trisodium 2-(2-sulfoethoxy)ethane-1-sulfonate ethenesulfonate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vitro

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2021

Reliability:

1 (reliable without restriction)

Data source

Reference

Title:

Unnamed

Year:

2021

Report date:

2021

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 442E (In Vitro Skin Sensitisation assays addressing the key event on activation of dendritic cells on the Adverse Outcome Pathway for skin sensitisation)

GLP compliance:

yes

Type of study:

U937 cell line activation test (U-SENS™)

Test material

Test material information

Constituent 1

Reference substance name:

trisodium 2-(2-sulfonatoethoxy)ethane-1-sulfonate ethenesulfonate

EC Number:

700-978-5

Molecular formula:

C2H3O3S.Na C4H8O7S2Na2

IUPAC Name:

trisodium 2-(2-sulfonatoethoxy)ethane-1-sulfonate ethenesulfonate

Test material form:

liquid

Details on test material:

Reaction mass of disodium 2,2 oxydiethanesulfonate and sodium ethenesulfonate
Purity/Composition correction factor: The test item is a multi-constituent dissolved in water, dry residue is 33.65% , the correction factor is 2.971.

COA Lot nr 202108300033 of the aqueous solution.

Sodium ethylene sulphonate content cas 3039-83-6 :25.24 % Dry content :33.65 % pH@ 25°C :9.88 Colour :130 Apha

Composition of the dry product.

Constituent 1: Sodium ethylene sulphonate cas nr 3039-83-6 : 75.1 % Constituent 2: Isethionate bisether, disodium salt cas nr 63440-92-6: 16.52 % Impurity 1: Isethionate sodium cas nr1562-00-1 : 4.12 % Impurity 2: Sodium ethionate, disodium salt cas nr 1562-03-4 : 0.62 % Impurity 3: Sodium sulfate cas nr 7757-82-6 : 2.76 % Impurity 4: Sodium ethandisulfonate disodium salt cas nr 5325-43-9 : 0.64 %

Specific details on test material used for the study:

Reaction mass of disodium 2,2 oxydiethanesulfonate and sodium ethenesulfonate
Batch lot number:202108300033
Dry weight content : 33.65 %

In vitro test system

Vehicle / solvent control:

cell culture medium

Negative control:

DL-Lactic acid

Positive control:

other: 2,4,6 Trinitrobenzenesulfonic acid

Results and discussion

In vitro / in chemico

Results

Results 1

Group:

test chemical

Run / experiment:

run/experiment 2

Parameter:

EC150, CD86 [442E]

Cell viability:

The test item showed no cytotoxicity, the viability of the cells was higher than 70% at all test concentrations and therefore no CV70 values could be calculated and is considered to be higher than 200 µg/mL.

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: • No biologically relevant increase in expression levels of CD86 compared to the vehicle control was observed at any of the test concentrations after treatment with the test item. No EC150 could be calculated and is considered to be higher than 200 µg/mL.

Results 2

Key result

Group:

test chemical

Run / experiment:

run/experiment 1

Parameter:

EC150, CD86 [442E]

Cell viability:

The test item showed no cytotoxicity, the viability of the cells was higher than 70% at all test concentrations and therefore no CV70 values could be calculated and is considered to be higher than 200 µg/mL.

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: No EC150 could be calculated and is considered to be higher than 200 µg/mL.

Outcome of the prediction model:

negative [in vitro/in chemico]

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In conclusion, Reaction mass of disodium 2,2’-oxydiethanesulfonate and sodium ethenesulfonate is classified as negative (no biologically relevant increase in the expression levels of CD86 cell surface marker in the U937 cell line) under the experimental conditions described in this report.

Executive summary:

The objective of this study is to evaluate the ability of the test item to increase the expression levels of CD86 cell surface marker in the U937 cell line activation Test (U-Sens™) assay.

The test item was evaluated for the ability to increase the expression levels of CD86 cell surface marker. An overview of the viability and CD86 cell surface marker activity is summarized in Table 1. The results of the positive, negative and vehicle controls are summarized in Table 2. An overview of EC₁₅₀ and CV₇₀ values is given in Table 3. The individual raw data are presented in Table 4 and Table 5.

Two independent experiments were performed. The cell viability before incubation with the test item was > 90% (98% and 97% in experiment 1 and 2, respectively). The cells were in these experiments incubated with the test item in a concentration range of 1.0 – 200 µg/mL. The increase of CD86 cell surface marker expression was assessed by measuring the amount fluorescent cell staining of the CD86 cell surface marker compared to the vehicle control. In addition, the viability was assessed with propidium iodide.

Experiment 1

• No precipitation was observed at the end of the incubation period in the 96-well plates.


• The test item showed no toxicity, the viability of the cells was higher than 70% at all test concentrations and therefore no CV₇₀ values could be calculated and is considered to be higher than 200 µg/mL.


• No increase in expression levels of CD86 compared to the vehicle control was observed at any of the test concentrations after treatment with the test item. No EC₁₅₀ could be calculated and is considered to be higher than 200 µg/mL.


• The test item showed no colour interference.


• The positive control (TNBS) showed a S.I. ≥ 1033% in all wells and was non-cytotoxic at all concentrations (cell viability ≥ 70%). The negative control (LA) showed a  I. ≤ 88% in all wells and was non-cytotoxic at all concentrations (cell viability ≥ 70%).

Experiment 2

• No precipitation was observed at the end of the incubation period in the 96-well plates.


• The test item showed no toxicity, the viability of the cells was higher than 70% at all test concentrations and therefore no CV₇₀ values could be calculated and is considered to be higher than 200 µg/mL.


• No biologically relevant increase in expression levels of CD86 compared to the vehicle control was observed at any of the test concentrations after treatment with the test item. No EC₁₅₀ could be calculated and is considered to be higher than 200 µg/mL. At the dose level of 100µg/mL, the test item showed a CD86 induction of 153%. However, as this is only just above the threshold of 150%, and only occurred at this dose level and not at the highest dose level, this minor increase is not considered to biologically relevant.


• The test item showed no colour interference.


• The positive control (TNBS) showed a S.I. ≥ 634% in all wells and was non-cytotoxic at all concentrations (cell viability ≥ 70%). The negative control (LA) showed a I. ≤ 148% in all wells and was non-cytotoxic at all concentrations (cell viability            ≥ 70%).

 

 

Both tests passed the acceptance criteria:

• At the end of the incubation treatment period, the mean viability of the triplicate untreated U937 cells was above the threshold of 90% (99% in experiment 1 and 2).


• The CD86 basal expression of untreated U937 cells is within the range of ≥ 2% and
  ≤ 25% in both experiments.


• At least two out of three IgG1 values of untreated U937 cells fell within the range of
  ≥ 0.6% and < 1.5% in both experiments.


• No drift in CD86 expression was observed in the untreated controls (RPMI) and negative (LA) controls.

In both experiments the positive and negative control were considered valid, and the positive control fell within the historical control data. Overall, it is concluded that the test conditions were adequate and that the test system functioned properly.