Sodium 3-(allyloxy)-2-hydroxypropanesulphonate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation / corrosion

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

23 April - 13. July 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

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:

EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Sodium 3-(allyloxy)-2-hydroxypropanesulphonate

EC Number:

258-004-5

EC Name:

Sodium 3-(allyloxy)-2-hydroxypropanesulphonate

Cas Number:

52556-42-0

Molecular formula:

C6H12O5S.Na

IUPAC Name:

sodium 2-hydroxy-3-(prop-2-en-1-yloxy)propane-1-sulfonate

additive 1

Reference substance name:

Sodium hydroxide

EC Number:

215-185-5

EC Name:

Sodium hydroxide

Cas Number:

1310-73-2

Molecular formula:

HNaO

IUPAC Name:

sodium hydroxide

impurity 1

Reference substance name:

3-(allyloxy)propane-1,2-diol

EC Number:

204-620-4

EC Name:

3-(allyloxy)propane-1,2-diol

Cas Number:

123-34-2

Molecular formula:

C6H12O3

IUPAC Name:

3-(allyloxy)propane-1,2-diol

impurity 2

Reference substance name:

disodium 2-hydroxy-3-(3-sulfonatopropoxy)propane-1-sulfonate

Molecular formula:

C6H12Na2O8S2

IUPAC Name:

disodium 2-hydroxy-3-(3-sulfonatopropoxy)propane-1-sulfonate

Test material form:

solid: particulate/powder

Details on test material:

light yellow powder

In vitro test system

Test system:

human skin model

Source species:

human

Vehicle:

water

Control samples:

yes, concurrent negative control

yes, concurrent vehicle

yes, concurrent positive control

Duration of treatment / exposure:

3 minutes

Number of replicates:

2

Test system

Amount / concentration applied:

25 mg in 25 µl water

Duration of treatment / exposure:

3 min, 60 min

Details on study design:

Preparations:
On the day of the start of the experiment, the MTT concentrate was thawed. The concentrate was diluted with the MTT solvent and the solution was stored at 4°C in the dark. The tissue plate was brought out of the fridge one hour before the treatment. The assay medium was wamed in the water bath to 37 °C.
Description ot the method
Four 6-well-plates were prepared with 0.9 l assay medium in each well. The inserts containing the tissues were transferred to the wells using sterile forceps and the 6-well-plates were set into the incubator at 37 °C and 5 % CO2 for one hour (pre-incubation).
For each experiment ("three minutes" and "one hour"), two 6-well-plates were used. After preincubation, the assay medium was replaced by fresh assay medium and the test was started, using two wells as negative control with 50 µl H2O demin., two wells as positive controls with 50 µL potassium hydroxide solution and two other wells for testing the test item. The solid test item was ground, and 25 mg test item were applied together with 25 µL H2O. At the start of each experiment (application of negative controls), a stop watch was started.
After the respective incubation time (three minutes +- 10 sec and one hour), the inserts were removed from the plates using sterile forceps. The inserts were thoroughly rinsed with PBS, blotted with sterile cellulose tissue and set into the respective holding plate, using the wells containing assay medium. After transfer of all inserts, they were immediately moved to the wells containing MTT reagen, blotting the bottom with cellulose tissue again before setting the insert into the MTT well. The tissues were incubated with MTT reagent for three hours. After this time, the MTT reagent was aspirated and replaced by PBS buffer. This was then aspirated, too, and replaced several times. At last, each insert was thoroughly dried and set into the empty, pre-warmed 24-well-plate. Into each well, 2 mL isopropanole werepipetted, taking careto reach the upper rim of the insert. The plate was then covered with Parafilm and left to stand over night at room temperature.
On the next day, the inserts in which formazan had been produced over night were pierced with an injection needle, taking care that all colour was extracted. the inserts were then discarded and the content of each well was thoroughly mixed in order to achieve homogenisation.
From each well, three replicates with 200 µL solution (each) were pipetted into a 96-wll-plate which was read in a plate spectral photometer at 570 nm.

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Value:

81.9

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

Measured Values
see table in "any other information"

Comparison of Formazan Production
3 min Incubation time: 81.9 % (test item); 20.2 % (Positive control)
60 min Incubation time: 42.6 % (test item); 10.2 % (Positive control)

Assessment:
The relative absorbance values were reduced to 81.9 % after three minutes treatment. This value is above the threshold for corrosivity (50 %). After one hour treatment, the relative absorbance values were reduced to 42.6 %, lying above the threshold for corrosivity (15%).
Therefore, the test item is considered as not corrosive.

Any other information on results incl. tables

Measured Values

The absorption values of negative control, test item and positive control are given in the following table:

Absorption Values

Negative Control		Test Item		Positive Control		Incubation
Tissue 1	Tissue 2	Tissue 1	Tissue 2	Tissue 1	Tissue 2
2.465	2.253	1.938	1.954	0.552	0.383	3 min
2.388	2.351	1.977	1.865	0.540	0.422
2.407	2.267	1.927	1.912	0.541	0.419
2.199	2.200	0.941	0.930	0.224	0.231	1 hour
2.222	2.217	0.950	0.926	0.219	0.243
2.125	2.226	0.939	0.929	0.219	0.213
Mean		Mean		Mean
2.355		1.929		0.476		3 min
2.198		0.936		0.225		1 hour

 

Comparison of Formazan Production

For the test item and the positive control, the following percentage values of mean formazan production were calculated in comparison to the mean of the negative controls:

  % Formazan Production

Test Item	Positive Control	Incubation
81.9%	20.2%	3 min
42.6%	10.2%	1 hour

 

Corrosivity of the Test Item

The relative absorbance values were reduced to 81.9% after three minutes treatment. This value is above the threshold for corrosivity (50%). After one hour treatment, the relative absorbance values were reduced to 42.6 %, lying above the threshold for corrosivity (15%). Therefore, the test item is considered as not corrosive.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

The study is well performed and gives following results for the Endpoint:
not corrosive

Executive summary:

Two tissues of the human skin model EpiDerm^TMwere treated with sodium 3-(allyloxy)-2-hydroxypropanesulphonate (HAPS) for three minutes and one hour, respectively. In average, 25.2 mg of the solid test item were applied to each tissue and spread to match the tissue size.

Deionised water was used as negative control, 8m KOH was used as positive control.

After treatment, the respective substance was rinsed from the tissue; then, cell viability of the tissues was evaluated by addition of MTT which can be reduced to a blue formazan. Formazan production was measured by measuring the optical density (OD) of the resulting solution.

After treatment with the negative control, the absorbance values were well above the required acceptability criterion of mean OD > 0.8 for both treatment intervals thus showing the quality of the tissues. The positive control showed clear corrosive effects for both treatment intervals.

After three minutes treatment with the test item, the relative absorbance values were reduced to 81.9 %. This value is well above the threshold for corrosion potential (50%). After one hour treatment, relative absorbance values were reduced to 42.6 %. This value, too, is well above the threshold for corrosion potential (15%). In the guideline, values greater or equal to the threshold are considered as “non-corrosive to skin”.

Therefore, sodium 3-(allyloxy)-2-hydroxypropanesulphonate (HAPS) is considered as

not corrosive in the Human Skin Model Test.