Registration Dossier

Administrative data

Endpoint:
skin sensitisation: in vivo (non-LLNA)
Type of information:
experimental study
Adequacy of study:
key study
Study period:
from 2000-01-17 into 2000-03-03
Reliability:
1 (reliable without restriction)

Data source

Reference
Reference Type:
other: Body responsible for the test
Title:
Unnamed
Year:
2000
Report Date:
2000

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: B.6 Skin sensitization OECD NR:406 Order of Ministry of the Health of Czech Republik No. 251/1998 Coll, B.6 Sensitization skin
GLP compliance:
yes
Type of study:
guinea pig maximisation test

Test material

Reference
Name:
Unnamed
Type:
Constituent

In vivo test system

Test animals

Species:
guinea pig
Strain:
Dunkin-Hartley
Sex:
female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Guinea pig white, female
- Housing: plastic cage, 2 animals per cage
- Diet (e.g. ad libitum): ad libidum standart food
- Water (e.g. ad libitum): ad libidum, ascorbic acid was added into water
- Acclimation period:1 week

ENVIRONMENTAL CONDITIONS
standart conditions
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Inductionopen allclose all
Route:
epicutaneous, semiocclusive
Vehicle:
olive oil
Concentration / amount:
Concentration of test material and vehicle used at induction:
1st injection: FCA in water 1:1
2nd injection: 1% sample in olive oil
3rd injection: 1% sample in FCA with water, 1:1
The sample was prepared in the mixture with the Freund
complete adjuvant (FCA). The olive oil was used as vehiculum
because the sample is almost insoluble in water.


Intradermal induction : zero day - test group

1st injection - FCA : water = 1:1

2nd injection - 1 % sample in olive oil

3rd injection - 1 % sample in FCA : water = 1:1


Intradermal induction : zero day - negative control group

1st injection - FCA : water = 1:1

2nd injection - olive oil

3rd injection - (50 % FCA : water = 1:1) : 50 % olive oil


Dermal induction : 7th day - test group

50 % sample in olive oil with fixation time : 48 hours


Dermal induction : 7th day - negative control group

olive oil with fixation time : 48 hours

Concentration of test material and vehicle used for each challenge:
20 % sample in olive oil.
Challengeopen allclose all
Route:
epicutaneous, semiocclusive
Vehicle:
olive oil
Concentration / amount:
Concentration of test material and vehicle used at induction:
1st injection: FCA in water 1:1
2nd injection: 1% sample in olive oil
3rd injection: 1% sample in FCA with water, 1:1
The sample was prepared in the mixture with the Freund
complete adjuvant (FCA). The olive oil was used as vehiculum
because the sample is almost insoluble in water.


Intradermal induction : zero day - test group

1st injection - FCA : water = 1:1

2nd injection - 1 % sample in olive oil

3rd injection - 1 % sample in FCA : water = 1:1


Intradermal induction : zero day - negative control group

1st injection - FCA : water = 1:1

2nd injection - olive oil

3rd injection - (50 % FCA : water = 1:1) : 50 % olive oil


Dermal induction : 7th day - test group

50 % sample in olive oil with fixation time : 48 hours


Dermal induction : 7th day - negative control group

olive oil with fixation time : 48 hours

Concentration of test material and vehicle used for each challenge:
20 % sample in olive oil.
No. of animals per dose:
Number of animals in test group: 20
Number of animals in negative control group: 10
Positive control substance(s):
yes
Remarks:
benzocain CAS 94-09-7

Results and discussion

Positive control results:
60% of positive skin reaction
Reliability of test was confirmed.

In vivo (non-LLNA)

Resultsopen allclose all
Reading:
1st reading
Hours after challenge:
24
Group:
test group
Dose level:
20 %
No. with + reactions:
7
Total no. in group:
20
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 20 %. No with. + reactions: 7.0. Total no. in groups: 20.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
test group
Dose level:
20 %
No. with + reactions:
7
Total no. in group:
20
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 20 %. No with. + reactions: 7.0. Total no. in groups: 20.0.
Reading:
1st reading
Hours after challenge:
24
Group:
negative control
Dose level:
%
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: %. No with. + reactions: 0.0. Total no. in groups: 10.0.
Reading:
2nd reading
Hours after challenge:
48
Group:
negative control
Dose level:
%
No. with + reactions:
0
Total no. in group:
10
Remarks on result:
other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: %. No with. + reactions: 0.0. Total no. in groups: 10.0.

Any other information on results incl. tables

Maximum concentration not causing irritating effects in preliminary test: 20 %

Signs of irritation during induction:
Test of primary irritation with different concentration of
the test sample was realised. The fixation time was 24
hours. One hour after removing bandage the sign of
irritation was observed. The highest concentration not
giving rise to irritating effects  applied epicutaneously
was 20% sample in olive oil.

The signs of irritation observation during induction have
not been executed.

Evidence of sensitisation of each challenge concentration:
1st Challenge 24 hour:

In 48 hours after application seven animals had the positive
skin reactions: two of the animals had the middle
repletion and the mild oedema, five of the animals had the
mild or the dappled repletion and the mild or the very
mildoedema.


In 72 hours after application the reaction was held over all
the previous reagents, the reaction was expressed as the
repletion, the oedema and the separating of skin surface
coats. For the animals numbered 28 and 30 the reaction was
held over the next 24 hours as the result of the allergic
dermatitis.

Other observations:
The sensitivity and reliability control results of the test:

Applied substance - Benzocain (CAS No.94-09-7)

Intradermal induction exposition: 5% conc.in olive oil.

Epicutan exposition: 25% concentration in vaseline alba.
Challange concentration: 5% concentration in vaseline alba.

Test results : 60% positive dermal reaction. The test
reliability was confirmed.

Applicant's summary and conclusion

Interpretation of results:
other: not classified