Sodium dimethyl 5-sulphonatoisophthalate

Endpoint summary

Description of key information

Sodium dimethyl 5-sulphonatoisophthalate was not irritating to skin using the
EPISKIN model and did not show any eye irritating effects relevant for C&L in
the BCOP assay and in vivo in the rabbit eye.  

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012-06-06 to 2012-06-08

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP and guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

other: EpiSkin™ Small Model

Type of coverage:

other: not applicable

Preparation of test site:

other: not applicable

Vehicle:

unchanged (no vehicle)

Amount / concentration applied:

The epidermal surface was first moistened with 10 μL deionised water (in order to improve further contact between powder and epidermis) and then 10 mg of the test item was applied evenly onto the skin.

Duration of treatment / exposure:

15 minutes at room temperature

Details on study design:

PERFORMANCE OF THE STUDY
Pre-incubation (day [-1])
The medium was pre-warmed to 37 °C. The appropriate number of assay plate wells were filled with the pre-warmed medium (2 mL per well). The epidermis units were placed, with the media below them in contact with the epidermis, into each prepared well and then incubated overnight at 37 °C in an incubator with 5 % CO2.

APPLICATION (DAY 0)
Test Item
The epidermal surface was first moistened with 10 μL deionised water* (in order to improve further contact between powder and epidermis) and then 10 mg of the test item was applied evenly onto the skin. The test item was spread gently with a curved flat spatula in order to cover evenly all the skin surface if necessary.

Positive and negative control
A volume of 20 μL positive control (SDS 5 % aq.) or negative control (1 x PBS) was applied to the skin surface by using a suitable pipette. Chemicals were spread gently with the pipette tip in order to cover evenly all the epidermal surface if necessary.

EXPOSURE (DAY 0)
The plates with the treated epidermis units were incubated for the exposure time of 15 minutes at room temperature.

RINSING (DAY 0)
After the incubation time the EPISKIN-SM units were removed and rinsed thoroughly with PBS 1 x solution to remove all of the test material from the epidermal surface. The rest of the PBS was removed from the epidermal surface with a suitable pipette tip linked to a vacuum source (care was taken to avoid damaging the epidermis).

POST-INCUBATION (DAY 0-2)
After rinsing the units were placed into the plate wells filled with fresh pre-warmed “Maintenance Medium” (2 mL/well) below them and then incubated for 42 hours at 37 °C in an incubator with 5 % CO2.

MTT TEST (DAY 2)
After the 42 hours incubation the EPISKIN-SM units were transferred into the MTT solution filled wells (2 mL of 0.3 mg/mL MTT per well) and then incubated for 3 hours at 37 °C in an incubator with 5 % CO2 protected from light.

FORMAZAN EXTRATION (DAY 2)
At the end of incubation with MTT a formazan extraction step was undertaken:
A defined disk of epidermis from each replicate was cut from the unit (this involves the maximum area of the disk) using a biopsy punch (supplied as part of the kit). The epidermis was separated with the aid of forceps and both parts (epidermis and collagen matrix) were placed into a tube of 500 μL acidified isopropanol (one tube corresponding to one well of the tissue culture plate).
The capped tubes were thoroughly mixed by using a Vortex Mixer to achieve a good contact of all of the material to the acidified isopropanol then incubated for about two hours at room temperature protected from light with gentle agitation (~150 rpm) for formazan extraction.

CELL VIABILITY (DAY 2)
Following the formazan extraction, 2 × 200 μL samples from each tube were placed into the wells of a 96-well plate (labelled appropriately). The OD’s of each well were recorded using a 96-well plate spectrophotometer (Thermo Scientific; Multiscan FC) at a wavelength of 570 nm while using an acidified isopropanol solution blank (6 × 200 μL).

VALIDITY OF THE TEST
– The mean OD value of the three negative control tissues should be between 0.6 and 1.5 and the standard deviation value (SD) of the % viability should be ≤ 18.
– The acceptable mean percentage viability range for positive controls is 0-40 % and the standard deviation value (SD) of the % viability should be ≤ 18.
– For test chemicals, the standard deviation value (SD) of the % viability should be ≤ 18.

CRITERIA FOR IN VITRO INTERPRETATION
Mean tissue viability % is ≤ 50 % = Irritant
Mean tissue viability % is > 50 % = Non Irritant

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Duration of exposure: 15 min.

Value:

96

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

The results of the optical density measured at 570 nm and the calculated % viability of the cells are presented below:

 

Substance	Mean Optical Density	Mean Viability (%)
Negative Control	0.816	100 (standard deviation: 2.18)
Positive Control	0.038	5 (standard deviation: 1.18)
Test Item	0.780	96 (standard deviation: 1.06)

 

Summary

All obtained test item viability results were above 50 % when compared to the viability values obtained from the negative control, therefore the test item was considered to be non-irritant to skin.

Interpretation of results:

GHS criteria not met

Conclusions:

The test item revealed no skin irritation in vitro (EPISKIN model).

Executive summary:

In a skin irritation study ( Toxi-Coop Zrt., 2012,EPISKIN model, OECD 439, GLP compliance) disks of epidermal units (three units / chemical) were treated with the test item Sodium dimethyl 5-sulphonatoisophthalate (SIM-Ester)and incubated for 15 minutes at room temperature. Exposure of the test material was terminated by rinsing the epidermal units with PBS 1x solution. Epidermis units were then incubated at 37 °C for 42 hours in an incubator with 5 % CO2. The viability of each disk was assessed by incubating the tissues for 3 hours with MTT solution at 37 °C in 5 % CO2 and protected from light. The resulting formazan chrystals were extracted with acidified isopropanol and quantified with the optical densities (OD) recorded spectrophotometrically. SDS 5 % aq. and 1 x PBS treated epidermis units were used as positive and negative controls, respectively. For each treated tissue viability was expressed as a percentage relative to negative control.

Positive and negative controls showed the expected cell viability values within acceptable limits. The experiment was considered to be valid. SIM-Ester did not show significantly reduced cell viability in comparison to the negative control. All obtained test item viability results were above 50 % when compared to the viability values obtained from the negative control, therefore the test item was considered to be non-irritant to skin using the in vitro EPISKIN model.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012-05-29 to 2012-06-05

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Justification for type of information:

In vivo eye irritation study was initiated before the entry into force of the amendments to Annexes VII and VIII

Qualifier:

according to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2400 (Acute Eye Irritation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: S & K LAP Kft. 2173 Kartal, Császár út 135, Hungary
- Age at study initiation: Young adult rabbits
- Weight at study initiation: 2838-3213 g
- Housing: Animals were housed individually in metal cages
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 12 days

ENVIRONMENTAL CONDITIONS
- Temperature: 20 ± 3 °C
- Humidity: 30 - 70 %
- Air changes: 9-12 changes/hour
- Photoperiod (hrs dark / hrs light): 12 hours artificial light, from 6 a.m. to 6 p.m.

Vehicle:

unchanged (no vehicle)

Remarks:

The test item was ground to a fine dust prior to application

Controls:

other: The contralateral eye served as control

Amount / concentration applied:

A weight of 0.1 g of the undiluted ground test item Sodium dimethyl 5-sulphonatoisophthalate (SIM-Ester) was used for the study in pure state, in a single dose.

Duration of treatment / exposure:

Continuous (the eyes of the test animals were not washed out after the application of test item).

Observation period (in vivo):

The eyes were examined at 1, 24, 48 and 72 hour(s) and then at 1 week after the application.

Number of animals or in vitro replicates:

3 male rabbits

Details on study design:

SCORING SYSTEM: according to the scoring system by Draize (1979) and OECD 405 (24 April 2002)

BODY WEIGHT: The body weights were recorded at the beginning and at the end of the experiment.

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: no effect

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: no effect

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: no effect

Irritation parameter:

iris score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: no effect

Irritation parameter:

iris score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: no effect

Irritation parameter:

iris score

Basis:

animal #3

Time point:

24/48/72 h

Score:

0

Max. score:

2

Reversibility:

other: no effect

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0

Max. score:

3

Reversibility:

fully reversible within: 1 week

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

3

Reversibility:

other: no effect

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

24/48/72 h

Score:

1

Max. score:

3

Reversibility:

fully reversible within: 1 week

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 1 week

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

fully reversible within: 1 week

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

24/48/72 h

Score:

2

Max. score:

4

Reversibility:

fully reversible within: 1 week

Irritant / corrosive response data:

- The animals individual mean score (considering reading at 24,48 and 72 hours after the treatment) for discharge was: 0.33, 0.00 and 2.67. However, conjunctival discharge does not represent a relevant endpoint with respect to classification for acute eye irritation.

- During the study the control eyes of the animals were symptom-free.

Other effects:

General state and the behaviour of the animals were normal throughout the study period. There were no effects on body weight attributable to the treatment with the test item during the contact and observation period.

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The test item, caused slight to moderate conjunctival irritant effects, fully reversible within 1 week and not relevant for C&L. There were no test item-related effects on cornea and iris.

Executive summary:

In an acute eye irritation study (Toxicoop Zrt., 2012, OECD 405, GLP compliance) three rabbits were exposed with the test item Sodium dimethyl 5-sulphonatoisophthalate (SIM-Ester). The test item was placed into the conjunctival sac of the left eye of each animal. The untreated right eye served as control. A weight of 0.1 g of the ground test item was used in pure state, as a single dose. The eyes of the test animals were not washed out after the application of test item. The eyes were examined at 1, 24, 48 and 72 hour(s) and then at 1 week after the application. One hour after the treatment, slight conjunctival redness and chemosis, as well as slight to moderate discharge were observed. 24 hours after the treatment, slight redness, slight to moderate chemosis, as well as slight and severe discharge were detected in two animals. 1 week after treatment the study was terminated, as all animals were free of symptoms of irritation. There were no test item-related effects on cornea and iris. In conclusion the test item SIM-Ester did not show any irritating effects relevant for C&L.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin

In a skin irritation study ( Toxi-Coop Zrt. (c), 2012,EPISKIN model, OECD 439, GLP compliance) disks of epidermal units (three units / chemical) were treated with the test item Sodium dimethyl 5-sulphonatoisophthalate (SIM-Ester) and incubated for 15 minutes at room temperature. Exposure of the test material was terminated by rinsing the epidermal units with PBS 1x solution. Epidermis units were then incubated at 37 °C for 42 hours in an incubator with 5 % CO2. The viability of each disk was assessed by incubating the tissues for 3 hours with MTT solution at 37 °C in 5 % CO2 and protected from light. The resulting formazan chrystals were extracted with acidified isopropanol and quantified with the optical densities (OD) recorded spectrophotometrically. SDS 5 % aq. and 1 x PBS treated epidermis units were used as positive and negative controls, respectively. For each treated tissue viability was expressed as a percentage relative to negative control. Positive and negative controls showed the expected cell viability values within acceptable limits. The experiment was considered to be valid. SIM-Ester did not show significantly reduced cell viability in comparison to the negative control. All obtained test item viability results were above 50 % when compared to the viability values obtained from the negative control, therefore the test item was considered to be non-irritant to skin using the in vitro EPISKIN model.

Eye

In an acute eye irritation study (Toxicoop Zrt. (d), 2012, OECD 405, GLP compliance) three rabbits were exposed with the test item Sodium dimethyl 5-sulphonatoisophthalate (SIM-Ester). 0.1 g of the test item was placed into the conjunctival sac of the left eye of each animal. The untreated right eye served as control. The eyes of the test animals were not washed out after the application of test item. The eyes were examined at 1, 24, 48 and 72 hour(s) and then at 1 week after application. One hour after treatment, slight conjunctival redness and chemosis, as well as slight to moderate discharge were observed. 24 hours after treatment, slight redness, slight to moderate chemosis, as well as slight and severe discharge were detected in two animals. 1 week after treatment the study was terminated, as all animals were free of symptoms of irritation. There were no test item-related effects on cornea and iris. In conclusion the test item SIM-Ester did not show any irritating effects relevant for C&L.

In an in vitro eye irritation study (Harlan Cytotest Cell Research GmbH (a), 2012, OECD 437, GLP compliance) the corneal irritation and damage potential of Sodium dimethyl 5-sulphonatoisophthalate (SIM-Ester) were assessed by means of the BCOP assay using fresh bovine corneae. After a first opacity measurement of the fresh bovine corneae (t0), the 20% (w/v) suspension in saline (0.9% (w/v) NaCl in deionised water) of the test item, the positive, and the negative controls were applied to corneae and incubated for 240 minutes at 32 ± 1 °C. The posterior chamber contained MEM medium supplemented with sodium bicarbonate and L-glutamine and 1% fetal calf serum (FCS) (complete medium = cMEM). After the incubation phase the test item, the positive, and the negative controls were each rinsed from the corneae and opacity was measured again (t240). After the opacity measurements permeability of the corneae was determined by measuring spectrophotometrically the transfer of sodium fluorescein after incubation in a horizontal position for 90 minutes at 32 ± 1 °C. With the negative control (saline) neither an increase of opacity nor permeability of the corneae could be observed. The positive control (10% (w/v) Benzalkonium chloride in saline) showed clear opacity effects corresponding to a classification as corrosive / severe irritant to the eye. Relative to the negative control, the test item SIM-Ester did not cause any increase of the corneal opacity or permeability. The calculated mean in vitro irritation score was 0.00 and thus the test item was classified as non corrosive / non severe irritant to the eye.

Respiratory tract

There is no study available. However, since the substance is neither a skin nor an eye irritant, an irritating effect on the respiratory tract is unlikely.

Justification for selection of skin irritation / corrosion endpoint:
well documented and reliable study.

Justification for selection of eye irritation endpoint:
well documented and reliable study.

Justification for classification or non-classification

Based on the results obtained in the skin and eye irritation studies, Sodium dimethyl 5-sulphonatoisophthalate

is not subject to classification for skin and eye irritation according to Directive 67/548/EEC and according to Regulation (EC) No 1272/2008. Furthermore, the available data on the irritation potential indicate that the substance is not subject to classification and labelling for respiratory tract irritation.