Oxydiethylene bis(chloroformate)

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

migrated information: read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study conducted in accordance with GLP. Read-across to a structural similar substance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

Principles of method if other than guideline:

Maximization test according to Magnusson et Kligman.

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

The assay is representing an OECD TG validated test method for a challenge assay.

Test material

Test material information

Constituent 1

Reference substance name:

Cyclopentyl chloroformate

EC Number:

411-460-0

EC Name:

Cyclopentyl chloroformate

Cas Number:

50715-28-1

Molecular formula:

C6 H9 Cl O2

Constituent 2

Reference substance name:

Cyclopentylchlorformate

IUPAC Name:

Cyclopentylchlorformate

Details on test material:

- Name of test material (as cited in study report): Chloroformiate de cyclopentyle
- Supplier: SNPE (study sponsor)
- Physical state: liquid, colourless
- Analytical purity: 98.2%
- Purity test date: 24.06.1992
- Lot/batch No.: 26/92
- Stability under test conditions: assumed
- Storage condition of test material: at 4°C under N2

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male/female

Details on test animals and environmental conditions:

TEST ANIMALS
- Thirty animals were received (15/sex)
- Source: Centre d'élevage Lebeau, 78950 Gambais, France
- Age at study initiation: not specified
- Weight at study initiation: the mean body weight was 392±26 g and 403±24 g for males and females, respectively
- Housing: individually, in polycarbonate cages (48 x 27 x 20 cm)
- Diet (e.g. ad libitum): granulated "Cobayes entretien référence 106" feed, ad libitum
- Water (e.g. ad libitum): 0.22 µm filtered tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 3° C
- Humidity (%): 50 ± 20 %
- Air changes (per hr): filtered air, not recycled
- Photoperiod (hrs dark / hrs light): 12 hrs / 12 hrs

ANALYSES FOR CONTAMINATION OF BEDDING, FEED AND DRINKING WATER
Analyses of bedding for contaminants were regularly done (Laboratoire Wolff, 92110 Clichy, France).
Analyses data for feed contamination were provided by the feed supplier.
Analyses for drinking water contamination was regularly done (Laboratoire Municipal et Régional de Rouen, 76000 Rouen, France - Centre de Nutrition Humaine . 54000 Nancy, France - Laboratoire Départemental d'Analyses, 27000 Evreux, France).
All analyses indicated that bedding, feed and drinking water were suitable.

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

maize oil

Induction 2

Concentration / amount:

The concentrations used for testing were selected on the basis of the results of a pretest.
For the intradermal induction, 0.1 mL of 0.1% test material in vehicle was injected.
For the epicutaneous induction, 0 .5 mL of 15% test material in vehicle was applied to the skin.
For the epicutaneous challenge, 0.5 mL of 20% test material in vehicle was applied to the skin.

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

maize oil

Challenge 2

Concentration / amount:

The concentrations used for testing were selected on the basis of the results of a pretest.
For the intradermal induction, 0.1 mL of 0.1% test material in vehicle was injected.
For the epicutaneous induction, 0 .5 mL of 15% test material in vehicle was applied to the skin.
For the epicutaneous challenge, 0.5 mL of 20% test material in vehicle was applied to the skin.

No. of animals per dose:

The thirty guinea-pigs (15 males and 15 females) were allocated in the following 2 groups:
- control group 1 ( 5 males and 5 females )
- treated group 2 (10 males and 10 females)

Details on study design:

PRETEST
Referring to the intradermal administration, the pre-test revealed that 0.1% of test material resulted in skin irritation whereas 1% test material induced necrosis.
Referring to the epicutaneous application, the pre-test revealed that under occlusive test conditions for 24 hours, 20% test material induced no skin changes whereas test material applied as such, i.e. undiluted, resulted in early skin necrosis.
The test concentrations for the main study were selected on the basis of these findings.

MAIN TEST
INDUCTION
On the first day, 0.1 mL of 0.1% test material in vehicle was administered to the animals by intradermal injection, in presence of Freund's adjuvant.
After 8 days, 0 .5 mL of 15% test material in vehicle was applied to the skin of each animal under occlusive conditions. For details, see the table below.

CHALLENGE
Challenge was done after 12 days following induction.
Challenge consisted of an epicutaneous application of 0.5 mL vehicle on the left flank and 0.5 mL of 20% test material in vehicle on the right flank of each animal. Both application sites were covered with a dressing and maintained under occlusive conditions for 24 hours.
Cutaneous reactions on the challenge application sites were evaluated 24 and 48 hours after removal of the dressing.
For details, see the table below.

SCORING OF SKIN FINDINGS
For details see the table below.

CLINICAL EXAMINATION
The animals were examined twice a day for possible clinical symptoms and/or mortality.
The animals were weighed the day of random distribution in the test groups, at test initiation (day 1) and thereafter, on day 8, 15 and 25.

SACRIFICE
After the final scoring period, the animals were sacrificed and cutaneous samples were taken from the challenge application sites from all the animals for gross examination; no histological examination was performed.

CRITERIA FOR DETERMINATION OF SENSITIZATION POTENTIAL
The skin reaction of the animals was considered positive in case of well-defined effects or in case of questionable findings which need to be confirmed at necropsy to be due to sensitization.
The microscopical examination of dermal reaction due to sensitization will reveal basal spongiosis and reactional acanthosis of the epidermis as well as an infiltration of mononucleated in the dermis (according to Duprat et al., Investigations histopathologiques et cytologiques lors de la mise en évidence, chez le cobaye, d'une allergie cutanée de type retardé. Revue Méd. Vét. 127(7): 1083-1101,1976).
The determination of the degree of sensitization of the test material is calculated as ratio between animals with positive reaction and total number of animals (for details, see table below). According to the EU Directive 91/325/CEE, the results of a sensitization test are considered to be positive when at least 30% of the experimental animals exhibit skin reactions; furthermore, only the findings obtained 48 hours after application were taken into account for the determination of the sensitization rate.

Positive control substance(s):

not specified

Results and discussion

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

0.5 ml of 20% test material in vehicle

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

After challenge in the treated animals, moderate or severe erythema (scores of 3 or 4) was observed in all animals.

Remarks on result:

other: see Remark

Remarks:

Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 0.5 ml of 20% test material in vehicle. No with. + reactions: 20.0. Total no. in groups: 20.0. Clinical observations: After challenge in the treated animals, moderate or severe erythema (scores of 3 or 4) was observed in all animals..

Results 2

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

0.5 ml of 20% test material in vehicle

No. with + reactions:

20

Total no. in group:

20

Clinical observations:

Moderate or severe erythema (scores of 3 or 4) was observed in all animals. In 2/20 animals, signs of tissue destruction of the skin were noted after 48 hrs; in the remaining 18 animals dryness of the skin was reported.

Remarks on result:

other: see Remark

Remarks:

Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 0.5 ml of 20% test material in vehicle. No with. + reactions: 20.0. Total no. in groups: 20.0. Clinical observations: Moderate or severe erythema (scores of 3 or 4) was observed in all animals. In 2/20 animals, signs of tissue destruction of the skin were noted after 48 hrs; in the remaining 18 animals dryness of the skin was reported..

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

negative control

Dose level:

Vehicle only

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

In the control group, moderate signs of a primary cutaneous irritation were noted in 4/5 males (erythema scores of 1 or 2); no erythema was noted in females.

Remarks on result:

other: see Remark

Remarks:

Reading: 1st reading. . Hours after challenge: 24.0. Group: negative control. Dose level: Vehicle only. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: In the control group, moderate signs of a primary cutaneous irritation were noted in 4/5 males (erythema scores of 1 or 2); no erythema was noted in females..

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

negative control

Dose level:

Vehicle only

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

After 48 hours, dryness of the skin appeared in 8 out of 10 animals.

Remarks on result:

other: see Remark

Remarks:

Reading: 2nd reading. . Hours after challenge: 48.0. Group: negative control. Dose level: Vehicle only. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: After 48 hours, dryness of the skin appeared in 8 out of 10 animals..

Any other information on results incl. tables

No clinical signs and no mortalities were observed during the study. The body weight gain of the treated animals was normal when compared to that of the controls. Referring to the skin reactions reported above, the severity and frequency of the cutaneous reactions recorded in all the animals of the treated group, when compared with those noted in 4 out of 10 animals of the control group, enabled to attribute local reactions to a

skin sensitization effect.

Applicant's summary and conclusion

Interpretation of results:

sensitising

Remarks:

Migrated information

Executive summary:

Cyclopentyl chloroformate was tested for sensitization properties in an guinea pig maximization test according to OECD 406 and GLP. Ten male and female DunkingHartley guinea pig were treated intra dermally with 0.1 % cyclopentyl chloroformate. 15 % cyclopentyl chloroformate was applied epicutaneously one week later (occlusive conditions). A control was treated in parallel with corn oil. Challenge application with 20 % cyclopentyl chloroformate was for control and test substance group 14 days later. In all treated animals, a moderate or severe erythema (scores of 3 or 4) was observed after 24 and/or 48 hours in all. In 2 out of the 20 animals effects were described as skin destruction after 48 hours . In the 18 remaining animals . dryness of the skin appeared after 48 hours . In animals of the control group, moderate signs of a primary cutaneous irritation were noted in 4/5 males (erythema . scores of 1 or 2). Effect disappeared within the first 24 h. No erythema was noted in females After 48 hours, dryness of the skin appeared in 8 out of 10 animals. The severity and frequency of the cutaneous reactions all animals of the treated group in comparisons to the control group, demonstrated strong skin sensitizing properties of cyclopentyl chloroformate.