Copper(II) carbonate-copper(II) hydroxide (1:1)

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: Already evaluated by the Competent Authorities for Biocides and Existing Substances Regulations.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2001

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

Test guideline 2

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:

The available study was conducted prior to the date on whch the LLNA became the method of choice.

Test material

Test material information

Constituent 1

Reference substance name:

Copper carbonate

EC Number:

214-671-4

EC Name:

Copper carbonate

Cas Number:

1184-64-1

Molecular formula:

CH2O3.Cu

IUPAC Name:

copper(2+) carbonate

Details on test material:

Substance: Copper carbonate
Lot/batch number: 26694/4/ROX
Description: Green powder
Purity: Not reported
Stability: Stable at room temperature.

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male

Details on test animals and environmental conditions:

Source: David Hall Limited, Burton-on-Trent, Staffordshire, UK.
Age/weight at the start of the study: At the start of the study the test animals weighed 300-357 g and were approximately 8-12 weeks old.

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

water

Remarks:

Distilled

Induction 2

Concentration / amount:

Intradermal induction: 0.1% w/w in distilled water (causing mild to moderate irritation)
Topical Induction: 50% w/w in distilled water (causing mild to moderate irritation)
Topical challenge: 5 % w/w and 2 % w/w in distilled water (usually maximum non-irritant concentration)

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

water

Remarks:

Distilled

Challenge 2

Concentration / amount:

Intradermal induction: 0.1% w/w in distilled water (causing mild to moderate irritation)
Topical Induction: 50% w/w in distilled water (causing mild to moderate irritation)
Topical challenge: 5 % w/w and 2 % w/w in distilled water (usually maximum non-irritant concentration)

No. of animals per dose:

10 test animals were used in the main study.

Details on study design:

State study type: Adjuvant
INDUCTION EXPOSURE: Days 0 – 7
On day 0, an area of 40 mm x 60 mm of hair was clipped from each animal using veterinary clippers and three pairs of 0.1 ml intradermal injections
were made on either side of the mid-line. The injections were:
a) Freund’s Complete Adjuvant plus distilled water at a ratio of 1:1
b) 0.1 % w/w formulation of the test material in distilled water
c) 0.1 % w/w formulation of the test material in a 1:1 preparation of Freund’s Complete Adjuvant plus distilled water.

Approximately 24 and 48 hours later, the degree of erythema at the test material injection sites (injection b) was evaluated. On day 7, the same area
was clipped again on each animal and treated with a topical application of test material (50 % w/w in distilled water) and held in place with occlusive
dressing for 48 hours. After 1 and 24 hours, the degree of erythema and oedema was evaluated after removal of the dressings.

Induction of the control animals was performed in an identical manner as for the test animals, except that the test material was ommitted.

The scoring schedule for erythema was derived from 'Modified OECD Test Guideline 406, 1992 and Method B6 Skin Sensitisation of Commission Directive 96/54/EEC' and the scoring schedule for oedema was taken from Draize, J.H. 1977. see Table 1.

CHALLENGE EXPOSURE: Day 21; see Table 1
On day 21, an area of 50 mm x 70 mm on both flanks was clipped free of hair and a filter paper patch loaded with test material at the maximum non-
irritant concentration (5 % w/w in distilled water) was applied to the right flank of each animal and held in place with surgical tape and an occlusive
dressing. To ensure the maximum non-irritant concentration was used at challenge, the test material was applied in a similar method to the left flank at a concentration of 2 % w/w in distilled water.

The dressings were kept in place for 24 hours, the dressing was then removed and the challenge sites swabbed with cotton wool soaked in distilled
water to remove residual material. After approximately 24 and 48 hours after challenge dressing removal, the degree of erythema and
oedema was evaluated. Any other reactions were also recorded. The scoring schedules used were the same as those outlined above for the
inductrion exposure stage (also see Table 1).

There was no rechallenge.

Challenge controls:

5 control animals were used in the main study.

Positive control substance(s):

yes

Remarks:

2-Mercaptobenzothiazole.

Results and discussion

Positive control results:

Six groups of 10 animals were tested the number f animals with signs of allergic reactions/number of animals in group gave an average of 9.5 animals per group.

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

2%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

See Table 1

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 2%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: See Table 1.

Results 2

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

2%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

See Table 1

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 2%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: See Table 1.

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

See Table 1

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: See Table 1.

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

5%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

See Table 1

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 5%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: See Table 1.

Any other information on results incl. tables

Pilot study

Intradermal induction sighting test:

The highest concentration causing only mild to moderate skin irritation which was well tolerated systemically (0.1% w/w) was selected for the intradermal induction stage of the main study.

Topical induction sighting test:

The highest concentration applied causing only mild to moderate dermal irritation which was well tolerated systemically (50% w/w) was selected for the topical induction stage of the main study.

Topical challenge sighting test: 

The highest non irritant concentration of the test material and one lower concentration were selected for the topical challenge stage of the main study (5% and 2% w/w in arachis oil BP).

Main study

After 24 hours: Number of animals with signs of allergic reactions / number of animals 0/10.

No test animals showed signs of erythema or oedema at either the 2 % or 5 % w/w challenge concentration. Green-coloured staining was noted at the challenge sites of all test and control animals.

After 48 hours: Number of animals with signs of allergic reactions / number of animals 0/10

No test animals showed signs of erythema or oedema at either the 2 % or 5 % w/w challenge concentration. Green-coloured staining was noted at the challenge sites of 5/10 test animals at the 2 % w/w challenge concentration and 2/10 animals at the 5 % w/w challenge concentration. Green-coloured staining was noted in 4/5 control animals at both the 2 % and 5 % w/w challenge concentrations. TABLE 1:

DETAILED INFORMATION INCLUDING INDUCTION/CHALLENGE/SCORING SCHEDULE FOR SENSITISATION TEST 

INDUCTION/ CHALLENGE	DAY OF TREATMENT	APPLICATION	OBSERVATIONS/REMARKS
Induction 1   Intradermal injection	0	3 intradermal injections made as follows; ·         FCA & distilled water in ratio 1:1 ·         0.1% w/w formulation of the test material in distilled water ·         0.1% formulation of the test material in a 1:1 preparation of FCA plus distilled water. Degree of erythema quantified at 24 and 48 hours following injections.	Moderate and confluent erythema was observed in all test animals at all time points except one animal at 48 hours which showed discrete or patchy erythema.   In the control animals, three animals at 24 hours showed discrete erythema and at 48 hours no signs of erythema were observed. No other signs of irritation were noted.
Pre-treatment for non irritating substance	There was no pre-treatment for non irritating substance
Induction 2   Topical induction	7	Filter paper with test material (50% w/w in distilled water) was applied to skin for 48 hours. Degree of erythema and oedema quantified 1 and 24 hours following removal of patch.	Green-coloured staining was noted at the challenge sites of all test animals. The staining did not affect evaluation of skin responses.   At 1-hour all test animals showed moderate and confluent erythema. At 24-hours test animals showed discrete/patchy erythema to moderate and confluent erythema. No signs of irritancy were noted in any of the controls.
Challenge   Topical challenge	21	Filter paper with 5 % w/w test material in distilled water was applied to each animal. To ensure a maximum non-irritant concentration was used at challenge, the test material was also applied at 2% w/w in distilled water.   The test material was removed after 24 hours. After 24 and 48 hours following challenge dressing removal, the degree of erythema and oedema was quantified.	Green-coloured staining was noted at the challenge sites of all test animals after 24 hours and in 5/10 test animals at the 2 % w/w challenge concentration and 2/10 animals at the 5 % w/w challenge concentration. All control animals showed green-coloured staining after 24 hours and4/5 control animals at both the 2 % and 5 % w/w challenge concentrations showed staining also. The staining did not affect evaluation of skin responses.   No skin reactions were noted at the challenge sites of the test or control animals at the 24 or 48-hour observations at the 2% and 5% w/w concentrations.

FCA – Freund’s Complete Adjuvant

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information

Conclusions:

The test material did not meet the criteria for classification as a sensitiser according to EU labelling regulations Commission Directive 93/21/EEC.

Executive summary:

Materials and methods

The study was performed to assess the contact sensitisation potential of copper carbonate in the albino guinea pig. Ten test and five control animals were used for the study. Two phases were involved; an induction of a response by intradermal injection and topical application, and a topical challenge of that response. Based on the results of sighting tests, the concentrations of the test material for the induction and challenge phases were selected as;
Intradermal induction: 0.1% w/w in distilled water
Topical Induction: 50% w/w in distilled water
Topical challenge: 2 and 5% w/w in distilled water

 

On day 0, approximately 24 and 48 hours after the initial intradermal induction injection (0.1% w/w), the degree of erythema was evaluated. Seven days later, the same area used for the intradermal injection was treated with a topical application of test material (50% w/w). The degree of erythema and oedema was evaluated 1 and 24 hours after removal of the patches. Induction of the control animals was performed in an identical manner as for the test animals, except that the test material was ommitted.

 

On day 21, test material was applied at the maximum non-irritant concentration (5% w/w) and a lower concentration (2% w/w) as challenge doses. Approximately 24 and 48 hours after removal of the challenge doses, the degree of erythema and oedema was evaluated and any other skin reactions were recorded.

 

The study was conducted according to Commission Directive 96/54/EC Method B6 Acute Toxicity (Skin Sensitisation) and OECD Guidelines for the Testing of Chemicals No. 406 'Skin Sensitisation' (adopted 17 July 1992). The study was also conducted according to GLP. No deviations from the test guidelines, or deficiencies in the method were reported.

 

Results and discussions

No skin reactions were noted at the challenge sites of the test or control animals at the 24 or 48-hour observations at the 2% and 5% w/w concentrations. Therefore, under the conditions of the test, the test material produced a 0% (0/10) sensitisation rate and was classified as a non-sensitiser to guinea pig skin.