Chloroacetone

Endpoint summary

Description of key information

The test item was determined to be corrosive to skin (reference 7.3.1-1).

A study for eye irritation does not need to be conducted as the test item is classified for skin corrosion (reference 7.3.2-1).

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:

2020-12-01 to 2020-12-03

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: Council Regulation 440/2008, Method B.40 BIS: “In Vitro Skin Corrosion: Human Skin Model Test”

Version / remarks:

May 30, 2008

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

18 June, 2019

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: donors not further specified, consent was obtained

Justification for test system used:

The EpiDerm Skin Model is a well-established organotypic, three-dimensional model of the human epidermis and is used for in vitro experiments since many years. It is known for its similarity to human skin.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: reconstituted three-dimensional human skin model EpiDerm
- Tissue batch number: 34106
- Production date: not specified
- Shipping date: not specified
- Delivery date: not specified
- Date of initiation of testing: 2020-12-01

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 3 min treatment: room temperature; 60 min treatment: 37 ± 1 °C
- Temperature of post-treatment incubation: 37 ± 1 °C,

REMOVAL OF TEST MATERIAL AND CONTROLS
-Volume and number of washing steps: gently rinsed about 20 times with PBS
- Observable damage in the tissue due to washing: No
- Modifications to validated SOP: No

MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL medium
- Incubation time: 3 h
- Spectrophotometer: plate spectrophotometer
- Wavelength: 570 nm
- Filter: not specified
- Filter bandwidth: not specified
- Linear OD range of spectrophotometer: not specified

FUNCTIONAL MODEL CONDITIONS WITH REFERENCE TO HISTORICAL DATA
As provided on CoA for tissue lot:
- Viability: Acceptance criteria: OD 1.0-3.0; result: 1.654+/-0.2
- Barrier function: Acceptance criteria: ET-50 4.77-8.72 h; result: 6.59 h
- Contamination: Acceptance criteria: no contamination; result: sterile

NUMBER OF REPLICATE TISSUES: 2

NUMBER OF INDEPENDENT TEST SEQUENCES / EXPERIMENTS TO DERIVE FINAL PREDICTION: 2

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

NEGATIVE CONTROL
- Amount applied: 50 μL

POSITIVE CONTROL
- Amount applied: 50 μL

Duration of treatment / exposure:

3 and 60 min

Duration of post-treatment incubation (if applicable):

3 h

Number of replicates:

2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

60 min exposure

Value:

9.2

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Remarks:

100

Positive controls validity:

valid

Remarks:

8.6

Remarks on result:

positive indication of irritation

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 min exposure

Value:

46.5

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Remarks:

100

Positive controls validity:

valid

Remarks:

13.7

Remarks on result:

positive indication of irritation

Other effects / acceptance of results:

OTHER EFFECTS
- Visible damage on test system: No
- Direct-MTT reduction: The mixture of 50 μL test item per 1 mL MTT medium showed no reduction of MTT as compared to the solvent.
- Colour interference with MTT: The mixture of 50 μL test item per 300 μL Aqua dest. and per 300 μL isopropanol showed no relevant colouring as compared to the solvent.

DEMONSTRATION OF TECHNICAL PROFICIENCY: not specified

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
- Range of historical values: Please refer to "Any other information on results"

Table 1: Results of 3 min Experiment

Name	Negative Control		Positive Control		Test Item
Replicate Tissue	1	2	1	2	1	2
Absolute OD570	2.414	1.891	0.316	0.312	1.039	0.898
	1.693	1.930	0.321	0.309	1.035	0.914
	2.392	1.910	0.331	0.314	1.041	0.908
Mean Absolute OD570	2.038****		0.317		0.972
OD570 - Blank Corrected	2.370	1.847	0.272	0.268	0.995	0.853
	1.649	1.866	0.277	0.265	0.990	0.870
	2.348	1.866	0.287	0.270	0.997	0.864
Mean OD570 of 3 Aliquots (Blank Corrected)	2.122	1.866	0.278	0.268	0.994	0.862
SD OD570 of 3 Aliquots	0.410	0.020	0.008	0.002	0.003	0.008
Total Mean OD570 of 2 Replicate Tissues (Blank Corrected)	1.994*		0.273		0.928
SD OD570 of 2 Replicate Tissues	0.181		0.0088		0.093
Mean Relative Tissue Viability [%]	100.0		13.7		46.5
Coefficient Of Variation [%]***	9.1		2.8		10.00

* corrected mean OD₅₇₀ of the negative control corresponds to 100 % absolute tissue viability

*** coefficient of variation (CV) (in the range of 20 – 100 % viability) between two tissues treated identically is ≤ 30 %

**** The mean absolute OD₅₇₀ of the negative control is ≥ 0.8 and ≤ 2.8

 

Table 2: Results of 60 min Experiment

Name	Negative Control		Positive Control		Test Item
Replicate Tissue	1	2	1	2	1	2
Absolute OD570	2.038	2.070	0.252	0.173	0.227	0.221
	1.963	2.094	0.259	0.179	0.234	0.224
	1.985	2.099	0.257	0.175	0.235	0.225
Mean Absolute OD570	2.042****		0.216		0.228
OD570 - Blank Corrected	1.994	2.026	0.208	0.129	0.183	0.177
	1.919	2.050	0.215	0.134	0.190	0.180
	1.941	2.055	0.212	0.131	0.191	0.181
Mean OD570 of 3 Aliquots  (Blank Corrected)	1.951	2.043	0.212	0.132	0.188	0.179
SD OD570 of 3 Aliquots	0.038	0.015	0.003	0.003	0.005	0.002
Total Mean OD570 of 2 Replicate Tissues (Blank Corrected)	1997*		0.172		0.184
SD OD570 of 2 Replicate Tissues	0.065		0.057		0.006
Mean Relative Tissue Viability [%]	100.0		8.6**		9.2
Coefficient Of Variation [%]***	3.3		33.1		3.4

* corrected mean OD₅₇₀ of the negative control corresponds to 100% absolute tissue viability

** mean relative tissue viability of the 60 min positive control is ≤ 15 %

*** coefficient of variation (CV) (in the range of 20 – 100 % viability) between two tissues treated identically is ≤ 30 %

**** The mean absolute OD₅₇₀ of the negative control is ≥ 0.8 and ≤ 2.8

 

Table 3: Test Acceptance Criteria

	Value	Cut off	Pass/fall
Mean Absolute OD570 nm NC (3 min Experiment)	2.038	0.8 ≤ NC ≤ 2.8	pass
Mean Absolute OD570 nm NC (60 min Experiment)	2.042	0.8 ≤ NC ≤ 2.8	pass
Mean Relative Tissue Viability [%] of PC (60 min experiment)	8.6	≤ 15 %	pass
CV [%] (in the range of 20 – 100 % viability)	3.3 – 10.0	≤ 30%	pass

 

Table 4: Historical Data

	Mean	SD	Range of LCL – UCL	n
OD570 of NC (3 min Experiment)	1.749	0.225	1.298 – 2.199	60
OD570 of NC (60 min Experiment)	1.800	0.202	1.395 – 2.204	61
Relative Tissue Viability [%] of PC (60 min experiment)	5.8	2.2	1.3 – 10.3	61
CV [%] (in the range of 20 – 100 % viability)	7.1	10.7	0.0 – 28.6	300

LCL:     Lower control limit (95 %, mean – 2*SD)

UCL:     Upper control limit (95 %, mean + 2*SD)

n:         number of control values

Historical control data were generated from 2015 - 2020.

Conclusions:

The test item was determined to be corrosive to skin.

Executive summary:

A study according to OECD 431 was conducted to assess the skin corrosivity potential the test item. Since corrosive chemicals are cytotoxic after a short time exposure to the stratum corneum of the epidermis the cytotoxic effects of the test item on EpiDerm, a reconstituted three-dimensional human epidermis model, were determined. Hereby, the test item was applied topically. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT after a 3 min and 60 min exposure period and compared to those of the concurrent negative controls.

The mixture of 50 μL test item per 1 mL MTT medium showed no reduction of MTT as compared to the solvent. The mixture did not turn blue/purple. Therefore, NSMTT equalled 0 %.

The mixture of 50 μL test item per 300 μL Aqua dest. and per 300 μL isopropanol showed no colouring as compared to the solvent. Therefore, NSC equalled 0 %.

The test item showed no non-specific reduction of MTT and no relevant colouring potential after mixture with aqua dest. and with isopropanol. Therefore, no additional controls for correction of possible false-negative results were necessary.

The controls confirmed the validity of the study. The mean OD570nm of the two negative control tissues was ≥ 0.8 and ≤ 2.8 for each exposure period (2.038, 2.042). The mean relative tissue viability (% negative control) of the positive control was ≤ 15 % (8.6 %) after 60 min treatment. The coefficient of variation (CV) (in the range of 20 – 100 % viability) of replicate tissues of all dose groups was ≤ 30 % (3.3 % - 10.0 %).

The test item showed corrosive effects. The mean relative tissue viability (% negative control) was reduced to 46.5 % after 3 min treatment and to 9.2 % after 60 min exposure to the test item.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (corrosive)

Eye irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin corrosion

A study according to OECD 431 was conducted to assess the skin corrosivity potential the test item. Since corrosive chemicals are cytotoxic after a short time exposure to the stratum corneum of the epidermis the cytotoxic effects of the test item on EpiDerm, a reconstituted three-dimensional human epidermis model, were determined. Hereby, the test item was applied topically. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT after a 3 min and 60 min exposure period and compared to those of the concurrent negative controls.

The mixture of 50 μL test item per 1 mL MTT medium showed no reduction of MTT as compared to the solvent. The mixture did not turn blue/purple. Therefore, NSMTT equalled 0 %.

The mixture of 50 μL test item per 300 μL Aqua dest. and per 300 μL isopropanol showed no colouring as compared to the solvent. Therefore, NSC equalled 0 %.

The test item showed no non-specific reduction of MTT and no relevant colouring potential after mixture with aqua dest. and with isopropanol. Therefore, no additional controls for correction of possible false-negative results were necessary.

The controls confirmed the validity of the study. The mean OD570nm of the two negative control tissues was ≥ 0.8 and ≤ 2.8 for each exposure period (2.038, 2.042). The mean relative tissue viability (% negative control) of the positive control was ≤ 15 % (8.6 %) after 60 min treatment. The coefficient of variation (CV) (in the range of 20 – 100 % viability) of replicate tissues of all dose groups was ≤ 30 % (3.3 % - 10.0 %).

The test item showed corrosive effects. The mean relative tissue viability (% negative control) was reduced to 46.5 % after 3 min treatment and to 9.2 % after 60 min exposure to the test item.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008

The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. Based on available data on skin corrosion, the test item is classified for skin corrosion Category 1 (H314: Causes severe skin burns and eye damage) and therefore also for causing serious damage to the eye Category 1 according to Regulation (EC) No 1272/2008 (CLP), as amended for the seventeenth time in Regulation (EU) 2021/849.