2-methyl-2H-isothiazol-3-one hydrochloride

Endpoint summary

Description of key information

Skin corrosion

MIT.HCl was found to be corrosive to the skin using the three-dimensional human skin model EPIDERM comprising a reconstructed epidermis with a functional stratum corneum.

 

In a GLP, OECD 431 study, 25 mg of test article or 50 uL of negative or positive control were applied to EpiDerm tissue 3 and 60 minutes followed by a 42 hour post-incubation period and immediate determination of cytotoxic effects via the MTT reduction assay.

 

Corrosivity potential of the test article was predicted from the relative mean tissue viabilities obtained compared to the corresponding negative control tissues concurrently treated with distilled water. The positive control, 8N potassium hydroxide viability was <20% following a 60 minute exposure, thereby confirming thatcorrosivitycould be detected in this test system.

 

The test article showed evidence ofcorrosivityduring both the short term (3 minute) and prolonged exposures (60 minutes). The mean relative tissue viability was <50% (22%) after the 3 minute exposure and <15% (5.6%) after the 60 minute treatment (+ 42 hour post-incubation).

 

Under the conditions of this study the MIT.HCl showed irritant effects. The mean relative tissue viability was <50% after 3 minute (22%) and 1 hour (5.6%) exposure. Therefore test article,MIT.HCl, was considered to be

corrosive (Sub-category 1A) according to the UN GHS classification system.

 

Skin irritation

A waiver for the skin irritation endpoint has been submitted as the available information indicates that the criteria are met for classification as corrosive to the skin.

 

Eye irritation

A waiver for the skin irritation endpoint has been submitted as the available information indicates that the criteria are met for classification as corrosive to the skin. Therefore, as MIT.HCl is confirmed to cause skincorrosivity, no further testing is required for ocular irritation/corrosive, with MIT.HCl considered corrosive to eyes.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20 September 2017 to 21 September 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Version / remarks:

(2016)

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

- Identification: MIT.HCl (Methyl isothiazolinone hydrochloric acid)
- CAS no.: 26172-54-3
- Source and lot/batch No.of test material: Sigma-Aldrich / 10013913
- Expiration date of the lot/batch: 30 Nov 2018
- Purity test date: 23 Nov 2016
- Purity: >99.9%
- Apperance: White to light beige powder
- Moelcular weight: 151.61 g/mol
- Storage condition of test material: 2 to 8°C, protected from light

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

foreskin from multiple donors

Source strain:

other: Normal human epidermal keratinocytes (NHEK), derived from neonatal-foreskin tissue

Details on animal used as source of test system:

TEST KIT
- Source: MatTex Corporation
- Type: Three-dimensional human skin model, comprising a reconstructed epidermis with a functional stratum corneum

Cell culture media:
- Assay medium (MatTex Corporation)
- Vehicle / postive controls: purifed water / 8N KOH

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 37°C, humidified incubator
- CO2 (%): 5%

IN-LIFE DATES: 20 September 2017 to 21 September 2017

Vehicle:

unchanged (no vehicle)

Details on test system:

Pre-experimental checks:
To check the non-specific MTT-reducing capability of the test article 25 mg of the test article was mixed with 1 mL MTT (1 mg MTT/mL) medium and incubated for 1 hour and observed visually after stirring.

Assessment for Colour Interference
25 mg of test article was added to 0.3 mL of both deionized water and isopropanol and incubated for 60 minutes at 37°C, 5% CO2, 95% RH. Neither solution became coloured, therefore it was deemed not to have the potential to stain the tissue.

Control samples:

yes, concurrent vehicle

Amount/concentration applied:

50 uL of test article

Duration of treatment / exposure:

single exposure / 3 and 60 minutes

Duration of post-treatment incubation (if applicable):

42 h

Number of replicates:

2 replicates/dose/time point

Details on test animals or test system and environmental conditions:

n/a, in vitro test system used

Amount / concentration applied:

n/a, in vitro test system used

Duration of treatment / exposure:

n/a, in vitro test system used

Observation period:

n/a, in vitro test system used

Number of animals:

n/a, in vitro test system used

Details on study design:

n/a, in vitro test system used

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 minute exposure

Value:

ca. 22

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: positive indication of corrosivity

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 minute exposure

Value:

ca. 5.6

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

other: positive indication of corrosivity

Other effects / acceptance of results:

refer to "Any other information on results incl. tables"

Table 7.3.1/01-1: Summary of in vitro EPIDERM corrosivity result following application of MIT.HCL

Test Substance	Mean OD 579	Mean relative tissue viability (%) ±CV
3 minute exposure
Negative	1.247	100 ±1.7
Test article	0.269	22 ±4.9
Positive	0.319	-6 ±0.8
Positive FK	0.391
60 minute exposure
Negative	1.260	100 ±2.4
Test article	0.071	5.6 ±6.5
Positive	0.168	-0.1 ±2.6
Positive FK	0.170

FK          Freeze killed (Positive control tissue mean)

 

Skin viability after a three minute or one hour exposure to the test article was 22% and 5.6%, respectively.

The OD values for the negative controls met the acceptance criteria.

 

Skin viability after a three minute or one hour exposure to the positive control article was -6% and -0.1%, respectively, demonstrating appropriate performance of the assay.

 

All CVs between replicates were less than 30% and met the acceptance criteria.

 

The negative and positive control values fell within the laboratory historical control values.

Interpretation of results:

Category 1A (corrosive) based on GHS criteria

Conclusions:

Under the conditions of this study the MIT.HCl showed irritant effects. The mean relative tissue viability was <50% after 3 minute (22%) and 1 hour (5.6%) exposure. Therefore test article, MIT.HCl, was considered to be corrosive (Sub-category 1A) according to the UN GHS classification system

Executive summary:

This study was conducted to determine whether the test article,MIT.HCl, causes corrosion in thein vitroskin model EpiDerm^TM.

 

Duplicate EpiDerm^TM inserts were treated with test article, purified water (negative control) and 8N potassium hydroxide (positive control) for 3 minutes and 1 hour. At the end of the treatment period, the tissues were washed with phosphate buffered saline (PBS) and cell viability was assessed using the MTT assay. The skin corrosivity potential was classified according to the remaining cell viability obtained after test material treatment with either of the two treatment times.

 

Skin viability after a three minute or one hour exposure to the test article was 22% and 5.6%, respectively.

The OD values for the negative controls met the acceptance criteria.

 

Skin viability after a three minute or one hour exposure to the positive control article was -6% and -0.1%, respectively, demonstrating appropriate performance of the assay.

 

All CVs between replicates were less than 30% and met the acceptance criteria.

 

The negative and positive control values fell within the laboratory historical control values.

 

Under the conditions of this study the MIT.HCl showed irritant effects. The mean relative tissue viability was <50% after 3 minute (22%) and 1 hour (5.6%) exposure. Therefore test article, MIT.HCl, was considered to be corrosive (Sub-category 1A) according to the UN GHS classification system.

Reference 2

Endpoint:

skin irritation: in vitro / ex vivo

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study need not be conducted because the available information indicates that the criteria are met for classification as corrosive to the skin or irritating to eyes

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (corrosive)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vitro / ex vivo

Study period:

n/a

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because the substance is classified as skin corrosion, leading to classification as serious eye damage (Category 1)

Endpoint conclusion

Endpoint conclusion:

no study available

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

No studies are available; however in accordance with the guidance on the application of the CLP criteria, in addition to classification for skin corrosivity, where there is a likelihood of the test article being inhaled the following labelling EUH071 – Corrosive to the respiratory tract should be applied.

Justification for classification or non-classification

Comparison with the CLP criteria

- Skin:

Oct-7-en-1-ol was deemed to be corrosive to the skin in the EpiDerm in vitro corrosivity model conducted under OECD 431. The mean relative tissue viability was <50% (22%) after the 3 minute exposure and <15% (5.6%) after the 60 minute treatment (+ 42 hour post-incubation). In accordance with STEP 2 of the judgement criteria, as tissue viability was <25% following the 3 minute exposure, MIT.HCl is deemed to be corrosive to skin, Category 1A. 

 

Based on the results of this study, MIT.HCl was deemed to be corrosive to the skin, according to the criteria listed in OECD 431. Therefore, according to Annex I Regulation (EC) 1272/2008 the active ingredient, MIT.HCl is classified as Skin corrosion, Category 1A. 

 

- Eye:

A waiver for the skin irritation endpoint has been submitted as the available information indicates that the criteria are met for classification as corrosive to the skin. Therefore, as MIT.HCl is confirmed to cause skincorrosivity, no further testing is required for ocular irritation/corrosive, with MIT.HCl considered corrosive to eyes.

 

-Respiratory tract:

No studies are available; however in accordance with the guidance on the application of the CLP criteria, in addition to classification for skin corrosivity, where there is a likelihood of the test article being inhaled the following labelling EUH071 – Corrosive to the respiratory tract should be applied.

 

Overall conclusion:

Based on the results of this study, MIT.HCl showed irritant effects. The mean relative tissue viability was <50% after 3 minute (22%) and 1 hour (5.6%) exposure. Therefore test article,MIT.HCl, was considered to be

corrosive (Sub-category 1A) according to the UN GHS classification system. Consequently, as MIT.HCl is confirmed to cause skin corrosivity, no additional testing is required for ocular irritation/corrosion, with MIT.HCl considered corrosive to the eye - Serious Eye Damage Category 1. In addition, the following labelling EUH071 – Corrosive to the respiratory tract should be applied.