2(1H)-Pyridinone, 1-(acetyloxy)-

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

03 December 2012 to 01 August 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Fully GLP compliant and in accordance with current test guidelines

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 429 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.42 (Skin Sensitisation: Local Lymph Node Assay)

Deviations:

no

GLP compliance:

yes

Type of study:

mouse local lymph node assay (LLNA)

Test material

Test material information

Constituent 1

Reference substance name:

2-oxo-1,2-dihydropyridin-1-yl acetate

EC Number:

689-986-7

Cas Number:

1742-79-6

Molecular formula:

C7H7NO3

IUPAC Name:

2-oxo-1,2-dihydropyridin-1-yl acetate

Constituent 2

Reference substance name:

1742-78-6

IUPAC Name:

1742-78-6

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

Name: Oxypyrionacetate
CAS number: 1742-78-6
Batch number: 11085913
Quantity received: 52 g
Purity: 100%
Date of receipt: 20 November 2012
Storage details: stored in a refrigerator at 2 to 8ºC, in a sealed container in the dark.

In vivo test system

Test animals

Species:

mouse

Strain:

other: CBA/CaCrl

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River (UK) Ltd., Margate
- Age at study initiation: 9 to 10 weeks old
- Weight at study initiation: 18 to 20 g
- Housing: The animals were group housed during acclimatisation and individually housed from Day –1 in cages that conformed to the 'Code of Practice for the Housing and Care of Animals Used in Scientific Procedures' (Home Office, London, 1989).
- Diet (e.g. ad libitum): SQC(E) Rat and Mouse Maintenance Diet No 1, from Special Diets Services Ltd, Witham, UK was freely available to the animals at all times. Each batch of diet had been analyzed for specific constituents and contaminants by the manufacturer.
- Water (e.g. ad libitum): Mains water was provided, ad libitum, via cage-mounted water bottles. The water had been periodically analysed for specific contaminants.
- Acclimation period: 8 to 15 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 to 24°C, on two occassions the minimum dropped to 19°C
- Humidity (%): 45 to 65%, on two occassions the minimum dropped to 43%
- Air changes (per hr): The animal rooms were designed to permit 15 to 20 air changes per hour
- Photoperiod (hrs dark / hrs light): The rooms were illuminated by fluorescent strip-lights for twelve hours daily.

IN-LIFE DATES: From: 15 January 2013 To: 08 February 2013

Study design: in vivo (LLNA)

Vehicle:

dimethylformamide

Concentration:

Vehicle control: 0% test substance concentration
Test - low concentration: 10% test substance concentration
Test - Intermediate concentration: 25% test substance concentration
Test- High concentration: 50% test substance concentration

No. of animals per dose:

4 animals per dose

Details on study design:

Preliminary test
As no toxicological information was available regarding the systemic toxicity/irritancy potential of the test article, a preliminary screening test was performed with one mouse. The mouse was treated by daily application of 25 µL of the test article at the maximum suitable concentration (50% w/v in DMF) to the dorsal surface of each ear for three consecutive days (Days 1, 2 and 3).
The mouse was observed daily for six days from the initiation of treatment. Any signs of toxicity or irritation during this period were recorded. Ear thickness measurements were made using a thickness gauge on Day 1 (pre-treatment), Day 3 and Day 6.
Both ears were also observed for erythema.
Definitve test
The four groups of four female mice were subjected to application of the vehicle or one of the test formulations to the outer aspect of the auditory pinnae, once daily on Days 1, 2 and 3.
On Day 6 the mice were placed in a warming cabinet in order to dilate the peripheral blood vasculature and thus facilitate intravenous dosing. Each mouse was transferred to a cylindrical restrainer. A plastic syringe and fine gauge hypodermic needle were used to administer 0.25 mL phosphate buffered saline incorporating 20 µCi of 3HTdR into a tail vein of each mouse by slow bolus injection. After this treatment the mice were returned to their cages.
Approximately five hours after intravenous injection of the 3HTdR, all mice were killed by exposure to a rising concentration of carbon dioxide. Killing was organised to minimise the interval between death and the recovery of the auricular lymph nodes to no more than fifteen minutes.

Positive control substance(s):

hexyl cinnamic aldehyde (CAS No 101-86-0)

mercaptobenzothiazole (CAS No 149-30-4)

Statistics:

Not applicable

Results and discussion

In vivo (LLNA)

Results

Results 1

Parameter:

SI

Value:

32.9

Test group / Remarks:

10% w/v

Results 2

Parameter:

SI

Value:

39.7

Test group / Remarks:

25% w/v

Results 3

Parameter:

SI

Value:

29.8

Test group / Remarks:

50% w/v

Results 4

Parameter:

other: disintegrations per minute (DPM)

Remarks on result:

other: Scintilation fluid: 58 DPM Vehicle control: 793 DPM Test article, 10% w/v: 26054 DPM Test article, 25% w/v: 31447 DPM Test article, 50% w/v: 23598 DPM

Any other information on results incl. tables

Mortality

All animals survived treatment

Clinical signs

There were no clinical signs indicative of a systemic effect

The vehicle and test formulation application sites remained free of irritation

Body weights

There was no indication of a treatment related effect on body weight

Applicant's summary and conclusion

Interpretation of results:

Category 1 (skin sensitising) based on GHS criteria

Remarks:

Migrated information

Conclusions:

The Local Lymph Node Assay demonstrated that Oxypyrion Acetate has the potential to cause skin sensitisation.
The test article was classified as a Category 1 sensitiser according to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS).

Executive summary:

This study was conducted to asses the potential of the test article, Oxypyrion-Acetate, to cause skin sensitization in the mouse.

Following a preliminary screening test using a 50% w/v formulation, the test article was prepared for administration in dimethylformamide. Groups of four female CBA / CaCrl mice were subjected to topical applications of vehicle or one of the test formulations to the outer aspect of the auditory pinnae once daily on Days 1, 2 and 3. On Day 6 a 20 µCidose of tritiated ³H-methyl thymidine was injected intravenously into each mouse. Approximately five hours later the auricular lymph nodes were recovered from each animal. The nodes from mice subjected to the same treatment were pooled and suspensions of the cellular components of the lymph nodes were prepared in 5% w/v trichloroaceticacid and processed through a scintillation counter.

Test results are expressed in terms of Stimulation Indices, the ratios of the mean scintillation count per group obtained from the test groups relative to the corresponding mean scintillation count from controls. The threshold level for the Stimulation Index to be considered a positive indicator of the potential to cause skin sensitisation is 3.0.

	Concentration of test article in applied formulation (% w/v)
	10%	25%	50%
Stimulation Index	32.9	39.7	29.8

The Local Lymph Node Assay demonstrated thatOxypyrion-Acetate has the potential to cause skinsensitisation.

The test article was classified as a Category 1 sensitiser according to the Globally Harmonised System of Classification and Labelling of Chemicals (GHS).