Flucytosine

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2005-05-17 to 2005-07-27

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

adopted 17 July 1992

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Version / remarks:

Commission Directive 96/54/EC

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

According to Annex XI section 1.1.2. of Regulation (EC) No 1907/2006 (REACH) testing of skin sensitisation in an in vivo LLNA is not necessary if there are data that is generated by studies not carried out according to GLP or the test methods reffered to in Article 13 (3) of the same regulation, if the data is sufficcient for the purpose of classification and labelling and/or risk assessment and if the key parameters foreseen to be investigated in the recommended methods are reliably covered, the exposure duration is similar to that of the recommended methods and if the study is adequate and reliable documented. The present GPMT was conducted according to OECD guideline 406, which is reliable without restrictions and adequate for classification and labelling purposes. It is well documented, measures the recommended key parameters and exposure duration is comparable to an in vivo LLNA. Thus, due to the provided reliable and adequate data the perfomance of an in vivo LLNA is not necessary.

Test material

Test material information

Constituent 1

Reference substance name:

Flucytosine

EC Number:

217-968-7

EC Name:

Flucytosine

Cas Number:

2022-85-7

Molecular formula:

C4H4FN3O

IUPAC Name:

flucytosine

Test material form:

solid: particulate/powder

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 300-450 g
- Housing: The animals were housed singly or in pairs in solid-floor polypropylene cages furnished with woodflakes.
- Diet (e.g. ad libitum): ad libitum, (Certified Guinea Pig Diet (Code 5026) supplied by BCM IPS Limited, London, UK)
- Water (e.g. ad libitum): ad libitum, mains tap water
- Acclimation period: At least 5 days
- Indication of any skin lesions: not reported

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 17-23
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

water

Concentration / amount:

5% (w/w)

Day(s)/duration:

single injection one week before the epicutaneous application

Adequacy of induction:

highest concentration used causing mild-to-moderate skin irritation and well-tolerated systemically

Induction 2

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

50% (w/w)

Day(s)/duration:

48 h

Adequacy of induction:

highest technically applicable concentration used

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

50 and 25 % (w/w)

Day(s)/duration:

24 h

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

10 in the main test and 5 control animals

Details on study design:

RANGE FINDING TESTS: Groups of guinea pigs were treated with various concentrations of test material, either intradermal or epicutaneously.
For intradermal induction 0.1 mL /injection site of a test solution with 5% (w/w) test substance was applied to one animal. Reading intervals were 24, 48, 72 h and 7 days after injection. The concentration caused only mild to moderate skin irritation, was well tolerated systemically, and was selected for the intradermal induction stage of the main test. For epicutaneous application two guinea pigs (intradermally injected with Freund's Complete Adjuvant ten days earlier) were treated with preparations of the test material (50%, 25%, 10% and 5% w/w in distilled water) under occlusive conditions for 48 h. The degree of erythema and oedema was evaluated approximately 1, 24 and 48 hours after dressing removal. The highest concentration producing only mild to moderate dermal initation was selected for the topical induction stage of the main test. Selection of the challenge concentration was performed by application of 4 concentrations of the test material (50, 25, 10 and 5% (w/w) in distilled water to the clipped flanks of two guinea pigs. The degree of erythema and oedema was evaluated approximately 1, 24 and 48 hours after dressing removal. The highest non-irritant concentration of the test material and one lower concentration were selected for the topical challenge stage of the main test.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: 2
- Exposure period: 7 days
- Test groups: test material in FCA
- Control group: FCA only
- Site: shoulder region
- Frequency of applications: single injection followed by a topical 48 h exposure after 7 days
- Concentrations: Intradremal injection with 0.1 mL (5% (w/w) of the test substance, after 7 days epicutaneous application of 50 % (w/w) of the test substance for 48 h

B. CHALLENGE EXPOSURE
- No. of exposures: 1
- Day(s) of challenge: 21
- Exposure period:
- Test groups: 50 and 25 % of the test material in distilled water
- Control group: 50 and 25 % of the test material in distilled water
- Site: shoulder region
- Concentrations: 50 and 25 % (w/w) test substance
- Evaluation (hr after challenge): 24 and 48 h

Positive control substance(s):

yes

Remarks:

2-Mercaptobenzothiazole

Results and discussion

Positive control results:

The results of the historical positive controls were depicted in table 1. 2-Mercaptobezothiazole produced a sensitisation 80 and 90 % of the male and female guinea pigs.

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

50% (w/w)

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

50 % (w/w)

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 3

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

25% (w/w)

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Results 4

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

25% (w/w)

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

none

Remarks on result:

no indication of skin sensitisation

Any other information on results incl. tables

Table 1. Summary of positive control Data for the Magnusson and Kligman Maximisation Study

Number of Animals and Sex		Positive Control Material	Concentration			Incidence of Sensitisation
Test	Control		Induction		Challenge
			Intradermal	Topical
10 Male	5 Male	2-Mercaptobenzothiazole	5% in arachis oil BP	50% in acetone/PEG 400 (70:30)	50 and25% in acetone/PEG 400 (70:30)	80% (8/10)
10 Male	5 Male	2-Mercaptobenzothiazole	5% in arachis oil BP	50% in acetone/PEG 400	50 and25% in acetone/PEG 400 (70:30)	80% (8/10)
10 Female	5 Female	2-Mercaptobenzothiazole	5% in arachis oil BP	50% in acetone/PEG 400 (70:30)	50 and25% in acetone/PEG 400 (70:30)	90% (9/10)

Table 2: Intradermal Induction – Individual Skin Reactions

Group	Animal Number	Grade of Erythema at Observation Site
		24 h		48 h
		Left site	Right site	Left site	Right site
Test	1	1	1	0	0
	2	2	2	1	1
	3	1	1	0	1
	4	1	1	1	1
	5	1	1	1	1
	6	1	1	0	0
	7	1	1	1	0
	8	1	1	1	1
	9	1	1	1	1
	10	1	1	1	1
Control	11	0	0	0	0
	12	0	0	0	0
	13	0	0	0	0
	14	0	0	0	0
	15	0	0	0	0

Table 3: Topical Induction – Individual Skin Reaction

Group	Animal Number	Skin reactions (Hours After Removal of Dressing)
		1 h			24 h
		Er	Oe	Other	Er	Oe	Other
Test	1	1	0	Bs	1	0	-
	2	1	0	-	1	0	-
	3	1	0	Bs	0	0	-
	4	1	0	Bs	1	0	-
	5	1	0	Bs	1	0	-
	6	1	0	Bs	0	0	-
	7	1	0	-	0	0	-
	8	1	0	-	0	0	-
	9	1	0	Bs	1	0	-
	10	1	0	-	0	0	-
Control	11	1	0	Bs	0	0	-
	12	1	0	-	0	0	-
	13	1	0	-	0	0	-
	14	1	0	-	0	0	-
	15	1	0	-	0	0	-

Bs = Bleeding from intradermal induction site

- = No other reactions noted

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In the present study conducted according to OECD guideline 406 (adopted 17 July 1992) 10 male guinea pigs experienced intradermal injections and epicutaneous applications of the test substance as induction followed by an epicutaneous challenge 14 days after the last induction. After challenge no signs of sensitisation were detected, thus the test item is considered to be no sensitizer under the conditions of the test. In accordance with Regulation (EC) No 1272/2008 (CLP) and the Globally Harmonized System for Classification and Labelling of Chemicals (GHS) the substance does not need to be classified and labeled as a sensitizer.