiration date of the lot/batch: 2 December 2013
- Stability under test conditions: stable
- Storage condition of test material: At room temperature in the dark. The material is hygroscopic and must be stored in a well-sealed container.

Specific details on test material used for the study:

Test item is hygroscopic and was therefore stored in well-sealed container.
Test item is stability at higher temperatures (maximum temperature: >500°C).
No correction was made for the purity/composition of the test compound.

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

skin obtained from plastic surgery from multiple donors

Justification for test system used:

The test is based on the experience that irritant chemicals show cytotoxic effects following short term exposure to the stratum corneum of the epidermis. The test is designed to predict and classify the skin irritation potential of a test substance by assessment of its effect on a three dimensional human epidermis model. The study procedures were based on a validated method according to OECD/EC guidelines.

Vehicle:

unchanged (no vehicle)

Details on test system:

The test system consisted of EPISKIN Small Model(TM) (EPISKIN-SM(TM), 0.38 cm^2, Batch no.: 12-EKIN-032).
Source: SkinEthic Laboratories, Lyon, France.

The test was performed on a total of 3 tissues per test substance together with negative and positive controls. The skin was moistened with 5 μl Milli-Q water to ensure close contact of the test substance to the tissue and the solid test substance (11.6 to 12.2 mg; with a small glass weight boat) was added into 12-well plates on top of the skin tissues. Three tissues were treated with 25 μl PBS (negative control) and 3 tissues with 25 μl 5% SDS (positive control), respectively. The positive control was re-spread after 7 minutes contact time. After the exposure period of 15 minutes at room temperature, the tissues were washed with PBS to remove residual test substance. After rinsing the cell culture inserts were each dried carefully and moved to a new well on 2 ml pre-warmed maintenance medium until all tissues were dosed and rinsed. Subsequently the skin tissues were incubated for 42 hours at 37°C.

After incubation, cell culture inserts were dried carefully to remove excess medium and were
transferred into a 12-wells plate prefilled with 2 ml MTT-medium (0.3 mg/ml). The tissues were
incubated for 3 h at 37°C. After incubation the tissues were placed on blotting paper to dry the tissues. Total biopsy was made by using a biopsy punch. Epidermis was separated from the collagen matrix and both parts were placed in prelabeled microtubes and extracted with 500 μl isopropanol. Tubes were stored refrigerated and protected from light for 70 hours. The amount of extracted formazan was determined spectrophotometrically at 570 nm in duplicate with the
Multiskan spectrum.
Cell viability was calculated for each tissue as a percentage of the mean of the negative control
tissues. Skin irritation potential of the test substance was classified according to remaining cell viability following exposure of the test substance.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

Negative control:
25 μl Phosphate buffered saline (PBS).

Positive control:
25 μl 5% (aq) Sodium dodecyl sulphate (SDS) in PBS.

Duration of treatment / exposure:

15 minutes

Duration of post-treatment incubation (if applicable):

42 hours

Number of replicates:

3

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Value:

96

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Any other information on results incl. tables

5-(Sodiosulfo)isophthalic Acid was checked for possible direct MTT reduction by adding the test substance to MTT medium. Because no colour change was observed it was concluded that 5 - (Sodiosulfo)isophthalic Acid did not interact with MTT.

The positive control had a mean cell viability after 15 minutes exposure of 46% and did not meet the acceptability criteria. Approximately 1 hour after the performance of this test another in vitro skin irritation test was performed using the same batch of skin and the same batch of 5% (aq) SDS. The positive control had a mean cell viability of 5% and met the acceptability criteria. The absolute mean OD570 of the negative control tissues (in both in vitro skin irritation tests) were within the laboratory historical control data range (See APPENDIX 3). The standard deviation value of the percentage viability of three tissues treated identically was less than 11%, indicating that the test system functioned properly. It can be concluded that the deviation in the mean cell viability of the positive control in the first in vitro skin irritation test was caused by a technical error.

Mean tissue viability in the in vitro skin irritation test with 5 -(Sodiosulfo)isophthalic Acid (see table):

	Mean tissue viability(percentage of control)
Negative control	100
test substance	96
Positive control (note 1)	5

Note 1: Based on second in vitro skin irritation test

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Based on the results of an in vitro skin irritation test with 5-(Sodiosulfo)isophthalic Acid using a human skin model it is concluded that this test is valid and that 5-(Sodiosulfo)isophthalic Acid is non-irritant in the in vitro skin irritation test under the experimental conditions described in this report.