Registration Dossier

Administrative data

Description of key information

Skin irritation: In Vitro EpiDermTM Skin Irritation Test, Reconstructed Human Epidermal Model (OECD 439, GLP): not irritating
Skin irritation: In vivo, rabbit: Not irritating
Eye irritation: In vitro Bovine Corneal Opacity and Permeability Assay (BCOP) (OECD 437, GLP): not serious eye damaging (CLP/EPA/GHS (Cat 1)) but a prediction for the damage hazard cannot be made (GHS)
Eye irritation: In vivo, New Zealand White rabbit (OECD 405, GLP): mild irritant (Class 4 on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system / not an eye irritant based on Regulation (EC) 1272/2008
Eye irritation: In vivo, rabbit: slightly irritating to the 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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study on the registered substance itself.
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Version / remarks:
In Vitro EpiDermTM Skin Irritation Test (EPI-200-SIT) for use with MatTek Corporation’s Reconstructed Human Epidermal Model EpiDerm (EPI-200), Rev. 3/26/2012
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
By Hessisches Ministerium für Umwelt, Energie, Landwirtschaft und Verbraucherschutz
Species:
other: human-derived keratinocytes
Strain:
other: not applicable
Details on test animals and environmental conditions:
not applicable
Type of coverage:
other: not applicable
Preparation of test site:
other: not applicable
Vehicle:
other: Dulbecco's Phosphate Buffered Saline (DPBS)
Controls:
yes, concurrent vehicle
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 mg ≙ 39 mg/cm²

VEHICLE
- Amount(s) applied (volume or weight with unit): wetted with 50µl DPBS
Duration of treatment / exposure:
60 min
Observation period:
post incubation for 41.5 h in total
Number of animals:
not applicable, 3 replicates / treatment group
Details on study design:
TEST SITE
- % coverage: 100%

REMOVAL OF TEST SUBSTANCE
- Washing (if done): Yes, tissues were gently rinsed with DPBS at least 15 times.
- Time after start of exposure: 60 min
Irritation / corrosion parameter:
other: other: relative mean absorbance, MTT assay
Value:
88.9
Remarks on result:
other:
Remarks:
Basis: mean. Time point: 60min + 41.5h. Reversibility: other: not applicable. Remarks: test item. (migrated information)
Irritation / corrosion parameter:
other: other: relative mean absorbance, MTT assay
Value:
100
Remarks on result:
other:
Remarks:
Basis: mean. Time point: 60min + 41.5h. Reversibility: other: not applicable. Remarks: negative control. (migrated information)
Irritation / corrosion parameter:
other: other: relative mean absorbance, MTT assay
Value:
2.9
Remarks on result:
other:
Remarks:
Basis: mean. Time point: 60min + 41.5h. Reversibility: other: not applicable. Remarks: positive control. (migrated information)
Other effects / acceptance of results:
The mean relative absorbance value of the test item, corresponding to the cell viability, decreased to 88.9% (threshold for irritancy: ≤ 50%), consequently the test item was not irritant to skin.

Results after treatment with 3-Methyl-thiazolidin-2-thion and the controls:

 

Dose Group

Treat-ment Interval

Absor-bance 570 nm Tissue 1*

Absor-bance 570 nm Tissue 2*

Absor-bance 570 nm Tissue 3*

Mean Absor-bance of 3 Tissues

Rel. Absor-bance [%] Tissue 1, 2 + 3**

Relative Standard Deviation

[%]

Mean Rel. Absorbance

[% of Negative Control]***

Negative Control

60 min

1.999

1.903

1.976

1.959

102.0

97.1

100.8

2.5

100.0

Positive Control

60 min

0.047

0.063

0.061

0.057

2.4

3.2 3.1

15.0

2.9

Test Item

60 min

1.927

1.541

1.757

1.742

98.4

78.7

89.7

11.1

88.9

 

* Mean of three replicate wells after blank correction

** relative absorbance per tissue [rounded values]: 100 x (absorbancetissue) / (mean absorbancenegative control)

*** relative absorbance per treatment group [rounded values]: 100 x (mean absorbancetest item / negative control) / (mean absorbancenegative control)

 

The optical pre-experiment (colour interference pre-experiment) to investigate the test item’s colour change potential in water did not led to a change in colour.

Optical evaluation of the MTT-reducing capacity of the test item after 1 hour incubation with MTT-reagent did not show blue colour.

The mean relative absorbance value of the test item, corresponding to the cell viability, decreased to 88.9% (threshold for irritancy: ≤ 50%), consequently the test item was not irritant to skin.

Interpretation of results:
GHS criteria not met
Remarks:
UN GHS and EU CLP regulation
Conclusions:
Testing was performed via a GLP guideline study on the registered substance itself. The present in vitro method is recommended in a tiered testing approach and the validity criteria are met, making the results sufficiently reliable to assess the skin irritating potential of 3-Methyl-thiazolidin-2-thion. In conclusion, it can be stated that in this study and under the experimental conditions reported, 3-Methyl-thiazolidin-2-thion is not irritant to skin according to UN GHS and EU CLP regulation. The present result suffices to conclude that no hazard arises from 3-Methyl-thiazolidin-2-thion, no in vivo study is required.
Executive summary:

This in vitro study was performed to assess the irritation potential of 3-Methyl-thiazolidin-2-thion by means of the Human Skin Model Test.

The test item did not reduce MTT (test for direct MTT reduction), and it did not change colour when mixed with deionised water (test for colour interference). Also its intrinsic colour was not intensive. Consequently, additional tests with freeze-killed or viable tissues were not necessary.

Each three tissues of the human skin model EpiDerm™ were treated with the test item, the negative or the positive control for 60 minutes.

Approximately 25 mg of the test item were applied to each tissue, wetted with 50 μL of DPBS, and spread to match the surface of the tissue.

30 μL of either the negative control (DPBS) or the positive control (5% SLS) were applied to each tissue.

After treatment with the negative control the absorbance values were well in the required range of the acceptability criterion of mean OD ≥ 0.8 and ≤ 2.8 for the 60 minutes treatment interval, thus assuring the quality of the tissues.

Treatment with the positive control induced a sufficient decrease in the relative absorbance as compared to the negative control for the 60 minutes treatment interval, and thus assuring the validity of the test system.

After treatment with the test item 3-Methyl-thiazolidin-2-thion the mean relative absorbance value decreased irrelevantly to 88.9% compared to the relative absorbance value of the negative control. This value is above the threshold for irritancy of ≤ 50%. Therefore, the test item is not considered to possess an irritant potential.

In conclusion, it can be stated that in this study and under the experimental conditions reported, 3-Methyl-thiazolidin-2-thion is not irritant to skin.

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:
weight of evidence
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Well-documented GLP OECD guideline study on the registered substance itself without deviations
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Version / remarks:
adopted 02 October 2012
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
from the Department of Health of the Government of the United Kingdom
Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories UK Ltd., Leicestershire, UK
- Age at study initiation: 12-20 weeks
- Weight at study initiation: 2.36 - 2.76 kg
- Housing: Animals were individually housed in suspended cages
- Diet (e.g. ad libitum): 2930C Teklad Global Rabbit diet supplied by Harlan Laboratories UK Ltd., Oxon, UK, ad libitum
- Water (e.g. ad libitum): ad libitum
- Acclimation period: at least five days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): set to achieve limits of 17 to 23 °C
- Humidity (%): set to achieve limits of 30-70%
- Air changes (per hr): at least fifteen changes per hour
- Photoperiod (hrs dark / hrs light): 12 hours continuous light (06:00 to 18:00) and 12 hours darkness
Vehicle:
unchanged (no vehicle)
Controls:
other: untreated second eye
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 mL of the test item, which was found to weigh approximately 89 mg (as measured by gently compacting the required volume into an adapted syringe)
Duration of treatment / exposure:
continuous as not washed out after instillation
Observation period (in vivo):
72 h
Number of animals or in vitro replicates:
3
Details on study design:
REMOVAL OF TEST SUBSTANCE
- Washing (if done): no wash-out

SCORING SYSTEM: numerical evaluation according to Draize, J.H, 1977: Dermal and Eye Toxicity Tests In: Principles and Procedures for Evaluating the Toxicity of Household Substances, National Academy of Sciences, Washington DC, p. 48-49.

TOOL USED TO ASSESS SCORE: ophthalmoscope
Irritation parameter:
overall irritation score
Basis:
mean
Time point:
other: 1h
Score:
8.7
Max. score:
110
Reversibility:
fully reversible within: 72h
Irritation parameter:
overall irritation score
Basis:
mean
Time point:
other: 24h
Score:
5.3
Max. score:
110
Reversibility:
fully reversible within: 72h
Irritation parameter:
overall irritation score
Basis:
mean
Time point:
other: 48h
Score:
2.7
Max. score:
110
Reversibility:
fully reversible within: 72h
Irritation parameter:
overall irritation score
Basis:
mean
Time point:
other: 72h
Score:
0
Max. score:
110
Reversibility:
other: not applicable
Irritant / corrosive response data:
Ocular Reactions
Individual and group mean scores for ocular irritation are given in Table 1 and Table 2. No corneal or iridial effects were noted during the study.
Moderate conjunctival irritation was noted in all treated eyes 1 hour after treatment. Moderate conjunctival irritation was noted in one treated eye and minimal conjunctival irritation was noted in two treated eyes at the 24-Hour observation with minimal conjunctival irritation noted in all treated eyes at the 48-Hour observation.
All treated eyes appeared normal at the 72-Hour observation.
Other effects:
Body Weight
Individual body weights and body weight change are given in Table 3.
One animal showed a slight body weight loss and the other two animals showed an expected gain in body weight during the study.

Table 1Individual Scores and Individual Total Scores for Ocular Irritation

Rabbit Number and Sex

75057Male

75064Male

75065Male

IPR = 0

IPR = 0

IPR = 0

Time After Treatment

1Hour

24Hours

48Hours

72Hours

1Hour

24Hours

48Hours

72Hours

1Hour

24Hours

48Hours

72Hours

CORNEA

 

 

 

 

 

 

 

 

 

 

 

 

E=Degree of Opacity

0

0

0

0

0

0

0

0

0

0

0

0

F=Area of Cornea Involved

0

0

0

0

0

0

0

0

0

0

0

0

Score (E x F) x5

0

0

0

0

0

0

0

0

0

0

0

0

IRIS

 

 

 

 

 

 

 

 

 

 

 

 

D

0

0

0

0

0

0

0

0

0

0

0

0

Score (D x5)

0

0

0

0

0

0

0

0

0

0

0

0

CONJUNCTIVAE

 

 

 

 

 

 

 

 

 

 

 

 

A=Redness

1

1

1

0

2

2

1

0

2

1

1

0

B= Chemosis

2

1

0

0

2

1

1

0

1

1

0

0

C=Discharge

1

0

0

0

1

1

0

0

1

0

0

0

Score (A+B+C) x2

8

4

2

0

10

8

4

0

8

4

2

0

Total Score

8

4

2

0

10

8

4

0

8

4

2

0

IPR=Initial pain reaction

 

Table 2 IndividualTotalScoresand Group Mean Scores for Ocular Irritation

Rabbit Number and Sex

Individual Total Scores At:

1 Hour

24 Hours

48 Hours

72 Hours

75057Male

8

4

2

0

75064Male

10

8

4

0

75065Male

8

4

2

0

Group Total

26

16

8

0

Group Mean Score

8.7

5.3

2.7

0.0

 

Table 3 Individual Body Weights and Body Weight Change

Rabbit Number and Sex

Individual Body Weight (kg)

Body Weight Change (kg)

Day0

Day3

75057Male

2.41

2.46

0.05

75064Male

2.36

2.34

-0.02

75065Male

2.76

2.79

0.03

Interpretation of results:
GHS criteria not met
Conclusions:
The study was conducted under GLP according to OECD guideline 405 on the registered substance itself without deviations. The method is to be considered scientifically reasonable. Hence, the results can be considered as reliable to assess the eye irritating properties of 3-Methyl-thiazolidin-2-thion. As a previous BCOP study did not allow to exclude completely any eye irritating properties of the substance in humans, this in vivo study had to be performed. In the present study, the test item produced a maximum group mean score of 8.7 and was classified as a mild irritant (Class 4 on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system. This result is consistent with the previous one from the BCOP assay. According to Regulation (EC) 1272/2008, Table 3.3.2, Category for reversible eye effects, a substance must be classified as irritating to eyes Cat. 2, “if, when applied to the eye of an animal, a substance produces:
— at least in 2 of 3 tested animals, a positive response of:
- corneal opacity ≥ 1 and/or
- iritis ≥ 1, and/or
- conjunctival redness ≥ 2 and/or
- conjunctival oedema (chemosis) ≥ 2
— calculated as the mean scores following grading at 24, 48 and 72 hours after installation of the test material, and which fully reverses within an observation period of 21 days”
The mean scores over 24, 48 and 72 h for each animal are 0/0/0 (corneal opacity), 0/0/0 (iritis), 0.7/1.0/0.7 (conjunctival redness), and 0.3/0.7/0.3 (chemosis). So, the classification criteria are not met and 3-Methyl-thiazolidin-2-thion does not need to be classified as irritating to the eye according to Regulation (EC) 1272/2008.
Executive summary:

In a primary in vivo eye irritation study (OECD 405), 0.1 ml / 89 mg of 3-Methyl-thiazolidin-2-thion (unchanged) was instilled into the conjunctival sac of the right eye of 3 male 12-20 weeks old New Zealand White rabbits. No wash-out was done, the left eye served as untreated control. Animals then were observed for 72h. Irritation was scored by the method of Draize.

Moderate conjunctival irritation was initially produced. All treated eyes appeared normal at the 72-hour observation. The test item produced a maximum group mean score of 8.7 and was classified as a mild irritant (Class 4 on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system.

In this study, 3-Methyl-thiazolidin-2-thion is not an eye irritant based on Regulation (EC) 1272/2008.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

There are two studies available on skin irritation. One in vitro key study according to OECD 439 under GLP was assessed as Klimisch 1 and performed to confirm older, not assessable in vivo data, which is the selected supporting study. As the older study indicated no irritation potential of 3-Methyl-thiazolidin-2-thion, in vitro skin irritation testing was performed first over corrosion testing in a bottom up strategy. As the test substance was not predicted a skin irritant, the test substance does not require any further skin irritation testing for submission to regulatory agencies utilizing the GHS system, which is in accordance with in vitro testing strategy recommendations, e.g. by PETA. No further in vivo testing is required, as no additional information can be expected and no indication is given that the result „not irritating to the skin” may underestimate the hazard in human risk assessment. Hence, testing can be omitted due to animal welfare. This conclusion is supported by the barely existing eye irritation potential (not classified under the EU CLP regulation), as will be outlined below, because the eye is considered to be the more sensitive organ. If no irritating effects are seen here, it can be reasonably concluded that the substance doesn’t bear any irritation hazard to the skin.

 

With regard to eye irritation, there are three studies available. The primarily available information (supporting study) resulted from a summary of an old in vivo study, whose reliability could not be assessed as only the result was given, i.e. slightly irritating to the rabbit eye. Hence, to fulfil the tonnage-driven data requirements under REACH, in a tiered testing approach first an in vitro study was conducted. A BCOP-assay was performed under GLP according to OECD 437, allowing no doubt that the results are not reliable (first WoE study). 3-Methyl-thiazolidin-2-thion is not serious eye damaging (CLP/EPA/GHS (Cat 1)) but a prediction for the damage hazard cannot be made (GHS). In consequence, the present study does not suffice to exclude completely any eye irritating properties of the substance in humans, and an in vivo study needs to be performed as subsequent step in a tiered approach. According to ECHA’s document on Eye irritation / corrosion, in the case of a positive and definitive result from BCOP, ICE or FL, the substance can be classified as inducing “serious eye damage” (Cat 1 of CLP), and no further in vivo test is necessary. If the test revealed that the substance does not require any classification for eye irritation/corrosion, no further in vivo testing is necessary. With regard to the tonnage-driven data requirements, at Annex VIII, if neither of these conclusions can be made, a further test conducted in vivo, to assess the eye irritation potential is needed. So, a further GLP OECD guideline study (OECD 405) was conducted in vivo. It was also assessed with Klimisch 1 and does not allow any doubt that the results are not reliable (second WoE study). In the present study, the test item produced a maximum group mean score of 8.7 and was classified as a mild irritant (Class 4 on a 1 to 8 scale) to the rabbit eye according to a modified Kay and Calandra classification system. This result is consistent with the previous one from the BCOP assay. According to Regulation (EC) 1272/2008, Table 3.3.2, Category for reversible eye effects, a substance must be classified as irritating to eyes Cat. 2, “if, when applied to the eye of an animal, a substance produces:

— at least in 2 of 3 tested animals, a positive response of:

- corneal opacity1 and/or

- iritis1, and/or

- conjunctival redness2 and/or

- conjunctival oedema (chemosis)2

— calculated as the mean scores following grading at 24, 48 and 72 hours after installation of the test material, and which fully reverses within an observation period of 21 days”

The mean scores over 24, 48 and 72 h for each animal are 0/0/0 (corneal opacity), 0/0/0 (iritis), 0.7/1.0/0.7 (conjunctival redness), and 0.3/0.7/0.3 (chemosis). So, the classification criteria are not met and 3-Methyl-thiazolidin-2-thion does not need to be classified as irritating to the eye according to Regulation (EC) 1272/2008.

There are no reasons to believe that this result is not relevant for human risk assessment, and no further testing is considered necessary.

 

In summary, the tonnage-driven data requirements under REACH are met, there were no data gaps identified and no indication was given that the results are not relevant for human risk assessment or may underestimate the irritating hazard of 3-Methyl-thiazolidin-2-thion. The substance does not need to be classified as irritating to the skin nor eye according to the CLP Regulation (EC) 1272/2008.

Justification for classification or non-classification

All available studies relevant for classification indicate that 3-Methyl-thiazolidin-2-thion does not need to be classified as irritating to the skin nor eyes, as outlined above. In an in vitro OECD 439 study, treatment with 3-Methyl-thiazolidin-2-thionthe mean relative absorbance value decreased irrelevantly to 88.9% compared to the relative absorbance value of the negative control. This value is above the threshold for irritancy of ≤ 50%. Therefore, the test item is not considered to possess an irritant potential, which is sufficient for non-classification. In an OECD 405 study, the mean scores over 24, 48 and 72 h for each animal were determined to be 0/0/0 (corneal opacity), 0/0/0 (iritis), 0.7/1.0/0.7 (conjunctival redness), and 0.3/0.7/0.3 (chemosis). So, the classification criteria are not met and 3-Methyl-thiazolidin-2-thion does not need to be classified as irritating to the eye according to Regulation (EC) 1272/2008.