Prometryn

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

11 Dec 1996 to 11 Jan 1997

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1997

Report date:

1997

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EPA OPP 81-6 (Skin Sensitisation)

GLP compliance:

yes

Type of study:

Buehler test

Justification for non-LLNA method:

Currently no LLNA study is available for assessment. The Buehler test has been carried out as an animal test to predict human sensitization for over a decade and is recommended by international test guidelines such as OECD.

Test material

Test material information

Constituent 1

Reference substance name:

Prometryn

EC Number:

230-711-3

EC Name:

Prometryn

Cas Number:

7287-19-6

Molecular formula:

C10H19N5S

IUPAC Name:

6-(methylsulfanyl)-N2,N4-di(propan-2-yl)-1,3,5-triazine-2,4-diamine

Test material form:

solid: particulate/powder

In vivo test system

Test animals

Species:

guinea pig

Strain:

Hartley

Remarks:

Albino

Sex:

male/female

Details on test animals and environmental conditions:

TEST ANIMALS
- Females nulliparous and non-pregnant: not specified
- Weight at study initiation: males 436-548 g, females 418-488 g
- Housing: Suspended, wire bottom, stainless steel; 1-4 per cages (males separate from females)
- Diet: Guinea Pig Diet, ad libitum
- Water: Municipal water supply in water bls, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°F): 72 ± 5 (22.2 ± 2.5 ºC)
- Humidity (%): 30 - 80
- Air changes (per hr): 10 - 12
- Photoperiod (hrs dark / hrs light): 12/12

- IN-LIFE DATES: From: 11 Dec 1996 To: 11 Jan 1997

Study design: in vivo (non-LLNA)

Induction

Route:

epicutaneous, occlusive

Vehicle:

unchanged (no vehicle)

Concentration / amount:

400 mg moistened with 0.4 mL of 80% ethanol

Day(s)/duration:

days 1, 8 and 14 / 6 hours

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

unchanged (no vehicle)

Concentration / amount:

400 mg moistened with 0.4 mL of acetone

Day(s)/duration:

day 29 / 6 hours

No. of animals per dose:

- Irritation screening: 2 males and 2 females
- Group I (naive control): 5 males and 5 females
- Group II (test group): 5 males and 5 females

Details on study design:

IRRITATION SCREENING
Two male and two female albino guinea pigs were selected for irritation screening to determine both the maximum dose producing no more than slight irritation, and the maximum non-irritating dose. Tested in the screening were 400 mg of test substance moistened with 80% ethanol, and 75%, 50% and 25 % w/v concentrations of the test substance in 80% ethanol, with each animal receiving 0.4 mL of each concentration at different test sites.

PREPARATION OF ANIMALS
Five males and five females were selected for each of two treatment groups. Group I animals served as a naive control group and Group II animals were designated as the test group. On the day prior to each treatment, the animals were prepared by clipping the back of the trunk free of hair to expose a longitudinal area at least 8 x 10 cm on each animal. Individual body weights were recorded on Days 0 and 28.

TEST SUBSTANCE PREPARATION AND ADMINISTRATION
Based on the results of the irritation screening, the test substance was administered by application of 400 mg of test substance moistened with 0.4 mL of 80% ethanol. The test substance was not wettable with deionized water. For each induction treatment, Group II animals were treated by introducing the test substance beneath a 3.8 x 5 cm patch (a 1.6 x 2.8 cm gauze pad secured to a 3.8 x 5 cm piece of adhesive) known as a Coverlet adhesive dressing. Each adhesive coverlet patch was placed laterally from the midline of the back on the left front quadrant of the exposure area with the edge of the gauze pad adjacent to, but not overlapping the midline of the back. A strip of clear polyethylene film was placed over the patch and taped in place to secure the patch. Each animal was then placed in a restrainer for approximately six hours. At the end of the exposure period, the animals were removed from the restrainers, the wrappings and patches were removed, and the animals were returned to their cages. Group II animals were treated once weekly for three weeks with 400 mg of test substance moistened with 0.4 mL of 80 % ethanol. Induction treatments were on Days 1, 8 and 14. The same treatment regimen and test site location was used for all three induction treatments. Group I animals remained untreated during the induction phase of the study.

CHALLENGE TREATMENT
After a two week rest period, all animals (Groups I and II) were each challenged at a virgin test site with an application of 400 mg of test substance moistened with 0.4 mL of acetone. The challenge treatment was on Day 29. The dose was applied in a manner identical to the induction treatments, except the test site was placed laterally on the right rear quadrant of the exposure area with the edge of the gauze pad adjacent to the midline of the back.

OBSERVATIONS AND SCORING METHOD
Observations for skin reactions at each test site were made approximately 24 hours after each treatment. In addition, observations for skin reactions were made approximately 48 hours after the first induction treatment and 48 hours after the challenge treatment. The scoring scale used for grading skin reactions is presented in Table 1 (See ‘Any other information on materials and methods incl. tables’). An average score for each time period was obtained by adding all of the scores for each time period and dividing by the number of test sites scored for that time period. The test substance is considered a sensitizer if the mean irritation scores, the total number of animals with scores, and/or the total number of scores for the virgin test site in the test group after the challenge treatment are appreciably greater than those for the naive challenge group.

Challenge controls:

10 naive (previously untreated) control animals

Positive control substance(s):

yes

Remarks:

1-chloro-2,4-dinitrobenzene

Results and discussion

Positive control results:

A mean score of 1.2 for the test group after challenge treatment, when compared with the naive control group mean challenge score of 0.0, confirmed the sensitivity of guinea pigs to the positive control material.

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

400 mg moistened with 0.4 mL acetone

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:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

400 mg moistened with 0.4 mL acetone

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:

24

Group:

test chemical

Dose level:

400 mg moistened with 0.4 mL acetone

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

None

Remarks on result:

no indication of skin sensitisation

Results 4

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

400 mg moistened with 0.4 mL acetone

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

None

Remarks on result:

no indication of skin sensitisation

Results 5

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

0.1% w/v in acetone

No. with + reactions:

10

Total no. in group:

10

Clinical observations:

Faint to moderate erythema in all ten animals

Remarks on result:

positive indication of skin sensitisation

Results 6

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

0.1% w/v in acetone

No. with + reactions:

10

Total no. in group:

10

Clinical observations:

Faint to moderate erythema in all ten animals

Remarks on result:

positive indication of skin sensitisation

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the study (EPA 81-6, GLP), the test substance produced no irritation in animals of the naive control group (Group I) after the single treatment at challenge. The test substance likewise produced no irritation in animals of the test group (Group II) after the challenge treatment and therefore did not elicit a sensitizing reaction in guinea pigs.

Executive summary:

A skin sensitization study was conducted according to GLP and EPA guideline 81-6, on 10 male and 10 female short-haired albino guinea pigs to determine if the test substance produced a sensitizing reaction. Five males and five females were assigned to each of two groups, designated Groups I and II. Group I animals remained untreated during the induction phase of the study and served as a naive control group. Group II animals were treated with 400 mg of test substance moistened with 0.4 mL of 80 % ethanol (selected from previous screening). The animals were treated once weekly for three weeks, for a total of three treatments. After a two week rest period, all animals (Groups I and II) were challenged at a virgin test site with an application of 400 mg of test substance moistened with 0.4 mL of acetone.

The test substance produced no irritation in animals of the naive control group (Group I) after the single treatment at challenge. The test substance likewise produced no irritation in animals of the test group (Group II) after the challenge and therefore did not elicit a sensitizing reaction in guinea pigs.