Formaldehyde, reaction products with m-phenylenediamine, sodium sulfide (Na2S) and sulfur

Endpoint summary

Description of key information

Eye: non-irritating according to OECD 437;

         slighty irritating according to OECD 405

 

Skin: non-irritating according to OECD 439

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:

05-Jun-2012 to 11 June 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Amount / concentration applied:

TEST MATERIAL
- Amount applied: C.I. Leuco Sulphur Brown 37 was applied and incubated with a cotton swab directly on top of the skin tissue

NEGATIVE CONTOL:
- Amount applied: 25 µL Phosphate buffered saline

POSITIVE CONTROL
- Amount applied: 25 µL
- Concentration: 5% (aq) Sodium dodecyl sulphate

Duration of treatment / exposure:

Exposure: 15 minutes
Post incubation period: 42 hours

Details on study design:

TEST SITE
- Area of exposure: human epidermis model
- % coverage: 0.38 cm2

REMOVAL OF TEST SUBSTANCE
- Washing: phosphate buffered saline
- Time after start of exposure: 15 minutes

POST INCUBATION PERIOD
- 42 hours

SCORING SYSTEM:
- After a 42 hour incubation period, determination of the cytotoxic (irritancy) effect was performed. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT at the end of the treatment. Cell viability was calculated for each tissue as a percentage of the mean of the negative control tissues.

Irritation / corrosion parameter:

% tissue viability

Remarks:

percentage of control / time point: 15 minutes

Run / experiment:

1

Value:

102

Interpretation of results:

GHS criteria not met

Conclusions:

C.I. Leuco Sulphur Brown 37 is non-irritant in the in vitro skin irritation test

Executive summary:

The study was performed to investigate the skin irritation properties of the test substance according to OECD 439. The in vitro tissue model EPISKIN-SM was used as the test system. 5% (aq) Sodium dodecyl sulphate served as positive control and phosphate buffered saline as the negative control. The mean tissue viability was determined at 15 min after application of test item or the controls.

The positive control had a mean cell viability of 4% after 15 minutes exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the laboratory historical control data range. The standard deviation value of the percentage viability of three tissues treated identically was less than 5%, indicating that the test system functioned properly.

Skin irritation is expressed as the remaining cell viability after exposure to the test substance. The relative mean tissue viability obtained after 15 minutes treatment with C.I. Leuco Sulphur Brown 37 compared to the negative control tissues was 102%. Since the mean relative tissue viability for C.I. Leuco Sulphur Brown 37 was above 50% after 15 minutes treatment C.I. Leuco Sulphur Brown 37 is considered to be non-irritant.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14-Aug-2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU method B.47 (Bovine corneal opacity and permeability test method for identifying ocular corrosives and severe irritants)

Deviations:

no

GLP compliance:

yes

Species:

cattle

Amount / concentration applied:

TEST MATERIAL
- Amount applied: Due to the consistency of the test substance, dark brown paste, the test substance was applied directly on the corneas with a cotton swab in such a way that the cornea was completely covered

NEGATIVE CONTROL
- Amount applied: 750 µL of physiological saline per cornea

POSITIVE CONTROL
Amount applied: 750 µL per cornea
Concentration: 10% (w/v) Benzalkonium Chloride

Duration of treatment / exposure:

- Exposure: 10 minutes
- Post incubation period: 120 minutes

Details on study design:

TEST SITE
- Isolated bovine cornea

REMOVAL OF TEST SUBSTANCE
- Washing: yes
- Time after start of exposure: 10 minutes

SCORING SYSTEM:
- After the 10 minutes exposure and washing step, the corneas were incubated for 120 +/- 10 minutes at 32 °C. After the completion of the incubation period opacity determination was performed.

- The mean opacity and mean permeability values (OD490) were used for each treatment group to calculate an in vitro score:

In vitro irritancy score (IVIS) = mean opacity value + (15 x mean OD490 value).

TOOL USED TO ASSESS SCORE:
- opacitymeter and microplate reader

DATA EVALUATION:
A test substance that induces an IVIS ≥ 55.1 is defined as a corrosive or severe irritant

Irritation parameter:

in vitro irritation score

Remarks:

time point: 10 minutes

Value:

0

Interpretation of results:

GHS criteria not met

Conclusions:

C.I. Leuco Sulphur Brown 37 is not irritant or corrosive in the Bovine Corneal Opacity and Permeability test.

Executive summary:

The the test item was tested for its potential eye irritation properties in a Bovine Corneal Opacity and Permeability (BCOP) test according to OECD 437.

The negative control responses for opacity and permeability were less than the upper limits of the laboratory historical range indicating that the negative control did not induce irritancy on the corneas, except the response of one cornea. However since the opacity and permeability value were just above the upper limit of the range and all other values were less than the upper limits of the laboratory historical range, this deviation in the opacity and the premeability score had no effect on the results of the study.

 

The mean in vitro irritancy score of the positive control (10% (w/v) Benzalkonium Chloride) was 117 and was within the historical positive control data range. It was therefore concluded that the test conditions were adequate and that the test system functioned properly.

 

C.I. Leuco Sulphur Brown 37 did not induce ocular irritation through both endpoints, resulting in a mean in vitro irritancy score of 0 after 10 minutes of treatment.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation / corrosion endpoint:

The study was performed to investigate the skin irritation properties of the test substance according to OECD 439. The in vitro tissue model EPISKIN-SM was used as the test system. 5% (aq) Sodium dodecyl sulphate served as positive control and phosphate buffered saline as the negative control. The mean tissue viability was determined at 15 min after application of test item or the controls.

The positive control had a mean cell viability of 4% after 15 minutes exposure. The absolute mean OD570 (optical density at 570 nm) of the negative control tissues was within the laboratory historical control data range. The standard deviation value of the percentage viability of three tissues treated identically was less than 5%, indicating that the test system functioned properly.

Skin irritation is expressed as the remaining cell viability after exposure to the test substance. The relative mean tissue viability obtained after 15 minutes treatment with C.I. Leuco Sulphur Brown 37 compared to the negative control tissues was 102%. Since the mean relative tissue viability for C.I. Leuco Sulphur Brown 37 was above 50% after 15 minutes treatment C.I. Leuco Sulphur Brown 37 is considered to be non-irritant.

 

 

Eye irritation endpoint:

In the key study, the test item was evaluated for its potential eye irritation properties according to OECD 405. Three Rabbits were treated by instillation of 100.0 mg (range 99.6 – 100.3 mg) of the test substance (a volume of approximately 0.1 mL) in the conjunctival sac of one of the eye. The other eye left untreated served as a reference control. 

The eyes of each animal were examined approximately 1, 24, 48 and 72 hours after instillation of the test substance. The irritation scores and a description of all other (local) effects were recorded. The irritation was assessed according to OECD 405.

The treatment resulted in irritation of the conjunctivae, which consisted of redness and discharge. The irritation had completely resolved within 48 hours in one animal and within 72 hours in the other two animals. No iridial irritation or corneal opacity were observed, and treatment of the eyes with 2% fluorescein 24 hours after test substance instillation revealed no corneal epithelial damage. The test item was concluded to be slightly irritating to the eyes.

 

In a supplementary study, the test item was tested for its potential eye irritation properties in a Bovine Corneal Opacity and Permeability (BCOP) test according to OECD 437. Bovine eyes from young cattle obtained from the slaughterhouse were used as the test system and the test material was applied directly on the corneas to cover them completely. Additionally, physiological saline was used as negative control and 10% (w/v) Benzalkonium Chloride as positive control.

The negative control responses for opacity and permeability were less than the upper limits of the laboratory historical range indicating that the negative control did not induce irritancy on the corneas, except the response of one cornea. However since the opacity and permeability value were just above the upper limit of the range and all other values were less than the upper limits of the laboratory historical range, this deviation in the opacity and the permeability score had no effect on the results of the study.

The mean in vitro irritancy score of the positive control (10% (w/v) Benzalkonium Chloride) was 117 and was within the historical positive control data range. It was therefore concluded that the test conditions were adequate and that the test system functioned properly.

C.I. Leuco Sulphur Brown 37 did not induce ocular irritation through both endpoints, resulting in a mean in vitro irritancy score of 0 after 10 minutes of treatment. Under the conditions of this study, the test item was non-irritating to eyes.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008
The available in vitro experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008.

 

Skin: Based on available data on skin irritation, the test item is not classified according to Regulation (EC) No 1272/2008 (CLP), as amended for the eighteenth time in Regulation (EU) 2022/692.

 

Eye: Based on available data on eye irritation, the test item is not classified as according to Regulation (EC) No 1272/2008 (CLP), as amended for the eighteenth time in Regulation (EU) 2022/692.