4-isopropyl-m-cresol

Endpoint summary

Description of key information

Skin irritation: irritating (OECD 404, GLP, K , rel. 1);
Eye irritation: corrosive (OECD 437, K, rel.1);
Respiratory irritation: no data available.

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From May 28 to June 20, 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study conducted in compliance with OECD Guideline No. 404 without any deviation.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

according to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

UK GLP Compliance Programme (inspected on July 10, 2012/ signed on November 30, 2012)

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Harlan Laboratories UK Ltd., Leicestershire, UK.
- Age at study initiation: 12-20 weeks
- Weight at study initiation: 2.66-3.40 kg
- Housing: Animals were housed individually in suspended cages.
- Diet: Food (2030 Teklad Global Rabbit diet supplied by Harlan Teklad, Oxon, UK), ad libitum
- Water: Mains drinking water, ad libitum
- Acclimation period: 5 days

ENVIRONMENTAL CONDITIONS
- Temperature: 17-23 °C
- Humidity: 30-70 %
- Air changes: 15 changes/h
- Photoperiod: 12 h dark / 12 h light

IN-LIFE DATES: From May 28 to June 20, 2014

Type of coverage:

semiocclusive

Preparation of test site:

clipped

Vehicle:

other: test item moistened sufficiently with 0.5 mL of distilled water

Controls:

not required

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 g
- For the purpose of the study the test item was ground to a powder prior to application.

Duration of treatment / exposure:

First animal: 3 minutes, 1 and 4 h
Second and third animal: 4 h

Observation period:

1, 24, 48 and 72 h after the removal of the patch

Number of animals:

3 males

Details on study design:

PRETRAEATMENT
- On the day before the test each of a group of three rabbits was clipped free of fur from the dorsal/flank area using veterinary clippers. Only animals with a healthy intact epidermis by gross observation were selected for the study.

TEST SITE
- Area of exposure: Dorsal/flank area
- Type of wrap if used: Test material moistened sufficiently with 0.5 mL distilled water to achieve a paste, was introduced under a 2.5 cm x 2.5 cm cotton gauze patch. Each patch was secured in position with a strip of surgical adhesive tape.
- To prevent the animal interfering with the patches, the trunk of the rabbit was wrapped in an elasticated corset for the duration of the exposure period.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): One patch was removed at each of three time points: 3 minutes, 1 and 4 h after application of test material. Any residual test material was removed by gentle swabbing with cotton wool soaked in distilled water.
- Time after start of exposure: 3 minutes, 1 and 4 h

SCORING SYSTEM: Draize scale, as described in OECD Guideline No. 404

Irritation parameter:

erythema score

Basis:

animal #1

Time point:

other: mean 24, 48 and 72 h

Score:

0

Max. score:

4

Irritation parameter:

edema score

Basis:

animal #1

Time point:

other: mean 24, 48 and 72 h

Score:

0

Max. score:

4

Irritation parameter:

erythema score

Basis:

animal #2

Time point:

other: mean 24, 48 and 72 h

Score:

0

Max. score:

4

Irritation parameter:

edema score

Basis:

animal #2

Time point:

other: mean 24, 48 and 72 h

Score:

0

Max. score:

4

Irritation parameter:

erythema score

Basis:

animal #3

Time point:

other: mean 24, 48 and 72 h

Score:

0

Max. score:

4

Irritation parameter:

edema score

Basis:

animal #3

Time point:

other: mean 24, 48 and 72 h

Score:

0

Max. score:

4

Irritant / corrosive response data:

No evidence of skin irritation was noted during the study.

Other effects:

All animals showed expected gain in body weight during the study.

Table 7.3.1/1: Mean irritant/corrosive response data for each animal at each observation time

Score at time point	Erythema (Animal no 1 / 2 / 3) Max. score 4	Oedema (Animal no 1 / 2 / 3) Max. score 4
Immediately	0 / 0 / 0	0 / 0 / 0
1 h	0 / 0 / 0	0 / 0 / 0
24 h	0 / 0 / 0	0 / 0 / 0
48 h	0 / 0 / 0	0 / 0 / 0
72 h	0 / 0 / 0	0 / 0 / 0
Average 24, 48 and 72 h	0 / 0 / 0	0 / 0 / 0

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the test conditions, Parathymol is not classified according to Regulation (EC) No. 1272/2008 (CLP) and Annex VI to the Directive 67/548/EEC.

Executive summary:

In a dermal irritation study performed according to the OECD Guideline No. 404, and in compliance with GLP, 0.5 g of test material moistened with distilled water was applied on the clipped skin of the dorsal flank area of 3 male New Zealand White rabbits. Test sites were covered with a semi-occlusive dressing for 4 h. Skin irritation was assessed and scored according to the Draize scale at 1, 24, 48 and 72 h after removal of the patch.

No evidence of skin irritation was observed following 3 minutes, 1 and 4 h exposure period. All animals showed expected gain in body weight during the study.

The individual scores for each animal within 3 scoring times (24, 48 and 72 h) were 0.0 / 0.0 / 0.0 for erythema and 0.0 / 0.0 / 0.0 for oedema.

Under the test conditions, Parathymoll is not classified according to Regulation (EC) No. 1272/2008 (CLP) and Annex VI to the Directive 67/548/EEC.

This study is considered as acceptable and satisfies the requirement for skin irritation endpoint.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not 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

Study period:

May 15, 2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study conducted according to OECD test Guideline No. 437 without any deviation.

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

according to guideline

Guideline:

OECD Guideline 437 (Bovine Corneal Opacity and Permeability Test Method for Identifying Ocular Corrosives and Severe Irritants)

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Remarks:

UK GLP Compliance Program (inspected on March 12 to 14, 2014 / Signed on May 12, 2014)

Species:

other: Bovine eye

Details on test animals or tissues and environmental conditions:

TEST SYSTEM
- Eyes from adult cattle (typically 12 to 60 months old) were obtained from a local abattoir as a by-product from freshly slaughtered animals. The eyes were excised by an abattoir employee after slaughter, and were placed in Hanks' Balanced Salt Solution (HBSS) supplemented with antibiotics (penicillin at 100 IU/mL and streptomycin at 100 mg/ml.) They were transported to the test facility over ice packs on the same day of slaughter. The corneas were prepared immediately on arrival.

Vehicle:

other: 0.9% w/v sodium chloride solution

Amount / concentration applied:

TEST MATERIAL
- Test Item Formulation and Experimental Preparation: For the purpose of this study the test item was prepared as a 20% w/v solution in 0.9% w/v sodium chloride solution. The test item was formulated within two hours of being applied to the test system. It is assumed that the formulation was stable for this duration.
- Amount(s) applied (volume or weight with unit): 0.75 mL was applied on each cornea

Duration of treatment / exposure:

Test item applied BCOP holder was incubated, anterior chamber uppermost at 32 ± 1 °C for 240 minutes.

Observation period (in vivo):

- Corneal opacity was measured pre-treatment, post-treatment and post-incubation (after 240 minutes of incubation).
- Application of Sodium Fluorescein: Following the opacity measurement the permeability of the corneas to sodium fluorescein (5 mg/mL) was evaluated and then corneal holders were incubated, anterior chamber uppermost at 32 ± 1 °C for 90 minutes.

Number of animals or in vitro replicates:

Total: 9 corneas - 3 corneas/group for test item, negative and positive controls

Details on study design:

Details of test procedure:
- Treatment of corneas: Corneas obtained from freshly slaughtered adult cattle (from a local abattoir) were mounted in corneal holders. Both chambers of each BCOP holder were filled with complete Eagle’s minimum essential medium (MEM) and incubated for 60 minutes at 32 ± 1 °C. Before the treatment, opacity measurement was performed using an opacitometer. MEM was removed from anterior chamber and the test item (0.75 mL) was applied on each cornea. The holders were incubated, anterior chamber uppermost, at 32 ± 1 °C for 240 minutes. At the end of the exposure period the test item and control items were removed from the anterior chamber and the cornea was rinsed three times with fresh complete MEM containing phenol red before a final rinse with complete MEM. The anterior and posterior chambers were refilled with fresh complete MEM. A post-treatment opacity reading was taken and each cornea was visually observed.
- Application of sodium fluorescein: Following the opacity measurement the permeability of the corneas to sodium fluorescein was evaluated. The medium from the anterior chamber was removed and replaced with 1 mL of sodium fluorescein solution (5 mg/mL). The dosing holes were plugged and the holders incubated, anterior chamber uppermost, at 32 ± 1 °C for 90 minutes.
- Permeability determinations: After incubation the medium in the posterior chamber of each holder was decanted and retained. 360 µL of medium representing each cornea was applied to a designated well on a 96-well plate and the optical density at 492 nm (OD492) was measured using the Anthos 2001 microplate reader.

Negative control: 0.9 % w/v sodium chloride solution
Positive control: Imidazole, 20% w/v solution in 0.9 % w/v sodium chloride solution

Irritation parameter:

other: In Vitro Irritancy Score (IVIS)

Basis:

mean

Time point:

other: 240 minutes

Score:

46.9

Max. score:

55

Irritant / corrosive response data:

- In Vitro Irritancy Score (IVIS) for test item, negative and positive controls were 46.9, 2.3 and 95.3, respectively.
- Corneal Epithelium Condition: The corneas treated with the test item were cloudy with sloughing post treatment. The corneas treated with the negative control item were clear post treatment and post incubation. The corneas treated with the positive control item were cloudy post treatment and post incubation.
- The IVIS for the test item was 46.9. This score is below the cut-off for classification of serious eye damage set at > 55.
- In the three test item treated corneas sloughing of the corneal epithelium was observed. However in two of the three test item treated corneas the sloughing is considered to be more advanced resulting in lower opacity values (upper cloudy layers of the cornea have sloughed away revealing clearer layers underneath) The opacity was markedly higher in the cornea that had not sloughed as significantly. It is therefore apparent that if the sloughing was not as advanced in the first two corneas the overall IVIS would have been higher.
- In addition all three test item treated corneas induced fluorescein permeability values greater than noted in the three positive control corneas.

Other effects:

None

The post treatment condition of the corneal epithelium was as follows:

 

Treatment	Epithelium Condition
Test Item	Clear in three corneas
Negative Control	Cloudy in three corneas
Positive Control	Cloudy & sloughing in three corneas

 

Criteria for an Acceptable Test

The positive control In Vitro Irritancy Score was within the range of 73.8 - 103.6. The positive control acceptance criterion was therefore satisfied.

 

The negative control gave opacity of ≤ 4.7 and permeability ≤ 0.085. The negative control acceptance criteria were therefore satisfied.

Interpretation of results:

corrosive

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Under the test conditions, Parathymol is classified as Category 1, H318 Causes serious eye damage according to the Regulation (EC) No. 1272/2008.

Executive summary:

In an in vitro eye irritation study performed according to the OECD Guideline 437 and in compliance with GLP, Parathymol was applied at a concentration of 20% w/v in 0.9% w/v sodium chloride solution for 240 minutes. Negative and positive control items were tested concurrently. The two endpoints, decreased light transmission through the cornea (opacity) and increased passage of sodium fluorescein dye through the cornea (permeability) were combined in an empirically derived formula to generate anIn VitroIrritancy Score (IVIS).

 

In Vitro Irritancy Score (IVIS) for test item, negative and positive controls were 46.9, 2.3 and 95.3, respectively. In the three test item treated corneas sloughing of the corneal epithelium was observed. However in two of the three test item treated corneas the sloughing was considered to be more advanced resulting in lower opacity values (upper cloudy layers of the cornea have sloughed away revealing clearer layers underneath.) The opacity was markedly higher in the cornea that had not sloughed as significantly. It is therefore apparent that if the sloughing was not as advanced in the first two corneas the overall IVIS would have been higher. In addition all three test item treated corneas induced fluorescein permeability values greater than noted in the three positive control corneas.

 

Under the test conditions,Parrathymol is classified as Category 1, H318 Causes serious eye damage according to the Regulation (EC) No. 1272/2008.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation / corrosion:

A stepwise approach was developped, in accordance with the ECHA R.7 guidance (ITS for skin irritation).

1/ An in vitro skin corrosion study was performed according to the OECD Guideline 431 and in compliance with GLP, using the EPISKIN(TM) reconstructed human epidermis model. The relative mean viabilities of the test item treated tissues were 119.2% for 240 minutes exposure; 116.9 % for 60 minutes exposure; and 183.7 % for 3 minutes exposure: 183.7%. All quality criteria required for acceptance of results in the test were satisfied. Under the test conditions, Parathymol is not classified as corrosive.

2/ An in vitro skin irritation study was performed according to the OECD Guideline 439 and in compliance with GLP, using the EPISKIN(TM) reconstructed human epidermis model. This test was run on two separate occasions. However both runs produced equivocal results. The equivocal results were due to significant variation measured between the identically treated tissues in test item groups of both the initial and repeated runs. Significant variation was also measured between identically treated tissues in the negative control group of the initial run. It was not possible to accurately predict the skin irritation potential of Parathymol.This study is therefore considered to be inconclusive.

3/ Based on the in vitro result, an in vivo study was performed. The dermal irritation study performed according to the OECD Guideline No. 404, and in compliance with GLP, 0.5 g of test material moistened with distilled water was applied on the clipped skin of the dorsal flank area of 3 male New Zealand White rabbits. Test sites were covered with a semi-occlusive dressing for 4 h. Skin irritation was assessed and scored according to the Draize scale at 1, 24, 48 and 72 h after removal of the patch. No evidence of skin irritation was observed following 3 minutes, 1 and 4 h exposure period. All animals showed expected gain in body weight during the study. The individual scores for each animal within 3 scoring times (24, 48 and 72 h) were 0.0 / 0.0 / 0.0 for erythema and 0.0 / 0.0 / 0.0 for oedema. Under the test conditions, Parathymol is not classified as skin irritant. This study is considered as the key study.

The acute dermal toxicity showed consistent results with the skin irritation study (see section 7.2.3). Indeed, only very slight erythema has been observed in the animals following 24-hour semi-occlusive exposure to the substance at 2000 mg/kg bw/d. The crust formation observed in females was likely to be a Keratin crust. It appeared as a dry, straw coloured crust which flakes to reveal normal or glossy skin. It’s generally stimulated by the over production of keratinocytes in the epidermis and the experts do not consider this reaction indicative of the full thickness tissue destruction that typifies dermal corrosion.

Eye irritation:

A key study was identified (Harlan, 2014). In this in vitro eye irritation study performed according to the OECD Guideline 437 and in compliance with GLP, Parathymol was applied at a concentration of 20% w/v in 0.9% w/v sodium chloride solution for 240 minutes. Negative and positive control items were tested concurrently. The two endpoints, decreased light transmission through the cornea (opacity) and increased passage of sodium fluorescein dye through the cornea (permeability) were combined in an empirically derived formula to generate anIn VitroIrritancy Score (IVIS).

In Vitro Irritancy Score (IVIS) for test item, negative and positive controls were 46.9, 2.3 and 95.3, respectively. In the three test item treated corneas sloughing of the corneal epithelium was observed. However in two of the three test item treated corneas the sloughing was considered to be more advanced resulting in lower opacity values (upper cloudy layers of the cornea have sloughed away revealing clearer layers underneath.) The opacity was markedly higher in the cornea that had not sloughed as significantly. It is therefore apparent that if the sloughing was not as advanced in the first two corneas the overall IVIS would have been higher. In addition all three test item treated corneas induced fluorescein permeability values greater than noted in the three positive control corneas.

Under the test conditions, Parrathymol is classified as corrosive to the eyes.

Justification for selection of skin irritation / corrosion endpoint:
The key study is GLP-compliant and of high quality (Klimisch score = 1)

Justification for selection of eye irritation endpoint:
The key study is GLP-compliant and of high quality (Klimisch score = 1)

Effects on eye irritation: corrosive

Justification for classification or non-classification

Harmonized classification:

The substance has no harmonized classification according to the Regulation (EC) No. 1272/2008.

Self-classification:

Based on the available information, Parathymol should be classified as Eye Dam. Category 1 (H318: Causes serious eye damage) according to the Annex VI of the Regulation (EC) No. 1272/2008 (CLP).

No additional self-classification is proposed regarding skin irritation according to the Annex VI of the Regulation (EC) No. 1272/2008 (CLP).

No information was available regarding respiratory irritation.