Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 15 November to 17 December 1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
EU Method B.6 (Skin Sensitisation)
Deviations:
not specified
Guideline:
other: Magnusson B., Kligman A.M. (1970) Allergic Contact Dermatitis in the Guinea pig: Identification of contact allergens, Thomas C.C., Spriengfield, Illinois, U.S.A.
Deviations:
not specified
GLP compliance:
yes
Type of study:
guinea pig maximisation test

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Atrazine technical
Chemical name: 6-chloro-N2-ethyl-N4-isopropyl-1,3,5-triazine-2,4-diamine
Appaerance: white powder
Intended use: herbicide
Batch number: 2489
Purity: 96.9%

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
male
Details on test animals and environmental conditions:
The animals were in the weight range of 296 to 340 g on arrival and approximately four to five weeks of age. All the guinea-pigs were acclimatised to the experimental environment for twelve days prior to allocation to the main study.
The guinea-pigs were housed in groups of 5 in suspended metal cages with wire mesh floors in building R 17 Room 14.
A vitamin C enriched guinea-pigs diet FD2 and drinking water were provided ad libitum. hay was given weekly.
Animal room temperature was maintained at approximately 21°C relative humidity at 30-70%. These environmental parameters were recorded daily. air exchange was maintained at approximately 15 air changes per hour and lighting was controlled by means of a time switch to give 12 hours of artificial light (0700-1900 hours) in each 24 hours period.
Each animals was identified by ear tatoo number, each cage was identified by a coloured label .

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
intradermal and epicutaneous
Vehicle:
other: Alebicol D (a product of coconut oil, supplied by Alembic Products, Saltney, Chester, Englad)
Concentration / amount:
Based on the results of the preliminary investigations, the following concentrations of Atrazine a.i.were selected:
Induction intradermal injection: 10% w/v in Alebbicol D
Induction topical application: 50% w/v in Alebbicol D
Topical challenge: 50 and 25% w/v in Alebbicol D
Challengeopen allclose all
Route:
epicutaneous, occlusive
Vehicle:
other: Alebicol D (a product of coconut oil, supplied by Alembic Products, Saltney, Chester, Englad)
Concentration / amount:
Based on the results of the preliminary investigations, the following concentrations of Atrazine a.i.were selected:
Induction intradermal injection: 10% w/v in Alebbicol D
Induction topical application: 50% w/v in Alebbicol D
Topical challenge: 50 and 25% w/v in Alebbicol D
No. of animals per dose:
15 healthy male albino guinea-pigs of the Dunkin/hartley strain were obtained from D. Hall, Newchurch, Staffordshire, England.
The animals on the main study were allocated without conscious bias to 2 groups as follows:

Control animals: 5 animals
test animals: 10 animals

An additional six anomals, from the same supplier, were used for the preliminary investigations.
Details on study design:
The procedure of the main study may be considered in 2 parts, Induction and Challenge.

Induction intradermal injections-test animals
A 40 x 60 mm area of dorsal skin on the scapular region of the guinea-pig was clipped free of hair with electric clippers. Three pairs of intradermal injections were made into a 20 x40 mm area within the clipped area
Injectahles for the test animals were prepared as follows:
1.Freunds complete adjuvant was diluted with an equal volume of water for irrigation (Ph. Eur.)
2. Atrazine a. i., 10% w/v in Alembicol D
3. Atrazine a. i., 10% w/v in a 50 : 50 mixture of Freund s complete adjuvant and Alembicol D.

Induction topical application - test animals
The prel minary investigations indicated that the maximum practical concentration of the test substance for topical application (50%) did not produce skin irritation. Therefore, six days after the injections, the same 40 x 60 mm interscapular area was clipped and shaved free of hair and the site was pretreated hy gentle rubbing with 0.2 ml per site of 10% w/w sodium lauryl sulphate in petrolatum. Twenty-four hours later a 20 x 40 mm patch of Whatman No. 3 paper was saturated with approximately 0.4 ml of Atrazine a.i., 50% w/v in Alembicol D. The patch was placed on the skin of the test animals and covered by a length of impermeable plastic adhesive tape (50 mm width). This in turn was firmly secured by elastic adhesive bandage (50 mm width) wound round the torso of the animal and fixed with an impervious plastic adhesive tape. The dressing was left in place for 48 hours.

Induction control animals
During the induction phase the control animals were treated similarly to the test animals with the exception that the test substance was omitted from the intradermal injections and topical application.

Challenge control and test animals
The control and test animals were challenged topically two weeks after the topical induction application using Atrazine a.i. 50 and 25% w/v in Alembicol D.
Hair was removed by clipping and then shaving from an area on the left flank of each guinea-pig. A 20 x 20 mm patch of Whatman No. 3 paper was saturated with approximately 0.2 ml of Atrazine a. i. 50% w/v in Alembicol D and applied to an anterior site on the flank. Atrazine a. i. 25% w/v in Alembicol D was applied in a similar manner to a posterior site. The patches were sealed to the flank for 24 hours under strips of a plastic adhesive tape
covered by an elastic adhesive bandage wonnd round the trunk and secured with an impervious plastic adhesive tape.

Challenge controls:
see "Details on study design (traditional tests)"
Positive control substance(s):
yes
Remarks:
hexyl cinnamic aldehyde

Results and discussion

Positive control results:
The sensitivity of the guinea-pig strain used was checked periodically with hexyl cinnamic aldehyde, a known sensitiser.
Number of test animals: 10
Number of control animals: 5
Dose level (%):
Induction intradermal injection: Atrazine 10% w/v in Alembicol D
Induction topical application: Atrazine 50% w/v in Alembicol D
Challenge: Atrazine 50 and 25% w/v in Alembicol D.

Results:
Firs test:
Positive: 10/10 Inconclusive: 0/10 Negative: 0/10
Second test:
Positive: 9/10 Inconclusive: 0/10 Negative: 0/10
Third test:
Positive: 10/10 Inconclusive: 0/10 Negative: 0/10

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
positive response both for erythema and oedema
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D. No with. + reactions: 10.0. Total no. in groups: 10.0. Clinical observations: positive response both for erythema and oedema.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
4
Total no. in group:
10
Clinical observations:
Positive response both for erythema and oedema
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 4.0. Total no. in groups: 10.0. Clinical observations: Positive response both for erythema and oedema.
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
6
Total no. in group:
10
Clinical observations:
positive response only for erythema, no oedema formation
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 6.0. Total no. in groups: 10.0. Clinical observations: positive response only for erythema, no oedema formation .
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D
No. with + reactions:
10
Total no. in group:
10
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D. No with. + reactions: 10.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
9
Total no. in group:
10
Clinical observations:
Positive response both for erythema and oedema
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 9.0. Total no. in groups: 10.0. Clinical observations: Positive response both for erythema and oedema.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
1
Total no. in group:
10
Clinical observations:
positive response only for erythema, no oedema formation.
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 1.0. Total no. in groups: 10.0. Clinical observations: positive response only for erythema, no oedema formation..
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
test group
Dose level:
Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D
No. with + reactions:
10
Total no. in group:
10
Clinical observations:
Positive response both for erythema and oedema.
Remarks on result:
other: see Remark
Remarks:
Reading: other: 3rd reading. . Hours after challenge: 72.0. Group: test group. Dose level: Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D. No with. + reactions: 10.0. Total no. in groups: 10.0. Clinical observations: Positive response both for erythema and oedema..
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
test group
Dose level:
Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
9
Total no. in group:
10
Clinical observations:
positive response both for erythema and oedema
Remarks on result:
other: see Remark
Remarks:
Reading: other: 3rd reading. . Hours after challenge: 72.0. Group: test group. Dose level: Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 9.0. Total no. in groups: 10.0. Clinical observations: positive response both for erythema and oedema.
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
test group
Dose level:
Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
1
Total no. in group:
10
Clinical observations:
positive response only for erythema, no oedema formation.
Remarks on result:
other: see Remark
Remarks:
Reading: other: 3rd reading. . Hours after challenge: 72.0. Group: test group. Dose level: Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 1.0. Total no. in groups: 10.0. Clinical observations: positive response only for erythema, no oedema formation..
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D, Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: see Remark
Remarks:
Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D, Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D, Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: see Remark
Remarks:
Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D, Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
other: 3rd reading
Hours after challenge:
72
Group:
negative control
Dose level:
Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D, Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: see Remark
Remarks:
Reading: other: 3rd reading. . Hours after challenge: 72.0. Group: negative control. Dose level: Anterior site, exposed to atrazine a.i., 50% w/v in Alebicol D, Posterior site, exposed to atrazine a.i., 25% w/v in Alembicol D. No with. + reactions: 0.0. Total no. in groups: 10.0.

Any other information on results incl. tables

Dermal reactions in the test animals elicited hy the challenge application were compared with the findings simultaneously ohtained i a the control animals.

A test animal was considered to show positive evidence of delayed contact hypersensitivity if the observed dermal reaction at challenge was definitely more marked and/or persistent than the maximum reaction seen in animals of the control group.

If the dermal reaction seen in a test animal at challenge was slightly more marked and/or persistent than (but not clearly distinguishable from) the maximum reaction seen in control animals, the result for that test animal was classified as inconclusive.

A test animal was considered to show no evidence of delayed contact hypersensitivity 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 animals of the control group.

Applicant's summary and conclusion

Interpretation of results:
sensitising
Remarks:
Migrated information
Conclusions:
In this study, Atrazine a.i. produced evidence of skin sensitization (delayed contact hypersesitivity) in all ten test animals.