1,2-Heptanediol

Endpoint summary

Description of key information

In an in vitro skin irritation study according to OECD guideline 439, the test item was irritant to skin under the experimental conditions reported.

 

In an in vitro skin corrosion study according to OECD guideline 431, the test item was non-corrosive to skin under the reported experimental conditions reported.

 

In an ex vitro eye corrosion study according to OECD guideline 437, the test item was serious eye damaging under the experimental conditions reported.

 

Conclusion: The test substance is skin irritating and eye damaging.

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:

weight of evidence

Study period:

2020-08-18

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: MatTek Corporation Protocol: In vitro EpiDermTM Skin Irritation Test (EPI-200-SIT) For use with MatTek Corporation’s Reconstructed Human Epidermal Model EpiDerm (EPI-200-SIT)

Version / remarks:

2019-02-10

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2012-07-06

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

2019-06-18

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: All cells used to produce EpiDerm are derived from tissue obtained by MatTek Corporation from accredited institutions.

Source strain:

other: not applicable, cells from human donors are used

Details on animal used as source of test system:

Not applicable, cells from human donors are used

Justification for test system used:

In an international prevalidation study performed by ECVAM, the in vitro skin irritation test using the human skin model EpiDerm™ and EpiSkin™ and measurement of cell viability by dehydrogenase conversion of MTT into a blue formazan salt have turned out as a sufficiently promising predictor for skin irritancy potential.

Vehicle:

unchanged (no vehicle)

Remarks:

The test item was tested neat.

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: Epi-200-SIT (MatTek Corporation, 82105 Bratislava, Slovakia) is a three-dimensional human epidermis model which develops a highly differentiated and stratified epidermis analogous to those found in vivo.
- Tissue batch number: 30880
- Date of initiation of testing: 12th May 2020

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: At 37 ± 1.5°C (first 35 min), room temperature (last 25 min)
- Temperature of post-treatment incubation: 37 ± 1.5°C

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: The tissues were gently rinsed with PBS several times in order to remove any residual test material

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL in DMEM
- Incubation time: 3 hours in MTT solution, 3.5 hours in isorpopanol
- Spectrophotometer: Versamax® (Molecular Devices)
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: OD 0.03 to OD 0.11 ( positive control, corresponds to 2.24% - 6-19% viability according to HCD of laboratory); ≥ OD 0.8 and ≤ OD 2.8 (negative control, according to OECD439/MatTek protocol); ≥ OD 1.28 and ≤ OD 2.0 (according to HCD of laboratory)
- Barrier function: The barrrier function has been tested by ET-50 assay using 100 µL Triton X-100, 4 time points, n=3 using MTT assay. The acceptance criterium was fulfilled when the ET-50 was reached between 4.77 and 8.72 hours. The test result of the tissue batch used was 5.51 hours.
- Contamination: The cells used to produce EpiDerm tissue were screened for potential biological contaminants (HIV-1 virus, Hepatitis B and C virus by oligonucleotide direct amplification, bacteria, yeast and other funghi were ensured by long-term antibiotic, antimycotic-free culture). None of the listed contaminants were detected in the tissue batch used.

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Fresh tissues / killed tissues: Since test item/ water and test item/ isopropanol solutions did not change color significantly and the test item did not interfere with MTT, no additional tissues were necessary in the main experiment.

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 1

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
- The test substance is considered to be corrosive or irritant to skin if the tissue viability after exposure and post-incubation is ≤50%.
- The test substance is considered to be non-irritant to skin if the tissue viability after exposure and post-incubation is ≥50%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

yes, concurrent MTT non-specific colour control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 30 µL
- Concentration: The item was applied undiluted

NEGATIVE CONTROL
- Amount applied: Not indicated
- Concentration: The negative control (DPBS) was applied undiluted

POSITIVE CONTROL
- Amount applied: Not indicated
- Concentration: 5% SDS in deionised water

Duration of treatment / exposure:

1 hour (35 min at 37°C and 25 min at room temperature)

Duration of post-treatment incubation (if applicable):

45.5 hours (including a medium change after 25.5 hours)

Number of replicates:

3 tissue replicates, one experiment

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Mean of 3 tissue replicates

Value:

2.84

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

positive indication of irritation

Other effects / acceptance of results:

OTHER EFFECTS:
- Visible damage on test system: Not indicated
- Direct-MTT reduction: The test item did not interfere with MTT.
- Colour interference with MTT: The test item/ water and test item/ isopropanol solutions did not change color significantly.

DEMONSTRATION OF TECHNICAL PROFICIENCY:
See "Attached background material"

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes
- Acceptance criteria met for variability between replicate measurements: Yes, the standard deviations between the three tissue percentage viability values of each group (test item, positive and negative controls) in the main test were below 18
- Range of historical values if different from the ones specified in the test guideline: See "Details on test system"

Interpretation of results:

study cannot be used for classification

Remarks:

The test item is identified as requiring classification and labelling according to UN GHS/ EU CLP “Category 2” or “Category 1”. Further testing is required to resolve between UN GHS Categories 1 and 2 and decide on the final classification of the test item.

Conclusions:

In conclusion, it can be stated that in this study and under the experimental conditions reported, the test item is irritant to skin.

Executive summary:

This GLP compliant in vitro study was performed to assess the irritation potential of the test item by means of the Human Skin Model Test according to OECD guideline 439. The test item did not prove to be a MTT reducer in the MTT interference pre-experiment. Also, its intrinsic color was not intensive and it did not change color when mixed with deionised water or isopropanol. Therefore, additional tests with freeze-killed tissues and/or viable tissues (without MTT addition) did not have to be performed. Each three tissues of the human skin model EpiDerm™ were treated with the test item, the negative control (DPBS) or the positive control (5% SDS) for 60 minutes. After treatment with the negative control the absorbance values were well within 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 viability as compared to the negative control for the 60 minutes treatment interval, thus assuring the validity of the test system. After treatment with the test item, the mean relative viability value was 2.84% compared to the relative absorbance value of the negative control. This value is below the threshold for irritancy of ≤ 50%. Therefore, the test item is considered to possess an irritant potential. In conclusion, it can be stated that in this study and under the experimental conditions reported, the test item is irritant to skin. The test item is identified as requiring classification and labelling according to UN GHS/ EU CLP “Category 2” or “Category 1”, since the tissue viability after exposure and post-incubation is less or equal to 50%. Further testing is required to resolve between UN GHS Categories 1 and 2 and decide on the final classification of the test item.

Reference 2

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

2020-07-30 to 2020-11-02

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: MatTek test protocol “In vitro EpiDermTM Skin Corrosion Test (EPI-200-SCT)”

Version / remarks:

2014-11-07

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted 2019-06-18

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: human

Source strain:

other: not applicable, human cells

Details on animal used as source of test system:

Not applicable

Justification for test system used:

Validation studies have shown that tests employing human skin models are able to discriminate between known skin corrosives (Optional subcategory 1A, optional subcategory 1B and 1C) and non-corrosives.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm model (Epi-200 SCT kit)
- Tissue batch number: 30883
- Date of initiation of testing: 2020-08-04

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: At room temperature (3-minute treatment period), at 37 ± 1.5°C (1-hour treatment period)
- Temperature of post-treatment incubation: No post-treatment incubation was used.

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: The wells were gently rinsed with PBS to remove any residual test material for several times. Excess PBS was removed by gently shaking the tissue insert and blotting the lower surface with blotting paper.
- Observable damage in the tissue due to washing: Not indicated

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL in DMEM
- Incubation time: 3 hours
- Spectrophotometer: Versamax®, Molecular Devices
- Wavelength: 570 nm

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
- Viability: See "Any other infomation on materials and methods, incl. tables, table 1+2"
- Barrier function: The barrrier function has been tested by ET-50 assay using 100 µL Triton X-100, 4 time points, n=3 using MTT assay. The acceptance criterium was fulfilled when the ET-50 was reached between 4.77 and 8.72 hours. The test result of the tissue batch used was 5.7 hours.
- Contamination: The cells used to produce EpiDerm tissue were screened for potential biological contaminants (HIV-1 virus, Hepatitis B and C virus by oligonucleotide direct amplification, bacteria, yeast and other funghi were ensured by long-term antibiotic, antimycotic-free culture). None of the listed contaminants were detected in the tissue batch used.

NUMBER OF REPLICATE TISSUES: 2

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
- Fresh tissues / killed tissues: The test item did not reduce MTT and it did not change colour when mixed with deionised water and isopropanol. Consequently, additional tests with freeze-killed and viable tissues to determine correction factors for calculating the true viability in the main experiment were not necessary.

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 2 (two different time points)

PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be corrosive to skin if the viability after 3 minutes exposure is less than 50%, or if the viability after 3 minutes exposure is greater than or equal to 50 % and the viability after 1 hour exposure is less than 15%.
- The test substance is considered to be non-corrosive to skin if the viability after 3 minutes exposure is greater than or equal to 50% and the viability after 1 hour exposure is greater than or equal to 15%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount applied: 50 μL
- Concentration: The test item was tested neat.

NEGATIVE CONTROL
- Amount applied: 50 μL

POSITIVE CONTROL
- Amount applied: 50 μL
- Concentration: 8.0 N Potassium Hydroxide

Duration of treatment / exposure:

3 minutes and 1 hour

Duration of post-treatment incubation (if applicable):

No post-treatment incubation was used.

Number of replicates:

Two tissue replicates were used.

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Experiment II (1 hour incubation)

Value:

20.06

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Experiment I (3 minutes incubation)

Value:

90.52

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

OTHER EFFECTS
- Direct-MTT reduction: No
- Colour interference with MTT: No

DEMONSTRATION OF TECHNICAL PROFICIENCY: According to OECD 431 the laboratory demonstrated technical proficiency by correctly classifying the twelve recommended Proficiency Substances (see OECD 431, Table 1).

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes
- Acceptance criteria met for positive control: Yes
- Acceptance criteria met for variability between replicate measurements: Yes

Interpretation of results:

study cannot be used for classification

Remarks:

The test item is not identified as requiring classification and labelling according to UN GHS/ EU CLP “Category 1”. Further testing is required to identify UN GHS Category 2 and decide on the final classification of the test item.

Conclusions:

In conclusion, it can be stated that in this study and under the reported experimental conditions, the test item is non-corrosive to skin according to EU CLP and UN GHS.

Executive summary:

This GLP compliant in vitro study was performed to assess the corrosive potential of the test item by means of the Human Skin Model Test with EpiDerm™ tissues models according to OECD 431. The test item did not reduce MTT (pre-test for direct MTT reduction), and it did not change colour when mixed with deionised water and isopropanol (pre-test for colour interference). Consequently, additional tests with freeze-killed and viable tissues to determine correction factors for calculating the true viability in the main experiment were not necessary. Independent duplicate tissues of EpiDerm™ were exposed to either the test item, the negative control (Dulbecco's Phosphate-Buffered Saline (DPBS)) or the positive control (8.0 N KOH) for 3 minutes and 1 hour, respectively. After exposure to the negative control the absorbance values met the required acceptability criterion of a mean OD570 ≥ 0.8 and ≤ 2.8 for both treatment intervals thereby confirming the acceptable quality of the tissues. Exposure to the positive control induced a decrease in the relative absorbance as compared to the negative control, both for the 3 minutes exposure period and for the 1 hour exposure period. The 1 hour exposure caused a decrease of the cell viability < 15% of the negative control. The CV in the range 20 – 100% viability between the tissue replicates is ≤ 30%, thus the validity of the test system and the specific batch of the tissue models is confirmed. After exposure of the tissues to the test item the relative absorbance value was 90.52% after 3 minutes exposure. After 1 hour exposure the relative absorbance value was 20.06%. Both values did not reach the threshold for corrosivity which is defined to be < 50% after the 3 minutes exposure or ≥ 50% after 3 minutes exposure and < 15% after the 1 hour exposure. Therefore, the test item is considered to be not corrosive. In conclusion, it can be stated that in this study and under the reported experimental conditions, the test item is non-corrosive to skin according to EU CLP and UN GHS.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (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

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)

Version / remarks:

2017-12-09

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)

Version / remarks:

2010-10-09

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Species:

cattle

Strain:

other: 14 month old donor cattle

Details on test animals or tissues and environmental conditions:

SOURCE OF COLLECTED EYES
- Number of animals: Not indicated, but 9 corneae in total were used
- Characteristics of donor animals: 14 month old donor cattle
- Storage, temperature and transport conditions of ocular tissue: The isolated eyes were stored in HBSS containing 1% (v/v) penicillin/streptomycin (100 units/mL penicillin and 100 μg/mL streptomycin) in the cooled slaughter-house and during transportation on the same morning to the laboratory using a Styrofoam box. The corneae were isolated on the same day after delivery of the eyes.
- Time interval prior to initiating testing: On the same day the cattle was slaughtered
- Indication of any existing defects or lesions in ocular tissue samples: All eyes were carefully examined macroscopically for defects.
- Indication of any antibiotics used: 1% (v/v) penicillin/streptomycin (100 units/mL penicillin and 100 μg/mL streptomycin) were used in the storage solution and incubation medium
- Selection and preparation of corneas: Only intact corneas were selected, those presenting defects such as vascularization, pigmentation, opacity and scratches were discarded. The cornea was carefully removed from the eye using scalpel and rounded scissors. A rim of about 2 mm of tissue (sclera) was left for stability and handling of the isolated cornea.
- Quality check of the isolated corneas: Only corneae with a value of the basal opacity < 7 were used.

Vehicle:

unchanged (no vehicle)

Controls:

yes, concurrent positive control

yes, concurrent negative control

Amount / concentration applied:

TEST MATERIAL
- Amount applied: 0.75 mL
- Concentration: The neat test substance was used.

Duration of treatment / exposure:

Ten minutes (± 30 seconds)

Duration of post- treatment incubation (in vitro):

210 minutes (120 minutes until the second second opacity reading, another 90 minutes until permeability measurements)

Number of animals or in vitro replicates:

3 corneas per test item/positive control/negative control

Details on study design:

NUMBER OF REPLICATES: Three replicates

NEGATIVE CONTROL USED: Saline (0.9% NaCl in deionised water)

POSITIVE CONTROL USED: 2-Ethoxyethanol (purity: 99%)

APPLICATION DOSE AND EXPOSURE TIME: 0.75 mL of the neat test item

TREATMENT METHOD: Not indicated

POST-INCUBATION PERIOD: Yes, 210 minutes

REMOVAL OF TEST SUBSTANCE
- Number of washing steps after exposure period: At least three times or more until phenol red was still discoloured (yellow or purple), or the test item was still visible. Once the medium was free of test item, the corneas were given a final rinse with cMEM without phenol red.
- POST-EXPOSURE INCUBATION: In cMEM at 32 ± 1 °C for further two hours in a vertical position

METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: Using an opacitometer (OP_KiT opacitometer, Electro Design, 63-Riom France). The opacity of each of the corneae was determined by reading each holder placed in the photoreceptor compartment for treated cornea.
- Corneal permeability: passage of sodium fluorescein dye measured with the aid of microtiter plate reader at OD490

SCORING SYSTEM: In Vitro Irritancy Score (IVIS)

DECISION CRITERIA: Decision criteria as indicated in the TG were used.

Irritation parameter:

in vitro irritation score

Run / experiment:

Mean of three eyes

Value:

64.55

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

positive indication of irritation

Interpretation of results:

Category 1 (irreversible effects on the eye) based on GHS criteria

Conclusions:

In conclusion, according to the current study and under the experimental conditions reported, the test item is serious eye damaging (CLP/EPA/GHS (Cat 1).

Executive summary:

This GLP compliant in vitro study according to OECD guideline 437, was performed to assess the corneal damage potential of the test item by means of the BCOP assay using fresh bovine corneae. After a first opacity measurement of the fresh bovine corneae (t0), the neat test item, the positive, and the negative controls were applied to the different corneae and incubated for 10 minutes at 32 ± 1 °C. After the incubation phase, the test item as well as the positive and the negative controls were each rinsed from the corneae. Further, the corneae were incubated for another 120 minutes at 32 ± 1 °C in incubation medium, and opacity was measured for a second time (t130). 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 (mean IVIS = 0.89).
The positive control (2-Ethoxyethanol) was tested undiluted and showed clear opacity and distinctive permeability of the corneae (mean IVIS = 108.29) corresponding to a classification as serious eye damaging (CLP/EPA/GHS (Cat 1)).
The test item was tested undiluted. Relative to the negative control, the test item caused an increase of the corneal opacity and permeability. The calculated mean IVIS was 64.55 (threshold for serious eye damage: IVIS > 55). According to OECD 437, the test item is classified as serious eye damaging.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irreversible damage)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation in vitro, RL1

A GLP compliant in vitro study was performed to assess the irritation potential of the test item by means of the Human Skin Model Test according to OECD guideline 439. The test item did not prove to be a MTT reducer in the MTT interference pre-experiment. Also, its intrinsic color was not intensive and it did not change color when mixed with deionised water or isopropanol. Therefore, additional tests with freeze-killed tissues and/or viable tissues (without MTT addition) did not have to be performed. Each three tissues of the human skin model EpiDerm™ were treated with the test item, the negative control (DPBS) or the positive control (5% SDS) for 60 minutes. After treatment with the negative control the absorbance values were well within 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 viability as compared to the negative control for the 60 minutes treatment interval, thus assuring the validity of the test system. After treatment with the test item, the mean relative viability value was 2.84% compared to the relative absorbance value of the negative control. This value is below the threshold for irritancy of ≤ 50%. Therefore, the test item is considered to possess an irritant potential. In conclusion, it can be stated that in this study and under the experimental conditions reported, the test item is irritant to skin. The test item is identified as requiring classification and labelling according to UN GHS/ EU CLP “Category 2” or “Category 1”, since the tissue viability after exposure and post-incubation is less or equal to 50%. Further testing is required to resolve between UN GHS Categories 1 and 2 and decide on the final classification of the test item.

 

Skin corrosion in vitro, RL1

A GLP compliant in vitro study was performed to assess the corrosive potential of the test item by means of the Human Skin Model Test with EpiDerm™ tissues models according to OECD 431. The test item did not reduce MTT (pre-test for direct MTT reduction), and it did not change colour when mixed with deionised water and isopropanol (pre-test for colour interference). Consequently, additional tests with freeze-killed and viable tissues to determine correction factors for calculating the true viability in the main experiment were not necessary. Independent duplicate tissues of EpiDerm™ were exposed to either the test item, the negative control (Dulbecco's Phosphate-Buffered Saline (DPBS)) or the positive control (8.0 N KOH) for 3 minutes and 1 hour, respectively. After exposure to the negative control the absorbance values met the required acceptability criterion of a mean OD570 ≥ 0.8 and ≤ 2.8 for both treatment intervals thereby confirming the acceptable quality of the tissues. Exposure to the positive control induced a decrease in the relative absorbance as compared to the negative control, both for the 3 minutes exposure period and for the 1 hour exposure period. The 1 hour exposure caused a decrease of the cell viability < 15% of the negative control. The CV in the range 20 – 100% viability between the tissue replicates is ≤ 30%, thus the validity of the test system and the specific batch of the tissue models is confirmed. After exposure of the tissues to the test item the relative absorbance value was 90.52% after 3 minutes exposure. After 1 hour exposure the relative absorbance value was 20.06%. Both values did not reach the threshold for corrosivity which is defined to be < 50% after the 3 minutes exposure or ≥ 50% after 3 minutes exposure and < 15% after the 1 hour exposure. Therefore, the test item is considered to be not corrosive. In conclusion, it can be stated that in this study and under the reported experimental conditions, the test item is non-corrosive to skin according to EU CLP and UN GHS.

 

Eye corrosion ex vivo, RL1

A GLP compliant in vitro study according to OECD guideline 437 was performed to assess the corneal damage potential of the test item by means of the BCOP assay using fresh bovine corneae. After a first opacity measurement of the fresh bovine corneae (t0), the neat test item, the positive, and the negative controls were applied to the different corneae and incubated for 10 minutes at 32 ± 1 °C. After the incubation phase, the test item as well as the positive and the negative controls were each rinsed from the corneae. Further, the corneae were incubated for another 120 minutes at 32 ± 1 °C in incubation medium, and opacity was measured for a second time (t130). 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 (mean IVIS = 0.89). The positive control (2-Ethoxyethanol) was tested undiluted and showed clear opacity and distinctive permeability of the corneae (mean IVIS = 108.29) corresponding to a classification as serious eye damaging (CLP/EPA/GHS (Cat 1)). The test item was tested undiluted. Relative to the negative control, the test item caused an increase of the corneal opacity and permeability. The calculated mean IVIS was 64.55 (threshold for serious eye damage: IVIS > 55). According to OECD 437, the test item is classified as serious eye damaging.

 

Conclusion

The test substance is skin irritating and eye damaging.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008
The available experimental test data is reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on the available data, the test substance is classified for skin irritation (category 2) and serious eye damage (category 1) according to Regulation (EC) No 1272/2008 (CLP), as amended for sixteenth time in Regulation (EU) No 2021/743.