2-chloro-N,N-dimethyl-3-oxobutyramide

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: OECD Guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1996

Report date:

1996

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Test material

Test material information

Constituent 1

Reference substance name:

2-chloro-N,N-dimethyl-3-oxobutyramide

EC Number:

227-372-9

EC Name:

2-chloro-N,N-dimethyl-3-oxobutyramide

Cas Number:

5810-11-7

Molecular formula:

C6H10ClNO2

IUPAC Name:

2-chloro-N,N-dimethyl-3-oxobutanamide

Details on test material:

Name: Dimetbyl- 2-ch1oroacetoacetamide
Batch No.: 502001
Physical state: clear yellowish liquid
Density: 1.20 g/m3
Storage: ambient, protected from light

In vivo test system

Test animals

Species:

guinea pig

Strain:

other: Pirbright white Hsd/Poc: DH

Sex:

male/female

Details on test animals and environmental conditions:

Species: guinea pig
Strain: Pirbright white
Substrain: Hsd/Poc: DH
Source: Harlan-Winkelmann GmbH, Postfach 11 61, Gartenstr.27; D-33178 Borchen
Date of receipt: January 10, 1996 (range finding), January 17, 1996 (main test)
Acclimatization period: 7 days (range finding) 10 days (main test)
Animal selection: random
Animal identification: with colored markings; cage labelled with sex, date of study initiation, project no
Weight range (main test): male: 324 - 387 g, female: 303 - 369 g

Housing: collective housing up to a maximum of 10 animals per cage in a battery of cages, each equipped with a paper roll system
Illurnination: artificial lighting (120 lux) from 7.00 a.m. - 7.00 p.m.
Temperature: 22+/-3 °C
Relative humidity: 30 - 70 %
Measurement: twice daily

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

water

Induction 2

Concentration / amount:

5%

Challenge

Challenge 1

Route:

epicutaneous, semiocclusive

Vehicle:

water

Challenge 2

Concentration / amount:

5%

No. of animals per dose:

10 males/10 females

Details on study design:

The main study was performed on 20 test and 10 control animals. On the basis of the results of the range finding the concentration of 5 % of the test article was considered to be suitable for intradermal injection and the concentration of 100 % for dermal applica-tion.
First stage - an area of 4 x 6 cm over the shoulders was clipped short with electric clip-pers and cleaned with 70 % (v/v) ethanol. Three pairs of intradermal injections were then made symmetrically in two rows on either side of the spine:
Test group:
1. 0.1 ml FCA 50 % (w/w) diluted in aqua ad inject.
2. 0.1 ml test article diluted in aqua ad inject. (final concentration: 5 %)
3. 0.1 ml test article diluted in FCA/aqua ad inject. (final concentration: 5 %)
Control group:
1. 0.1 ml FCA 50 % (w/w) diluted in aqua ad inject.
2. 0.1 ml aqua ad inject.
3. 0.1 ml aqua ad inject. 50 % (w/w) diluted in FCA)
Second stage - 7 days after the intradermal injections, dermal application was initiated. Because the test article was non-irritating at the maximum concentration in the pilot study, the area was reclipped and pretreated with 10 % sodium lauryl sulfate (SLS) in vaseline 24 hours before application of the test article to induce a mild inflammation. The test article (at a concentration of 100 %) was spread in a thick layer [to saturation] over a 4 x 5 cm patch (filter paper). The latter was firmly secured over the previous in-jection sites by an occlusive dressing for 48 h. Control animals received a patch loaded with aqua ad inject.
Challenge procedure
Both control and test animals were subjected to a challenge exposure 14 days after the second stage of induction. The challenge test was performed on a 5 x 5 cm clipped area on each flank of the animals. The maximal non-irritating concentration of the test article (100 %) was applied to the left flank and aqua ad inject. to the right in a volume of 0.5 ml using the patch technique . In each case the duration of exposure was 24 h under an occlusive dressing. 24 and 48 h after patch removal, allergic responses were evaluated on a numerical scale according to Draize.
Rechallenge procedure
Although the contact sensitizing potential of the test article was assessed in the chal-lenge procedure, a rechallenge exposure was initiated 7 days later to confirm the results obtained. The rechallenge procedure was initiated because of the pre-terminal death of guinea pigs and the reaction in one control animal. In both control and test animals, two concentrations of the test article (50 and 25 % in aqua ad iniec!.) were applied to the right flank of each animal in a volume of 0.5 ml each, using the patch technique de-scribed above. Additionally a new control group was treated in the same manner. Ex-treme findings were observed in all guinea pigs of the main test with almost the same intensity of reactions in test and control animals. Slight primary skin reactions were also observed in animals of the new control group.
The severe findings after rechallenge in the animals of the main test confirmed the re-sults obtained after challenge.

Positive control substance(s):

yes

Remarks:

4-Aminobenzoic acid ethyl ester (benzocaine)

Results and discussion

Positive control results:

The reaction to the positive control substance is tested periodically. The last test with 4-Aminobenzoic acid ethyl ester (benzocaine) with an acceptable level of response to this substance was performed in November, 1995.

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

100%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 100%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: none.

Results 2

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

100%

No. with + reactions:

19

Total no. in group:

20

Clinical observations:

none

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 100%. No with. + reactions: 19.0. Total no. in groups: 20.0. Clinical observations: none.

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

The substance is sensitizing to the skin under the conditions used.

Executive summary:

The potential skin sensitizing properties of Dimetbyl-2-chloroacetoacetamide were assessed in the guinea pig maximization test according to OECD Guideline 406 using 20 test and 10 control animals. Following epicutaneous and intradermal induction exposure to the test article or the vehicle, the animals were subjected two weeks later to a challenge exposure with the test article. Allergic responses to the challenge procedure were evaluated 24 and 48 h after the end of the exposure period. 95% of the animals showed an allergic skin reaction.