diethyl 1-(2,4-dichlorophenyl)-5-methyl-4,5-dihydro-1H-pyrazole-3,5-dicarboxylate

Endpoint summary

Description of key information

Skin irritation (OECD 404), rabbit: not irritating

Eye irritation (OECD 405), rabbit: not irritating

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records

Reference

Reference 1

Endpoint:

skin irritation: in vivo

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Qualifier:

according to guideline

Guideline:

other: OECD Guidelines for Testing of Chemicals "Phototoxicity after cutaneous application" DRAFT Guideline, 14 March 1990

Deviations:

not specified

Principles of method if other than guideline:

Following systemic or local application chemical substances may produce an increased reactivity of the skin to irradiation with ultraviolet and/or visible light. This phenomenon is called "phototoxicity" or "photoirritancy", arises after the first exposure and is not related with the immune system. The pathogenic reaction is based on a cytotoxic photooxidation of the respective compound on the site of irradiation. Therefore the localization of signs of dermal irritation is limited to the light-exposed area.

GLP compliance:

yes

Species:

guinea pig

Strain:

other: Pirbright-White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: HOECHST AG, Kastengrund, SPF breeding colony
- Age at study initiation: approx. 8 weeks
- Weight at study initiation: 261 g mean
- Housing: in fully air-conditioned rooms in Makrolon cages (Type 4) on soft wood granulate, in groups of 5 animals
- Diet (e.g. ad libitum): Altromin 3112 mixed diet for guinea pigs and rabbits ad libitum
- Water (e.g. ad libitum): tap water in plastic bottles, ad libitum
- Acclimation period: at least 1 day


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3
- Humidity (%): 50 ± 20
- Photoperiod (hrs dark / hrs light): 12/12

Type of coverage:

open

Preparation of test site:

shaved

Vehicle:

other: DAE

Controls:

other: solvent control area on each of 5 animals

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.06 mL
- Concentration (if solution): 20, 10 and 5% in vehicle


VEHICLE
- Concentration (if solution): 40% dimethylacetamid, 30% acetone, 30% ethanol

Duration of treatment / exposure:

4, 24 and 48 h (substance remains not removed)

Observation period:

4, 24 and 48 h

Number of animals:

10 females

Details on study design:

TEST SITE
- Area of exposure: 6 cm by 8 cm area on the back, subdivided into 4 sites of equal size


REMOVAL OF TEST SUBSTANCE
- Washing (if done): no


SCORING SYSTEM: Draize

Irritation parameter:

erythema score

Remarks:

20 % test substance

Basis:

mean

Time point:

other: 4 h

Score:

1.8

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Remarks:

20 % test substance

Basis:

mean

Time point:

other: 24 h

Score:

1.4

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Remarks:

20 % test substance

Basis:

mean

Time point:

other: 48 h

Score:

1

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Remarks:

10 % test substance

Basis:

mean

Time point:

other: 4 h

Score:

1.3

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Remarks:

10 % test substance

Basis:

mean

Time point:

other: 24 h

Score:

1.3

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Remarks:

10 % test substance

Basis:

mean

Time point:

other: 48 h

Score:

0.9

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Remarks:

5 % test substance

Basis:

mean

Time point:

other: 4 h

Score:

0

Max. score:

4

Reversibility:

other: no effects

Irritation parameter:

erythema score

Remarks:

5 % test substance

Basis:

mean

Time point:

other: 24 h

Score:

0.3

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Remarks:

5 % test substance

Basis:

mean

Time point:

other: 48 h

Score:

0.3

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

edema score

Basis:

mean

Time point:

other: 4, 24, 48 h

Score:

0

Max. score:

4

Reversibility:

other: no effects

Irritant / corrosive response data:

Positive reactions were seen on all treated spots in an unequivocal concentration-dependent manner in an incidence of 100 , 100 and 40 % in the three treatment groups, reactions in the form of slight to moderate erythema were mostly pronounced in the highest treatment groups of the 20 and 10 % solutions. In the positive control with 0.02 % 8-methoxypsoralen 4 out of 5 treated animals showed slight erythema.

Other effects:

No clinical signs of intoxication were observed at any time during the study. The condition of the animals showed no signs of any disturbances of food consumption or body weight gains.

Interpretation of results:

other: phototoxic

Reference 2

Endpoint:

skin irritation: in 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 404 (Acute Dermal Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPP 81-5 (Acute Dermal Irritation)

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: HOECHST AG, Kastengrund, conventional breed
- Age at study initiation: approx. 3 - 5 months
- Weight at study initiation: 2.9 - 4.0 kg
- Housing: individually, in fully air-conditioned rooms, separate cages (arranged in a battery)
- Diet: Altromin 2123 maintenance diet - rabbits (Altromin GmbH, Lage/Lippe), ad libitum and hay (approx. 15 g daily)
- Water: deionised, chlorinated water from automatic water dispensers, ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20±3
- Humidity (%): 50±20
- Photoperiod (hrs dark / hrs light): 12/12

Type of coverage:

semiocclusive

Preparation of test site:

shaved

Vehicle:

other: sesame oil

Controls:

no

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.5 g
- Concentration (if solution): moistened


VEHICLE
- Amount(s) applied (volume or weight with unit): 0.35 mL

Duration of treatment / exposure:

4 h

Observation period:

1, 24, 48, 72 h

Number of animals:

6 (4 males, 2 females)

Details on study design:

TEST SITE
- Area of exposure: 6.25 cm²
- Type of wrap if used: surgical plaster with a 2.5 x 2.5 cm cellulose patch (specially produced by Beiersdorf AG, Hamburg) was fixed onto the shaved skin area of each animal. 0.5 g moistened test substance was applied under each patch. The area was then covered with a semi-occlusive bandage.


REMOVAL OF TEST SUBSTANCE
- Washing (if done): with warm tap water
- Time after start of exposure: 4 h

SCORING SYSTEM: Draize

Irritation parameter:

erythema score

Basis:

animal: #1, #2. #3, #5, #6

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritation parameter:

erythema score

Basis:

animal #4

Time point:

24/48/72 h

Score:

0.6

Max. score:

4

Reversibility:

fully reversible within: 72 hours

Irritation parameter:

edema score

Basis:

animal: #1, #2. #3, #4, #5, #6

Time point:

24/48/72 h

Score:

0

Max. score:

4

Reversibility:

other: not applicable

Irritant / corrosive response data:

The test substance caused minor effects on the skin of two animals in the form of very slight erythema (grade 1, in one animal only at 1 hour after removal of the test substance). These signs of irritation were completely reversible within 3 days.

Other effects:

No clinical signs of systemic toxicity were observed.

Interpretation of results:

other: CLP/EU GHS criteria not met, no classification required according to Regulation (EC) No. 1272/2008

Conclusions:

CLP: not classified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records

Reference

Endpoint:

eye irritation: in 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 405 (Acute Eye Irritation / Corrosion)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPP 81-4 (Acute Eye Irritation)

Deviations:

no

GLP compliance:

yes

Species:

rabbit

Strain:

New Zealand White

Details on test animals or tissues and environmental conditions:

TEST ANIMALS
- Source: HOECHST AG, Kastengrund, conventional breed
- Age at study initiation: approx. 3 - 5 months
- Weight at study initiation: 3.4 - 4.2 kg
- Housing: in fully air-conditioned rooms, separate cages (arranged in a battery)
- Diet (e.g. ad libitum): Altromin 2123 maintenance diet - rabbits (Altromin GmbH, Lage/Lippe), ad libitum, and hay (approx. 15 g daily)
- Water (e.g. ad libitum): deionised, chlorinated water from automatic
water dispensers, ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20±3
- Humidity (%): 50±20
- Photoperiod (hrs dark / hrs light): 12/12

Vehicle:

unchanged (no vehicle)

Controls:

other: untreated eyes served as controls

Amount / concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 0.1 g

Duration of treatment / exposure:

24 h

Observation period (in vivo):

1, 24, 48, 72 h

Number of animals or in vitro replicates:

6 (5 females, 1 male)

Details on study design:

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes, physiological saline
- Time after start of exposure: 24 hours


SCORING SYSTEM: Draize


TOOL USED TO ASSESS SCORE: fluorescein

Irritation parameter:

cornea opacity score

Basis:

mean

Remarks:

of 6 animals

Time point:

24/48/72 h

Score:

0.38

Max. score:

4

Reversibility:

fully reversible within: 72 hours

Irritation parameter:

iris score

Basis:

mean

Remarks:

of 6 animals

Time point:

24/48/72 h

Score:

0.11

Max. score:

2

Reversibility:

fully reversible within: 48 hours

Irritation parameter:

conjunctivae score

Basis:

mean

Remarks:

of 6 animals

Time point:

24/48/72 h

Score:

0.83

Max. score:

3

Reversibility:

fully reversible within: 72 hours

Irritation parameter:

chemosis score

Basis:

mean

Remarks:

of 6 animals

Time point:

24/48/72 h

Score:

0.33

Max. score:

4

Reversibility:

fully reversible within: 72 hours

Irritant / corrosive response data:

The test substance caused effects in the eyes in the form of conjunctival reddening, chemosis and discharge, inflammation and opacity of the iris, ranging in degree from slight to moderate. All signs of irritation were completely reversible within 72 hours.

Other effects:

No clinical signs of systemic toxicity were observed.

Means (24 - 72 hours) of individual eye irritation scores:

Animal #	Cornea opacity (max. 4)	Iris (max. 2)	Conjunctiva redness (max. 3)	Conjunctiva chemosis (max. 4)
1	0	0	0.33	0
2	0.33	0	1.00	0
3	0.33	0	0.66	0
4	0.66	0.33	0.66	0
5	0.66	0.33	1.33	1.00
6	0.33	0	1.00	1.00
Mean	0.38	0.11	0.83	0.33

Interpretation of results:

other: CLP/EU GHS criteria not met, no classification required according to Regulation (EC) No. 1272/2008

Conclusions:

CLP: Not classified

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Skin irritation

A study on skin irritation was performed in rabbits according to OECD guideline 404 and in compliance with GLP. 0.5 mg of moistened test substance was applied to the shaved skin of six New Zealand White rabbits. The skin was exposed for 4 h under semi-occlusive conditions. Local skin reactions were assessed according to the Draize scoring system 1, 24, 48 and 72 h post administration.

Very slight erythema (grade 1) was observed in 2/6 animals. The effect was observed in one animal at the 24 and 48 h observation time point and in one animal one hour after removal of the test substance only. All effects were fully reversible within 72 h after administration. There were no skin reactions in any other animal. Based on the study results, the test item is not considered irritating to the skin.

 

Additionally, a study on phototoxicity was performed in guinea pigs at concentrations of 20, 10 and 5% in vehicle, in which the ability of the test substance to induce an increased reactivity of the skin to irradiation with UV and/or visible light was investigated. Positive reactions in the form of slight to moderate erythema, mostly pronounced in the highest treatment groups, were observed on all treated spots in an unequivocal concentration-dependent manner in an incidence of 100, 100 and 40 % in the three treated groups. Incidence in the positive control group was 80 %. No other signs of intoxication were observed. Therefore, the test substance has to be regarded as phototoxic.

 

Eye irritation

The eye irritating potential of the test item was analysed in New Zealand White rabbits according to OECD guideline 405 and compliant with GLP. A single tests item dose of 100 mg was installed into one eye of each animal, and washed out after 24 h. The other eye remained untreated and served as control. Local irritations of the eye were evaluated according to Draize criteria 1, 24, 48 and 72 h after administration.

The test substance caused effects in the eyes in the form of conjunctival reddening, chemosis and discharge, inflammation and opacity of the iris, ranging in degree from slight to moderate. All signs of irritation were completely reversible within 72 h. Based on these results and under the experimental conditions of the study, the test item was considered not irritating tot the eye.

Justification for classification or non-classification

The available data on skin and eye irritation/corrosion do not meet the criteria for classification according to Regulation (EC) No. 1272/2008 and are therefore conclusive but not sufficient for classification. The test substance has to be regarded as phototoxic.