Bordeaux mixture

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: Study conducted in compliance with Good laboratory Practice and internationally accepted guidelines.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

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

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.2600 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

An LLNA study was not conducted because adequate data from a guine-pig maximisation test are already available. 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:

calcium tetracopper(2+) dioxocopperbis(olate) hexahydroxide sulfate

EC Number:

910-853-9

Cas Number:

8011-63-0

Molecular formula:

Ca3Cu4H6O22S4.nH2O where n = 1 to 6

IUPAC Name:

calcium tetracopper(2+) dioxocopperbis(olate) hexahydroxide sulfate

Details on test material:

- Name of test material (as cited in study report): Bordeaux Mixture
- Composition of test material, percentage of components: 26.35% w/w as copper
- Lot/batch No.: 44

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male/female

Details on test animals and environmental conditions:

Adult Dunkin-Hartley guinea pigs, weighing 374 + 17 g (males) and 360 + 21 g (females) at initiation were used for the main test.

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

other: 0.9% sodium chloride solution

Induction 2

Concentration / amount:

See Details on Study Design.

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

other: 0.9% sodium chloride solution

Challenge 2

Concentration / amount:

See Details on Study Design.

No. of animals per dose:

See Details on Study Design.

Details on study design:

RANGE FINDING TESTS:

An initial irritation screening test was performed to determine the highest non-irritant concentration for the challenge phase of the study and an irritant concentration for the induction phase. A concentration of 0.1% w/w in the vehicle produced moderate irritation by injection and was selected for the intradermal induction phase. By the topical route, 30% w/w in the vehicle produced no irritation (the highest non-irritant concentration) and was selected for the topical induction phase and challenge.

MAIN STUDY:

A. INDUCTION EXPOSURE

Intradermal injections of test substance (0.1% w/w) in 0.9% sodium chloride, FCA at 50% v/v dilution with 0.9% sodium chloride and test substance (0.1% w/w) in FCA/0.9% sodium chloride (50:50 v/v) were administered to the interscapular region of 20 animals (10 male, 10 female). Ten control animals (five males, five females) received FCA at 50% v/v dilution with 0.9% sodium chloride, 0.9% sodium chloride and 0.9% sodium chloride at 50% w/w in FCA/0.9% sodium chloride (50:50 v/v). After six days, the interscapular region of the animals was clipped and treated with 0.5 mL of sodium lauryl sulphate at a concentration of 10% w/w in vaseline to induce local irritation. The next day, the topical induction phase was performed on the test animals: a filter paper pad (8 cm2 in area) was fully loaded with a 30% concentration of the test substance and applied to the shaved interscapular area previously injected and held in place for 48 hours with an occlusive dressing. Control animals received a filter paper pad fully loaded with 0.9% sodium chloride for the same period.

B. CHALLENGE EXPOSURE

21 days after initiation, the filter paper pad of a ‘Finn Chamber’ was fully loaded with the challenge dose of 30% w/w test substance in vehicle and applied to a naïve shaved site on test and control animals and held in place for 24 hours with an occlusive dressing. The challenge patch was removed and the animals were examined for erythema as indication of a sensitisation response 24 and 48 hours after removal of the dressings. Results were compared with earlier controls run at the same laboratory with mercaptobenzothiazole.

Results and discussion

Positive control results:

Positive reactions were recorded with mercaptobenzothiazole consistent with its known sensitising potential.

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

30% w/w

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: negative control. Dose level: 30% w/w. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: None.

Results 2

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

30% w/w

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

None

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: 30% w/w. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: None.

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

30% w/w

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

None

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 30% w/w. No with. + reactions: 0.0. Total no. in groups: 20.0. Clinical observations: None.

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

30% w/w

No. with + reactions:

4

Total no. in group:

20

Clinical observations:

Discrete or moderate erythema (score 1 or 2)

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 30% w/w. No with. + reactions: 4.0. Total no. in groups: 20.0. Clinical observations: Discrete or moderate erythema (score 1 or 2).

Any other information on results incl. tables

No skin reaction was recorded in the control animals 24 hours or 48 hours after challenge or in the test animals 24 hours after challenge. 48 hours after challenge, a discrete or moderate erythema (score 1 or 2) was recorded in four of the 20 test animals. See Table 1.

Table 1. Skin Sensitisation Challenge Response

Group	Topical induction treatment	Incidence of significant response		Total responders
		24 hours	48 hours
Control	vehicle (sodium chloride)	0/10	0/10	0/10
	Bordeaux Mixture (30% w/w)	0/10	0/10	0/10
Treated	vehicle (sodium chloride)	0/20	0/20	0/20
	Bordeaux Mixture (30% w/w)	0/20	4/20	4/20

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Bordeaux Mixture induced skin sensitisation in 20% of animals in the maximisation test i.e. less than the 30% threshold.
Classification according to Directive 67/548/EEC: Not classified.
Classification according to CLP/GHS: Not classified.

Executive summary:

A GLP-compliant guinea-pig maximisation test was carried out in accordance with the requirements of OECD 406, EU B.6 and US EPA OPPTS 870.2600 without significant deviation. An initial irritation screening test was performed to determine the highest non-irritant Bordeaux Mixture concentration for the challenge phase of the study and an irritant concentration for the induction phase. A concentration of 0.1% w/w in the vehicle (0.9% sodium chloride solution) produced moderate irritation by injection and was selected for the intradermal induction phase. By the topical route, 30% w/w in the vehicle produced no irritation (the highest non-irritant concentration) and was selected for the topical induction phase and challenge. Adult Dunkin-Hartley guinea pigs, weighing 374 + 17 g (males) and 360 + 21 g (females) at initiation were used for the main test. Intradermal injections of test substance (0.1% w/w) in 0.9% sodium chloride, FCA at 50% v/v dilution with 0.9% sodium chloride and test substance (0.1% w/w) in FCA/0.9% sodium chloride (50:50 v/v) were administered to the interscapular region of 20 animals (10 male, 10 female). Ten control animals (five males, five females) received FCA at 50% v/v dilution with 0.9% sodium chloride, 0.9% sodium chloride and 0.9% sodium chloride at 50% w/w in FCA/0.9% sodium chloride (50:50 v/v). After six days, the interscapular region of the animals was clipped and treated with 0.5 mL of sodium lauryl sulphate at a concentration of 10% w/w in vaseline to induce local irritation. The next day, the topical induction phase was performed on the test animals: a filter paper pad (8 cm²in area) was fully loaded with a 30% concentration of the test substance and applied to the shaved interscapular area previously injected and held in place for 48 hours with an occlusive dressing. Control animals received a filter paper pad fully loaded with 0.9% sodium chloride for the same period. 21 days after initiation, the filter paper pad of a ‘Finn Chamber’ was fully loaded with the challenge dose of 30% w/w test substance in vehicle and applied to a naïve shaved site on test and control animals and held in place for 24 hours with an occlusive dressing. The challenge patch was removed and the animals were examined for erythema as indication of a sensitisation response 24 and 48 hours after removal of the dressings. Results were compared with earlier controls run at the same laboratory with mercaptobenzothiazole.

No skin reaction was recorded in the control animals 24 hours or 48 hours after challenge or in the test animals 24 hours after challenge. 48 hours after challenge, a discrete or moderate erythema (score 1 or 2) was recorded in four of the 20 test animals. As skin sensitisation was induced in only 20% of animals in the maximisation test, it is concluded that Bordeaux Mixture is not classified as a skin sensitizer.