Phosphoric acid, mono- and di-isotridecyl esters, potassium salts

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Remarks:

Guinea-pig maximimisation test (GPMT)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

From June 25, 2008 to August 09, 2008

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

GPMT with read across substance was available before LLNA method implementation

Test material

Reference

Name:

Unnamed

Type:

Constituent

Type:

Constituent

Type:

Constituent

Type:

Constituent

Type:

Constituent

Test material form:

solid

Specific details on test material used for the study:

Appearance: semi-solid (amorphous): gel

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

Test animals:
- Source: Charles River Deutschland GmbH, Kisslegg, Germany
- Age at study initiation: 4-6 weeks
- Weight at study initiation: 341-370g (pretest); 328-384g (test and control group)
- Housing: individually in Makrolon type-4 cages with standard softwood bedding
- Diet (e.g. ad libitum): pelleted standard Kliba 3418 guinea pig breeding/maintenance diet (Provimi Kliba AG, Kaiseraugst, Switzerland), ad libitum
- Water (e.g. ad libitum): communitytap water, ad libitum
- Acclimation period (main study): 25. June - 22 July 2008

Environmental conditions:
- Temperature (°C): 22 +/- 3
- Humidity (%): 37-70
- Air changes (per hr): 10-15
- Photoperiod (hrs dark / hrs light): 12

In-life datesS: From: 25.06 2008 To: 19.08.2008

Study design: in vivo (non-LLNA)

Induction

Route:

intradermal and epicutaneous

Vehicle:

water

Concentration / amount:

intradermal induction: 1%
epidermal induction: 75%

Day(s)/duration:

48h

Adequacy of induction:

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

Challenge

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

10%

Day(s)/duration:

24h

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

10 test and 5 control animals

Details on study design:

Induction
- Intradermal injection / test day 1
An area of dorsal skin fron the scapular region (approx. 6x8 cm) was clipped free of hair. Three pairs of intradermal injections (0.1 mL/site) were made just within the boundaries of a 4x6 cm area in the clipped region as follows:
Test Group:
1) 1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline.
2) The test substance at 1% in purified water.
3) The test substance at 1% in a 1:1 (v/v) mixture of Fraund's Complete adjuvant and physiological saline.

Control Group:
1) 1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline.
2) purified water
3) 1:1 (w/w) mixture of purified water in a 1:1 (v/v) mixture of Freund's Complete Adjuvant and physiological saline.

- Epidermal induction / test day 8
One week after the injection, the scapular area (approx. 6x8 cm) was again clipped and shaved free of hair to the application. A 2x4 cm patch of filter paper was saturated with the test substance at 75% in purified water and placed over the injcetion sites of the test animals. The amount of the test substance preparation applied was approx. 0.3g. The patch was covered with aluminium foil and firmly secured by an plaser wrapped around the trunk of the animal and secured with impervious tape. The occlusive dressings were left in place for 48h. The epidermal application procedure described ensured intensive contact of the test substance.

The guinea pigs of the control group were treated as described above with purified water only, applied at a volume of aprox. 0.3 mL.

The reaction sites were assessed 24 and 48h after removal of the bandage for erythema and oedema according to the method of Magnusson and Kligman.

- Challenge / test day 22
The test and control guinea pigs were challenged two weeks after the epidermal induction application and were treated in the same way.

Hair was cliped and shaved from a 5x5 cm area on the left and right flank of each guinea pig just prior to the application. Two patches (3x3 cm) of filter paper were saturated with the test substance at the highest tested non-irritating concentration of 10% (applied to the left flank) and the vehicle only (purified water applied to the right flank) using the same method as for the epidermal application. The volume of the test substance preparation was approx. 0.2 mL and a volume of approx. 0.2 mL was used for the vehicle. The dressings were left in place for 24h.

21h after removal of the dressing the test sites treated with the test substance were depilated as described in the epidermal pretest.

Approx. 3h later (approx. 4h from start of the challenge application) the skin reaction was observed and recorded according to the numerical grading system following Magnusson and Kligman grading scale.

Approx. 24h after this observation (72h from the start of the challenge application) was made and once again recorded.

Challenge controls:

Vehicle only

Positive control substance(s):

yes

Remarks:

alpha-Hexylcinnamaldehyde in PEG 300

Results and discussion

Positive control results:

Discrete/patchy to moderate/confluent erythema with or without scaling was observed in nine out of ten test animals at the 24- and 48-hour reading after the challenge treatment with alpha-Hexylcinnamaldehyde at 1% in PEG 300 (left shoulder). Five test animals showed discrete/patchy erythema at the 24-hour reading after treatment with alpha-Hexylcinnamaldehyde at 0.1% in PEG 300 (left flank). No skin effect was observed in the control group.

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

other: test group and control group

Dose level:

10%

No. with + reactions:

0

Total no. in group:

15

Clinical observations:

no effects

Remarks on result:

other: number of animals: 10 and 5 for test and control groups, respectively

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

48

Group:

other: test group and control group

Dose level:

10%

No. with + reactions:

0

Total no. in group:

15

Clinical observations:

no effects

Remarks on result:

other: number of animals: 10 and 5 for test and control groups, respectively

Applicant's summary and conclusion

Interpretation of results:

other: CLP criteria not met

Remarks:

not classified

Conclusions:

Under the study conditions, the test substance was considered te be not sensitizing to guinea-pig skin (GPMT).

Executive summary:

A study was conducted to determine the skin sensitization potential of the test substance according to OECD Guideline 406 (guinea-pig maximisation test), in compliance with GLP. The test was performed in 15 (10 test and 4 control) female albino Dunkin Hartley guinea pigs. The intradermal induction of sensitization in the test group was performed in the nuchal region with a 1% dilution of the test substance in purified water in an emulsion of Freund's Complete Adjuvant (FCA)/physiological saline. The epidermal induction of sensitization was conducted for 48 hours under occlusion with the test substance at 75 % in purified water one week after the intradermal induction. The animals of the control group were intradermally induced with purified water under occlusion. Two weeks after epidermal induction the control and test animals were challenged by epidermal application of the test substance at 10 % in purified water and purified water alone under occlusive dressing. Cutaneous reactions were evaluated at 24 and 48 hours after removal of the dressing. No toxic signs were evident in the guinea pigs of the control or test group. No deaths occurred. No skin effect/reaction was observed in the control and test group after challenge treatment with the test substance at 10% in purified water. Under the study conditions, the test substance was considered to be not sensitizing to guinea-pig skin (2008).