Imidazole

Endpoint summary

Description of key information

Imidazole is corrosive to skin under occlusive conditions and irritating to corrosive to the rabbit eye. The test methods were comparable with the corresponding OECD test guidelines 404 and 405. 

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:

1979

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable deviation (occlusive dressing). Occlusive dressing represents worst case conditions. The study was performed prior to the implementation of GLP.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 404 (Acute Dermal Irritation / Corrosion)

Principles of method if other than guideline:

Study was performed prior to the implementation of OECD Guidelines and GLP, but is in compliance with the principles described in OECD Guideline 404.

GLP compliance:

no

Species:

rabbit

Strain:

Vienna White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Zuchtbetrieb M. Gauckler, 63065 Offenbach, Germany
- Weight at study initiation: mean 2,7 kg
- Diet (ad libitum): Ssniff K standard diet for rabbits and guinea pigs from IMTERMAST GmbH, Germany
- Water: ad libitum

Type of coverage:

occlusive

Preparation of test site:

shaved

Vehicle:

water

Controls:

no

Amount / concentration applied:

Concentration: 80 % active substance
Ammounts applied: 0.5 ml

Duration of treatment / exposure:

1 hour (1 h experiment)
4 hour (4 h experiment)

Observation period:

The animals were observed 8 days and skin changes were recorded on working days.

Number of animals:

A total of 6 Vienna White rabbits; 4 animals (1 male, 3 females) in the 1h experiment, 2 females in the 4h experiment.

Details on study design:

- Area of exposure: clipped dorsal skin

REMOVAL OF TEST SUBSTANCE
- Washing (if done): water or a mixture of water/lutrole (1:1 v/v)
- Time after start of exposure: 1h and 4h

SCORING SYSTEM:
Score/Erythema/Oedema
0/not present/not present
1/very slight/very slight
2/mild/mild
3/moderate/moderate
4/severe/severe

Irritation parameter:

erythema score

Remarks:

4 h treatment

Basis:

animal #1

Time point:

other: mean of 1, 2 and 8 days

Score:

4

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Remarks on result:

other: comprehensive, parchment like skin necrosis at the end of the observation period

Irritation parameter:

erythema score

Remarks:

4 h treatment

Basis:

animal #2

Time point:

other: mean of 1, 2 and 8 days

Score:

4

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Remarks on result:

other: comprehensive, leathery skin necrosis at the end of the observation period

Irritation parameter:

edema score

Remarks:

4 h treatment

Basis:

animal #1

Time point:

other: mean of 1, 2 and 8 days

Score:

2

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Remarks on result:

other: light edema at the end of the observation period

Irritation parameter:

edema score

Remarks:

4 h treatment

Basis:

animal #2

Time point:

other: mean of 1, 2 and 8 days

Score:

2.33

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Remarks on result:

other: light edema at the end of the observation period

Irritation parameter:

erythema score

Remarks:

1 h treatment

Basis:

animal #1

Time point:

other: mean of 1, 2 and 8 days

Score:

2.67

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Remarks on result:

other: necrotic spots on the skin surface at the end of the observation period

Irritation parameter:

erythema score

Remarks:

1 h treatment

Basis:

animal #2

Time point:

other: mean of 1, 2 and 8 days

Score:

0

Max. score:

4

Remarks on result:

other: desquamation at the end of the observation period

Irritation parameter:

erythema score

Remarks:

1h treatment

Basis:

animal #3

Time point:

other: mean of 1, 2 and 8 days

Score:

1.33

Max. score:

4

Reversibility:

fully reversible within: 8 d

Remarks on result:

other: necrotic spots on the skin surface at the end of the observation period

Irritation parameter:

erythema score

Remarks:

1h treatment

Basis:

animal #4

Time point:

other: mean of 1, 2 and 8 days

Score:

1.67

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Remarks on result:

other: necrotic spots on the skin surface and desquamation at the end of the observation period

Irritation parameter:

edema score

Remarks:

1h treatment

Basis:

animal #1

Time point:

other: mean of 1, 2 and 8 days

Score:

< 1

Max. score:

4

Reversibility:

fully reversible within: 8 d

Irritation parameter:

edema score

Remarks:

1 h treatment

Basis:

animal #2

Time point:

other: mean of 1, 2 and 8 days

Score:

0

Max. score:

4

Reversibility:

fully reversible within: 8 d

Irritation parameter:

edema score

Remarks:

1 h treatment

Basis:

animal #3

Time point:

other: mean of 1, 2 and 8 days

Score:

1.33

Max. score:

4

Reversibility:

fully reversible within: 8 d

Irritation parameter:

edema score

Remarks:

1 h treatment

Basis:

animal #4

Time point:

other: mean of 1, 2 and 8 days

Score:

1.33

Max. score:

4

Reversibility:

fully reversible within: 8 d

Irritant / corrosive response data:

Skin reactions after 1-hr exposure time:
Mild erythema was seen in all (4/4) animals 1 hr after removal of the occlusive dressing. Erythema subsided in one animal overnight. Focal necrosis developed in the other 3 animals within 24 and 48 hr. Findings at the end of the observation period (8 d) were as follows: 2/4 animals showed no erythema but showed desquamation (1/4) and focal necrosis (1/4). Slight erythema, desquamation and extended focal necrosis was seen in the 3rd animal, and severe erythema with superficial necrosis was noted in the 4th animal. Oedema were absent after patch removal but grade 2 oedema were noted in 2/4 and 3/4 animals after 24 and 48 hr, respectively. Oedema had resolved on day 8 of observation.

Skin reactions after 4-hr exposure time:
Well defined, moderate erythema and oedema were seen in both animals after removal of the occlusive dressing. Erythema became severe overnight and rested severe until the end of the observation period. Soft necroses developed overnight in both animals. Leather-like necrosis indicative of full thickness necrosis was noted in both animals and confirmed by pathology after sacrifice of the animals. Oedema started to resolve overnight but rested as grade 2 until day 8.

Other effects:

General toxicity: No signs noted after 1-hr or 4-hr exposure.

On the basis of the results no substance specific destruction of skin tissue, namely, visible necrosis through the epidermis and into the dermis, after exposure up to a 1 hour duration was observed.

Interpretation of results:

corrosive

Remarks:

Migrated information

Conclusions:

After 4-hr occlusive exposure moderate erythema and oedema were noted within 1 hr. Severe erythema developed overnight and persisted until day 8, the end of the observation period. Soft necroses developed overnight. They hardened and changed into persistent leather-like necroses in both animals as noted on day 8. Slight oedema were noted at the end of the observation period.
No signs of general toxicity were noted but Imidazole was found to be corrosive.

Executive summary:

When imidazole was applied to the intact rabbit skin as a 80% aqueous paste (0.5 ml) for 1 or 4 hours under occlusive dressing skin reactions were noted as early as one hour after removal of the dressing .

After the 4 hour exposure period focal necrosis developed overnight in all animals and was described as leather-like at the end of the observation period when pathology confirmed full thickness necrosis after sacrifice of the animals.

After the 1 hour exposure under occlusive dressing mild erythema was seen in all (4/4) animals. Mild erythema and mild oedema were observed on the following two days of the study. The oedema resolved completely by day 8 of the post-exposure observation period. Residual signs included patchy, superficial necrotic lesions in addition to scaling.

On the basis of the results obtained after 1 hour exposure no substance specific destruction of skin tissue, namely, visible necrosis through the epidermis and into the dermis, was observed.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (corrosive)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1979

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Comparable to guideline study with acceptable deviations. The study was performed prior to the implementation of OECD TG 405 and GLP.

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 405 (Acute Eye Irritation / Corrosion)

Deviations:

yes

Remarks:

: 1h reading was not performed, observation period was 8 days (instead of 21 days)

Principles of method if other than guideline:

Study was performed prior to the implementation of OECD Guidelines and GLP, but is in compliance with the principles described in OECD Guideline 405. Deviations from the guideline were: 1-h reading was not performed, observation period was 8 days (instead of 21 days).

GLP compliance:

no

Species:

rabbit

Strain:

Vienna White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: Zuchtbetrieb M. Gauckler, 63065 Offenbach
- Weight at study initiation: mean 2.8 kg
- Diet (ad libitum): Ssniff K standard diet for rabbits and guinea pigs from INTERMAST GmbH, Soest, Germany
- Water: ad libitum

Vehicle:

unchanged (no vehicle)

Controls:

other: non-treated adjacent eye served as a control.

Amount / concentration applied:

Amount applied: volume of 0.1 ml

Duration of treatment / exposure:

After instillation of the test item the treated eye was not rinsed.

Observation period (in vivo):

Readings for eye irritation were done 24, 48 and 72 hours as well as 8 days after instillation.
The animals were observed several times on the treatment day and up to 8 days afterwards, and findings were recorded on working days.

Number of animals or in vitro replicates:

3

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing: The eyes were not washed out after 24 hours as specified in OECD Guideline 405

SCORING SYSTEM:
Scoring was performed according to the sytem descirbed in OECD 405.
Based on the scores the MMAS (Modified Maximum Average Score) was calculated.

Irritation parameter:

cornea opacity score

Basis:

animal #1

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

cornea opacity score

Basis:

animal #2

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

cornea opacity score

Basis:

animal #3

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

iris score

Basis:

animal #1

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

1

Max. score:

2

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

iris score

Basis:

animal #2

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

1

Max. score:

2

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

iris score

Basis:

animal #3

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

1

Max. score:

2

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

conjunctivae score

Basis:

animal #1

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

3

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

conjunctivae score

Basis:

animal #2

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

3

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

conjunctivae score

Basis:

animal #3

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

3

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

chemosis score

Basis:

animal #1

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

chemosis score

Basis:

animal #2

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

chemosis score

Basis:

animal #3

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2

Max. score:

4

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

other: secretion

Basis:

animal #1

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

2.33

Max. score:

3

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

other: secretion

Basis:

animal #2

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

1.33

Max. score:

3

Reversibility:

not fully reversible within: 8 d

Irritation parameter:

other: secretion

Basis:

animal #3

Time point:

other: 24, 48 and 72 hours after substance instillation

Score:

3

Max. score:

3

Reversibility:

not fully reversible within: 8 d

Irritant / corrosive response data:

Application of 0.1g unchanged imidazole to the rabbit's eye (Draize test ; according to Federal Register 38 no. 178 § 1500 .42 (1973)) affected iris, conjunctiva, cornea, and the nictating membrane of the animals. Grade 2 reddening and swelling of the conjunctiva was noted along with chemosis which aggravated and persisted to grade 3 until day 8. A slight grade 2 cornea opacity persisted until the end of the observation period on day 8. The affected corneal area comprised more than 3/4.

Interpretation of results:

Category 1 (irreversible effects on the eye)

Remarks:

Migrated information based on Regulation 1272/2008/EC

Executive summary:

Application of 0.1 g unchanged imidazole to the rabbit's eye affected iris, conjunctiva, cornea, and the nictating membrane of the animals. Grade 2 reddening and swelling of the conjunctiva was noted along with chemosis which aggravated and persisted to grade 3 until day 8. A slight grade 2 cornea opacity persisted until the end of the observation period on day 8. The affected corneal area comprised more than 3/4.

Endpoint conclusion

Endpoint conclusion:

adverse effect observed (irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation

In a patch test, the clipped dorsal skin of six rabbits (White Vienna) was exposed to a patch (2 cm x 2 cm) loaded with 0.5 ml of an aqueous paste of imidazole (imidazole concentration 80%) for 1 or 4 hours (“corrosion test”). Upon removal of the patch, the treated skin area was washed with polyethylene glycol 400 and subsequently with a mixture of polyethylene glycol 400 and water (1:1). Immediately after 4-hour exposure, the 2 exposed rabbits exhibited severe reddening of the area of exposure and beyond, accompanied by severe oedema. Soft necrosis and marked oedema were observed 24h after application. Mild oedema and necrosis with a parchment-like or leathery appearance were still visible at the end of the 8-day post exposure observation period. No signs of absorptive intoxication were observed after 4 hours of exposure. Imidazole was considered corrosive based on the results obtained after 4-hours of exposure. After the 1 hour exposure under occlusive dressing mild erythema was seen in all (4/4) animals. Mild erythema and mild oedema were observed on the following two days of the study. The oedema resolved completely by day 8 of the post-exposure observation period. Residual signs included patchy, superficial necrotic lesions in addition to scaling. On the basis of the results obtained after 1 hour exposure no substance specific destruction of skin tissue, namely, visible necrosis through the epidermis and into the dermis, was observed (BASF AG, 1979a).

 

Eye irritation

Application of 0.1 g unchanged imidazole to the rabbit's eye (BASF AG, 1979b) affected iris, conjunctiva, cornea, and the nictating membrane of the animals. Grade 2 reddening and swelling of the conjunctiva was noted along with chemosis which aggravated and persisted to grade 3 until day 8. A slight grade 2 cornea opacity persisted until the end of the observation period on day 8. The affected corneal area comprised more than 3/4. The observed manifestations of irreversible tissue damage and persistent large size cornea opacity indicate, that imidazole is severely irritating to corrosive to the rabbit eye.

In addition several in vitro assays were available:

Hayashi et al. (2012) used a Short Time Exposure test (STE), EpiOcular assay and Hen's Egg Test Chorioallantoic Membrane assay (HET-CAM) to assess eye irritation of several test substances including imidazole. A 5 minute exposure to 5 % imidazole solution in the STE test resulted in a cell viability of 1.4 ± 1.6 (% ± SD) and a classification as irritant. 90 minutes of exposure to 50 mg imidazole in theEpiOcular assay resulted in 6.8 ± 0.1(% ± SD) cell viability and based on this result imidazole was classified as irritant. The HET–CAM irritation score was 12 after an exposure period of 20 seconds to 0.2 g imidazole and was also classified as irritant. Kaluzhny et al. (2011) used the EpiOcular assay to assess potential eye irritation. Inserts from 4 different suppliers were dosed with 50µL imidazole (20% w/v) in duplicate for 30 minutes. Results were displayed as mean tissue viability ± difference in tissue viability. The cell viability after 30 min exposure in inserts 1, 2, 3 and 4 was 42.73% ± 14.98, 34.53 % ± 3.96, 37.03% ± 0.09 and 32.54 ± 0.00, respectively. Based on these results the test substance was classified in GHS categories 1 or 2. Kolle et al. (2011) used the EpiOcular assay to assess potential eye irritation. Two tissue were treated in two independent experiments for 90 ± 5 minutes exposure to 50 mg imidazole. This resulted in a mean cell viability of 2%. Imidazole was therefore categorized as a Category 1/2 irritant substance. Takahashi et al. (2011) tested used a Short Time Exposure test (STE) to assess eye irritation potential. A 5 minute exposure to 5% and 0.05% test substance resulted in a cell viability of 1.4% and 95.3% respectively. The test substance was moderate irritant based on the STE method and score. In accordance with OECD Guideline 437, Verstraelen et al. (2013) assessed 2 different methods to measure the opacity in the Bovine Corneal Opacity and Permeability assay. Tests were performed in duplicate. Using the standard OP-KIT an in vitro irritancy score (IVIS) of 89.5 ± 9.1 and 84.9 ± 2.1 (average IVIS ± SD) was obtained. Using a laser light-based opacitometer (LLBO) resulted in IVIS scores of 235.7 ± 20.0 and 204.4 ± 3.3. The in vitro classification based on the IVIS was corrosive/severe eye irritant.

Effects on skin irritation/corrosion: corrosive

Effects on eye irritation: highly irritating

Justification for classification or non-classification

Imidazole is corrosive to skin based on the results obtained from a patch test after 4 -hours occlusive exposure. Imidazole caused severely irritating to corrosive effects to the rabbit eye. Based on these results Imidazole is classified as follows: Skin corrosion / Irritation Category 1C / H314 , "Causes severe skin burns and eye damage" (Regulation 1272/2008/EC).