Reaction mass of (3R)-3-[(1R)-4-methyl-3-cyclohexen-1-yl]-1-butanol and (3S)-3-[(1R)-4-methyl-3-cyclohexen-1-yl]-1-butanol

Endpoint summary

Description of key information

Skin irritation: irritating, OECD 439, Warren 2013
Eye irritation: non-irritating, OECD 437, Warrent 2013
Eye irritation: non-irritating, rabbit, OECD 405 Prince 1983

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:

2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study performed under GLP.

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes

Species:

other: EPISKIN MODEL KIT

Strain:

not specified

Details on test animals or test system and environmental conditions:

The EPISKINTM model is a three-dimensional reconstructed human epidermis model consisting of adult human-derived epidermal keratinocytes seeded on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen. A highly differentiated and stratified epidermis model is obtained after a 13-Day culture period comprising of the main basal, supra basal, spinous and granular layers and a functional stratum corneum.

Incubations were at 37°C and 5% CO2.

Vehicle:

unchanged (no vehicle)

Controls:

other: Negative group treated with DPBS; Positive group treated with SDS (5% w/v).

Amount / concentration applied:

10 micolitre of test material was applied unchanged (no vehicle).

Duration of treatment / exposure:

15 minutes

Number of animals:

For each test group, triplicate tissues were used.

Details on study design:

TEST SITE
- Area of exposure: 10 μl of the undiluted test substance was added on top of the skin tissues.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes
- Time after start of exposure: 15 minutes.

SCORING SYSTEM: Skin irritation is expressed as the remaining cell viability after exposure to the test substance.

Irritation / corrosion parameter:

other: other: Mean tissue viability

Value:

9.6

Remarks on result:

other:

Remarks:

Basis: mean. Time point: 15 minutes. Reversibility: no data. Remarks: Score in terms of percentage of negative control group. (migrated information)

Other effects:

The MTT solution containing the test item did not turn blue which indicated that the test item did not directly reduce MTT.

Table 1. Mean OD₅₄₀ Values and Percentage Viabilities for the Negative Control Item, Positive Control Item and Test Item

Item	OD540of tissues	Mean OD540of triplicate tissues	± SD of OD540	Relative individual tissue viability (%)	Relative mean viability (%)	± SD of Relative mean viability (%)
Negative control	0.748	0.751	0.027	99.6	100	3.6
	0.725			96.5
	0.779			103.7
Positive control	0.092	0.066	0.023	12.3	8.8	3.1
	0.048			6.4
	0.057			7.6
Test material	0.07	0.072	0.002	9.3	9.6	0.3
	0.073			9.7
	0.074			9.9

Interpretation of results:

irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the conditions of this study, the test material is considered to be irritating to skin.

Executive summary:

The study was performed to assess the skin irritation potential of the test material to a human three dimensional epidermal model. The study was performed under GLP and followed the OECD guideline 439. Tissues were treated with the test material for 15 minutes and then washed with phosphate buffered saline to remove residual test material. Subsequently the skin tissues were incubated for 42 hours at 37°C in maintenance medium. After incubation, cell culture inserts were dried carefully to remove excess medium and were transferred into a 12-wells plate prefilled with 2 ml MTT-medium (0.3 mg/ml). The tissues were incubated for 3 h at 37°C. The amount of extracted formazan was determined spectrophotometrically at 540 nm. Cell viability was calculated for each tissue as a percentage of the mean of the negative control tissues. The relative mean tissue viability obtained after 15 minutes treatment with the test substance compared to the negative control tissues was 9.6%. Since the mean relative tissue viability for the test substance was below or equal to 50% after 15 minutes treatment it is considered to be a skin irritant.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Eye irritation

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation:

OECD 439 2013 - The study was performed to assess the skin irritation potential of the test material to a human three dimensional epidermal model. The study was performed under GLP and followed the OECD guideline 439. Tissues were treated with the test material for 15 minutes and then washed with phosphate buffered saline to remove residual test material. Subsequently the skin tissues were incubated for 42 hours at 37°C in maintenance medium. After incubation, cell culture inserts were dried carefully to remove excess medium and were transferred into a 12-wells plate prefilled with 2 ml MTT-medium (0.3 mg/ml). The tissues were incubated for 3 h at 37°C. The amount of extracted formazan was determined spectrophotometrically at 540 nm. Cell viability was calculated for each tissue as a percentage of the mean of the negative control tissues. The relative mean tissue viability obtained after 15 minutes treatment with the test substance compared to the negative control tissues was 9.6%. Since the mean relative tissue viability for the test substance was below or equal to 50% after 15 minutes treatment it is considered to be a skin irritant.

 

Human RIPT 1983 - The purpose of the test was to determine the irritation and/or sensitization potential of the test product after repeated application under occlusive patch test conditions to the skin of human subjects. The test was not performed under GLP and did not follow a recognised test method. A total of 58 subjects, 10 male and 48 female, ranging in age from 16 to 71 years were empaneled for this test. The test material was diluted to 25% in anhydrous alcohol prior to patching. The quantity of test material applied to each test patch was approximately 0.2 ml. For the induction, the test material was placed onto a Parke-Davis Readi-BandR (Webril) occlusive patch, which was applied to the back of each subject between scapulae and waist, adjacent to spinal mid-line. The site was scored 24 and 48 hours after application - a total of 47 subjects completed the test. Scattered, transient, barely perceptible to mild nonspecific skin responses were observed in 9/47 test panellists during the induction or challenge phases of the study. These non-specific patch test responses are not considered irritant or allergic in nature. Barely perceptible to mild irritant reactions were observed on 2/47 test panellists during the induction and/or challenge phases of the study. Neither the number of subjects reacting nor the severity of the reactions are considered indicative of a significant irritant effect. There was no evidence of induced allergic contact dermatitis. Under the conditions of this study the test material did not induce significant skin irritation in human subjects.

 

Eye irritation:

OECD 437 2013 - The study was performed to assess the eye irritancy potential of the test material in isolated bovine corneas. The study was performed under GLP and followed a method equivalent to OECD guideline 437. The ocular irritancy of the test substance was tested through topical application for 10 ± 1 minutes. 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. The mean in vitro irritancy score of the positive control (ethanol), was 36.2 and was within the historical positive control data range (22.5 to 60.9). The test substance did not induce ocular irritation through both endpoints, resulting in a mean in vitro irritancy score of 23.5 after 10 minutes of treatment. Under the conditions of this study the test material is not considered to be a severe irritant or corrosive in the Bovine Corneal Opacity and Permeability test.

 

equivalent to OECD 405 1983 - The study was performed to assess the eye irritancy potential of the test material in adult albino rabbits. The study was performed pre-GLP and followed a method similar to OECD guideline 405. The test substance was applied to the eye of each test animal without rinsing followed by observations for 14 days. No corneal or iris effects were observed in any animal. Redness, chemosis and discharge were observed but these effects had reversed by day 4. Under the conditions of this study the test material is not considered to be an eye irritant.

Justification for selection of skin irritation / corrosion endpoint:
Two studies available, one in vitro GLP compliant Klimisch 1; one non-GLP Human RIPT study Klimisch 4 where reliability cannot be assigned. Selected study is a validated in vitro study.

Justification for selection of eye irritation endpoint:
Two studies available, one in vitro GLP compliant Klimisch 1; one in vivo non-GLP study Klimisch 4 where reliability cannot be assigned. No study is selected since a Weight of Evidence determination is made based on in vitro data with supporting in vivo data. The available information satisfies the tonnage driven information requirements of the REACH Regulation (EC) 1907/2006.

Effects on skin irritation/corrosion: irritating

Justification for classification or non-classification

The substance meets classification criteria under EU Directive 67/548/EEC for dermal irritation: R38.

The substance meets classification criteria under Regulation (EC) No 1272/2008 for dermal irritation category 2.

 

The substance does not meet classification criteria under EU Directive 67/548/EEC for eye irritation.

The substance does not meet classification criteria under Regulation (EC) No 1272/2008 for eye irritation.