Ammonium bromide

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1997-08-04 to 1997-09-05

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

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:

EPA OPP 81-6 (Skin Sensitisation)

Deviations:

no

Test guideline 3

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 study was conducted prior to existance of OECD 429 (LLNA) testing guidelines

Test material

Test material information

Constituent 1

Reference substance name:

Ammonium bromide

EC Number:

235-183-8

EC Name:

Ammonium bromide

Cas Number:

12124-97-9

Molecular formula:

BrH4N

IUPAC Name:

Bromide activated chloramine (BAC) generated from ammonium bromide and sodium hypochlorite

Details on test material:

- Name of test material (as cited in study report): Ammonium Bromide
- Description: white powder
- Analytical purity: 98.5%
- Lot/batch No.: 970027/1
- Storage condition of test material: test material was stored in the dark under ambient conditions
- Other: requisite quantity of ammonium bromide was added to an appropriate amount of distilled water and mixed

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: David Hall Limited, Darley Oarks, Newchurch, Staffordshire, United Kingdom
- Age at study initiation: less than one year
- Weight at study initiation: 299-321 g
- Housing: in aluminium cages (48 x 61 x 25 cm) with a grid floor (dose-range finding: 2 per cage, main study: 5 per cage)
- Diet (e.g. ad libitum):
- Water (e.g. ad libitum):
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18 - 20°C
- Humidity (%): mean: 67%
- Air changes (per hr): 15-20
- Photoperiod (hrs dark / hrs light): 12 hour light/dark cycle

IN-LIFE DATES: From: 1997-08-04 To: 1997-09-05

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

water

Induction 2

Concentration / amount:

Induction:
5% test substance for intradermal injection,
55% test substance for topical application (maximum practicable concentration)

Challenge:
55% test substance for topical application (maximum practicable concentration)

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

water

Challenge 2

Concentration / amount:

Induction:
5% test substance for intradermal injection,
55% test substance for topical application (maximum practicable concentration)

Challenge:
55% test substance for topical application (maximum practicable concentration)

No. of animals per dose:

20 in test group
10 in control group

Details on study design:

RANGE FINDING TESTS:

Two animals each were used in the dose range finding phase for the selection of appropriate concentrations for intradermal and topical induction.

Dose range finding for induction:
Dose ranging for induction was conducted via intradermal injections and topical application of formulations of the test material. For the dose range finding study, hair was clipped from an area of the scapular region (intradermal injection of 1, 2, 5 or 10% ammonium bromide solution) or of both flanks (topical induction with 25, 40, 50 or 55% ammonium bromide solution) of the animals on the day prior to administration.

Dose ranging for challenge:
Concentrations used took account of the maximum practicable concentration for administration. Three weeks prior to administration of the test material, hair was clipped from an area across the scapular region of the animals. On the following day, each animal was treated via intradermal injection.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: Two
- Exposure period: day 0 intradermal injection, day 7 topical application
- Control group: 2-mercaptobenzothizole (MBT) as the positive control substance
- Site: scapula region
- Duration: Removal of test substance 48 hours after topical application with sterile distilled water.
- Concentrations: 5% test substance for intradermal injection, 55% test substance for topical application


B. CHALLENGE EXPOSURE
- No. of exposures: One
- Day(s) of challenge: day 21 topical application
- Exposure period: Removal of test substance 48 hours after topical application with sterile distilled water
- Site: scapula region
- Concentrations: 55% test substance
- Evaluation (hr after challenge): 24h, 48h after challenge; dermal reactions were also assessed 24h after intradermal and topical induction application.

OTHER:
- FCA was prepared as a 50% v/v emulsion with sterile distilled water

Positive control substance(s):

yes

Remarks:

2-mercaptobenzothizole (MBT)

Results and discussion

Positive control results:

For positive control, animals were treated with MBT at a dose level of 75% w/v in maize oil. Following challenge, 100% of test group animals and none of the control group animals reacted positively. This demonstrates the ability of the test method to identify a mild/moderate sensitizer.

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

5 % / 55 %

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

no clinical signs

Remarks on result:

no indication of skin sensitisation

Results 2

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

5 % / 55 %

No. with + reactions:

0

Total no. in group:

20

Clinical observations:

no clinical signs

Remarks on result:

no indication of skin sensitisation

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

100 %

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

no clinical signs

Remarks on result:

no indication of skin sensitisation

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

100 %

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

no clinical signs

Remarks on result:

no indication of skin sensitisation

Results 5

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

5 %/ 75 %

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

-

Remarks on result:

positive indication of skin sensitisation

Remarks:

Based on results of 6-monthly check

Results 6

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

5 %/ 75 %

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

-

Remarks on result:

positive indication of skin sensitisation

Any other information on results incl. tables

Result of pilot study:

Intradermal Injection:

For the selection of appropriate concentrations for induction, intense reactions were noted at a concentration of 10% ammonium bromide. Discrete to moderate reactions were noted at concentrations of 1, 2 and 5% ammonium bromide. No reactions were seen after challenge.

Topical Application:

No reactions were noted at concentrations up to 55% ammonium bromide both for the selection of inducation and challenge concentration. Animals pretreated with Sodium Lauryl Sulphate showed dry and/or flaky and scabbed test sites after challenge application.

Overall result (main study):

No positive skin responses were noted following induction or challenge and no skin sensitising potential could be identified.

Body weight performance was considered to have been satisfactory.

Two animals /1 test group and 1 control group), were humanely killed before challenge as the condition of their scapular region had exceeded the severity limit set by the project licence governing the study. There were no other clinical signs, other than skin reactions, noted.

There were no premature deaths noted.

Table A6.1.5/01-1     Result of skin sensitisation pretest with Ammonium Bromide
Application method	Concentration % (v/v)	Skin reaction after intracutaneous or epicutaneous application
		Animal no. 1			Animal no. 2
		24h	48h	72h	24h	48h	72h
intracutaneous	10	3	3	3	3	3	3
	5	2	2	2	1 1	1	0
	2	1	1	0	1	0	0
	1	0	0	0	1	0	0
epicutaneous		Animal no. 3			Animal no. 4
	55	0a	0a	n.d.	0	0	n.d.
	50	0	0	n.d.	0	0	n.d.
	40	0	0	n.d.	0	0	n.d.
	25	0	0	n.d.	0	0	n.d.

0= no visible change

1= discrete or patchy erythema

2= moderate and confluent erythema

3= intense erythema and swelling

a= scabbing on test site

n.d.= not determined

Table A6.1.5/01-2   Result of intradermal induction with Ammonium Bromide (main study)

 

Group and concentration (% ammonium bromide)	Animal number	Reaction Scores 24 hours after injection
Control (0)	1	0
	2	0
	3	0
	4	0
	5	0
	6	0
	7	0
	8	0
	9	0
	10	0
Test (5)	1	0
	2	0
	3	0
	4	0
	5	0
	6	0
	7	0
	8	0
	9	0
	10	0
	11	0
	12	0
	13	0
	14	0
	15	0
	16	0
	17	0
	18	0
	19	0
	20	0

Table A6.1.5/01-3     Result of topical induction with Ammonium Bromide (main study)

 

Group and concentration (% ammonium bromide)	Animal number	Reaction Scores 24 hours after injection
Control (0)	1	0
	2	0
	3	0
	4	0
	5	0
	6	0
	7	0
	8	0
	9	0
	10	0
Test (55)	1	0
	2	0
	3	0
	4	0
	5	0
	6	0
	7	0
	8	0
	9	0
	10	0
	11	0
	12	0
	13	0
	14	0
	15	0
	16	0
	17	0
	18	0
	19	0
	20	0

Table A6.1.5/01-4     Result of challenge with Ammonium Bromide (main study)

 

group	Animal number	Concentration ammonium bromide (%)/ Time after patch removal				Concentration (% ammonium bromide)/response		Concentration (% ammonium bromide)/ % positive
		55		0
		24h	48h	24h	48h	55	0	55	0
control	1	0	0	0	0	-	-	0	0
	2	0	0	0	0	-	-
	3	0	0	0	0	-	-
	4	0	0	0	0	-	-
	5	0	0	0	0	-	-
	6	0	0	0	0	-	-
	7	0	0	0	0	-	-
	8	0	0	0	0	-	-
	9	0	0	0	0	-	-
	10	a	a	a	a	a	a
test	1	0	0	0	0	-	-	0	0
	2	0	0	0	0	-	-
	3	0	0	0	0	-	-
	4	0	0	0	0	-	-
	5	0	0	0	0	-	-
	6	0	0	0	0	-	-
	7	0	0	0	0	-	-
	8	0	0	0	0	-	-
	9	0	0	0	0	-	-
	10	0	0	0	0	-	-
	11	0	0	0	0	-	-
	12	0	0	0	0	-	-
	13	0	0	0	0	-	-
	14	0	0	0	0	-	-
	15	a	a	a	a	a	a
	16	0	0	0	0	-	-
	17	0	0	0	0	-	-
	18	0	0	0	0	-	-
	19	0	0	0	0	-	-
	20	0	0	0	0	-	-

 

-             Negative

a             Animal dead before assessment

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Under the conditions of the study, ammonium bromide is considered not to be a sensitizer (0/20 sensitisation rate) in guinea pigs. In accordance with CLP Regulation (EC) No 1272/2008, no classification and labelling with respect to skin sensitisation is required.

Executive summary:

Materials and Methods

The objective of the study was to determine the delayed contact hypersensitivity of ammonium bromide using the protocol of the Magnusson-Kligman Maximisation Test in guinea pigs. The induction procedure for the test group of 20 guinea pigs consisted of exposure to the test material via two routes, intradermal injection (5% ammonium bromide) and topical application (55% ammonium bromide; the maximum practicable concentration). Animals were also exposed to an adjuvant via intradermal injection. The control group was exposed to vehicle, sterile distilled water, only. Prior to a topical challenge application with 55% ammonium bromide, all animals were treated with a 10% SLS solution the day before. The appropriate concentrations for intradermal injection and topical induction as well as for the topical challenge application were selected in a dose range finding study.

Results and Discussion

Prior to challenge, 2 animals (1 test group, 1 control group) were humanely killed as the condition of their scapular region had exceeded the severity limit set by the project licence governing the study. Following challenge no positive responses were observed in any of the remaining control or test group animals after challenge with 55% ammonium bromide.