3,5-dimethylpyrazole

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

16th Februrary to 22nd March 1994

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP conditions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Study is over 12 years old and precludes LLNA method

Test material

Test material information

Constituent 1

Reference substance name:

3,5-dimethylpyrazole

EC Number:

200-657-5

EC Name:

3,5-dimethylpyrazole

Cas Number:

67-51-6

Molecular formula:

C5H8N2

IUPAC Name:

3,5-dimethyl-1H-pyrazole

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

Appearance: White crystalline powder.
Storage conditions: Room temperature in the dark.
Date received: 11th February 1994.

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: D.Hall, Newchurch, Staffordshire, England.
- Age at study initiation: 6 - 7 weeks old.
- Weight at study initiation: 291 to 341g.
- Housing: In groups of 5 in suspended metal cages with wore mesh floors.
- Diet (e.g. ad libitum): Vitamin C enriched guinea-pig diet FDI, ad libitum. Hay provided weekly.
- Water (e.g. ad libitum): drinking water, ad libitum.
- Acclimation period: 6 days.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21ºC.
- Humidity (%): 30-70 %.
- Air changes (per hr): 15 charges per hour.
- Photoperiod (hrs dark / hrs light): 12 hours of artificial light (0700 - 1900) in each 24 hour period.

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

other: Alembicol D a product of coconut oil.

Induction 2

Concentration / amount:

Induction intradermal injection: 0.25 w/v in Alembicol D
Induction topical application: 60% w/v in Alembicol D
Challenge application: 60 and 30% w/v Alembicol D

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

other: Alembicol D a product of coconut oil.

Challenge 2

Concentration / amount:

Induction intradermal injection: 0.25 w/v in Alembicol D
Induction topical application: 60% w/v in Alembicol D
Challenge application: 60 and 30% w/v Alembicol D

No. of animals per dose:

10 test and 5 control animals.

Details on study design:

RANGE FINDING TESTS: A preliminary study was performed to identify a suitable concentration for the main test.

MAIN STUDY
A. INDUCTION EXPOSURE
- The concentration used was the maximum practical concentration.
- Site: An area of 40 x 60 mm on the dorsal was prepared by clipping the hair.

Intradermal Injection:
- No. of exposures: 3 intradermal injections per site.
- Area of site: 2 x 4 cm.
- Injection 1 = Freud's complete adjuvant, diluted with equal volume of water for irrigation.
- Injection 2 = 0.25% w/v of the test material in Alembicol D.
- Injection 3 = 0.25 w/v of the test material in a 50:50 mixture of Freud's complete adjuvant and Alembicol D.

Topical application:
- 6 days after the intradermal injections the area was prepared by clipping the hair and rubbing with 0.2 ml of 10% w/w sodium lauryl sulphate in petrolatum. 24 hours later a patch saturated in 0.4 ml of the test material, 60% w/v in Alembicol D, was attached to the test site. The patch was fixed with an impermeable adhesive tape.
- Exposure period: 48 hours.
- Area of site: 20 x 40 mm.

- Control group:
Animals were treated the same as in the induction phase, except for the test material was omitted.

B. CHALLENGE EXPOSURE
- 2 weeks after the topical induction both the control and the test group were exposed to the test material. The site was prepared by clipping the hair and a patch saturated in 0.2 ml of the test material was fixed at two sites, the anterior flank and posterior.
- Exposure period: 24 hours.
- Site area: 20 x 20 mm
- Concentration: 60% w/v at the anterior site and 30% w/v to the posterior site.
- Evaluation (hr after challenge): 24, 48 and 72 hours after the removal of the patches.

OBSERVATION:
- Clinical signs: all animals were observed daily.
- Bodyweight: Recorded on Day 1 and on the last day of the observation period.
- Dermal response: Scored according to the Draize scale (1997) which can be seen in table 1 in the field "any other information on materials and methods incl. tables".

INTERPRETATION OF RESULTS
- Positive reaction criteria: If the reaction at challenge was more marked and/or persistent than the maximum reaction seen in the control group, the result would be considered positive.
- Inconclusive reaction criteria: If the reaction was slightly more marked and/or persistent than (but not clearly distinguishable from) the maximum reaction seen in control animals, the result is considered inconclusive.
- Negative reaction criteria: If the dermal reaction resulting from the challenge application was the same as or less marked and/or persistent than the maximum reaction seen in the control animals, the result is considered negative.

Challenge controls:

Same as test challenge.

Positive control substance(s):

yes

Remarks:

Formalin

Results and discussion

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

72

Group:

test chemical

Dose level:

60% w/v anterior site

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 2

Key result

Reading:

1st reading

Hours after challenge:

72

Group:

test chemical

Dose level:

30% w/v posterior site

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 3

Key result

Reading:

1st reading

Hours after challenge:

72

Group:

negative control

Dose level:

60% w/v anterior site

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 4

Key result

Reading:

1st reading

Hours after challenge:

72

Group:

negative control

Dose level:

30% w/v posterior site

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Any other information on results incl. tables

Clinical Signs:

No signs of ill health or toxicity were recorded.

Bodyweight:

All animals showed the expected increase in bodyweight.

Intradermal Injections:

Necrosis was recorded at sites receiving Freund's Complete Adjuvant in the test and control animals.

Slight irritation was seen in test animals at sites receiving 0.25% w/v in Alembicol D and was also observed in control animals receiving Alembicol D.

Topical Application:

Very slight erythema was observed in test animals following topical application with 60% w/v in Alembicol D.

Very slight erythema was also seen in the control animals.

Challenge:

There was no dermal reaction seen in any of the test or control animals.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the test the test material was determined to be not sensitising.

Executive summary:

In a GLP compliant study which was performed according to the standardised guideline EU Method B.6, the potential for the test material to cause skin sensitisation was determined in a Guinea-pig maximisation test. Ten Guinea-pigs were exposed to the test material, none of which displayed a dermal reaction. No other signs of toxicity were observed in any animal.

Under the conditions of the test the test material is considered to be non-sensitising and therefore according to Regulation (EC) No. 1272/2008 no classification is required.