Phosphoric acid, mono- and di-C16-18 (even numbered) alkyl esters, salts with 2,2'-iminodiethanol

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Remarks:

In vivo, performed before 2018. Presented OECD 406 GPMT study was initially conducted for non-REACH related use of the registered substance. To avoid further vertebrate animal testing (i.e. conduct a new OECD 429 LLNA) this OECD 406 study is presented.

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1989

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Justification for type of information:

Presented OECD 406 GPMT study was initially conducted for non-REACH related use of the registered substance. In order to avoid further vertebrate animal testing (i.e. conduct a new OECD 429 LLNA) this OECD 406 study is presented.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1989

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

yes

Remarks:

- no results on positive control reported. - FCA mixture preapred with paraffin not water or saline. - Temperature range 20 - 25 °C. - minor methodical deviations due to pre-OECD 406 guidance release execution date of study

GLP compliance:

not specified

Remarks:

Statement on periodic checks by quality control unit attached to the study report.

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Presented OECD 406 GPMT study was initially conducted for non-REACH related use for a structural analogue of the registered substance.
In order to avoid further vertebrate animal testing (i.e. conduct a new OECD 429 LLNA) this OECD 406 study is presented.

Test material

Test material information

Constituent 1

Reference substance name:

Phosphoric acid, mixed decyl and octyl esters, compds. with diethanolamine

Cas Number:

68425-57-0

Molecular formula:

not applicable

IUPAC Name:

Phosphoric acid, mixed decyl and octyl esters, compds. with diethanolamine

Test material form:

solid: crystalline

Details on test material:

Name: Phosphoric acid, mixed decyl and octyl esters, compds. with diethanolamine
CAS No.: 68425-57-0
Physical state: white solid at 20 °C (turbid, non-pourable paste)

Specific details on test material used for the study:

Name: Phosphoric acid, mixed decyl and octyl esters, compds. with diethanolamine (Phosphorsaeure (C8-C10-Alkyl)-ester, DEA-Salz)
CAS No.: 68425-57-0 (68186-45-8)
Physical state: white solid at 20 °C (turbid, non-pourable paste)

In vivo test system

Test animals

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

male

Details on test animals and environmental conditions:

Species / Strain: Guinea pig / Pirbright white
Breeder: Winkelmann
Number of animals: 20
Sex: Female
Age (at start of administration): 29 days
Body weight (at start of administration): 330 – 480 g
Identification of animals: By cage label and coloured marks
Adaptation period: At least 5 days

Diet
Commercial diet Altromin served as food. This food was offered ad libitum. This diet contains an adequate amount of ascorbic acid as required by the OECD Testing Guideline 406.

Housing
The animals were kept in groups of 5 in MAKROLON cages at a room temperature of 20°C - 25 °C (maximum range) and a relative humidity of 45% - 75% (maximum range). Rooms were lit on a 12-hour light/12-hour dark cycle.
Granulated textured wood was used as bedding material in the cages. The cages were changed and cleaned twice a week.

Drinking water
Tap water (in drinking bottles) was offered ad libitum.

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

paraffin oil

Concentration / amount:

0.1 mL of a 1 % dispersion of test item in paraffin DAB 8

Day(s)/duration:

0 d

Adequacy of induction:

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

Induction 2

Route:

intradermal

Vehicle:

paraffin oil

Concentration / amount:

0.1 mL of test item mixture (mixture of FCA and test item in paraffin DAB 8; end concentration of test item 1% and 50 % FCA)

Day(s)/duration:

0 d

Adequacy of induction:

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

Induction 3

Route:

intradermal

Vehicle:

paraffin oil

Concentration / amount:

0.1 ml of Freund's complete adjuvant (FCA) diluted 1 : 1 paraffin DAB 8 (50 % FCA).

Day(s)/duration:

0 d

Adequacy of induction:

other: adjuvant

Induction 4

Route:

epicutaneous, occlusive

Vehicle:

paraffin oil

Concentration / amount:

20% concentration / 0.5 mL of the test preparation was spread over a wound plaster (Hansapor steril) and applied to the test area and held in contact by an occlusive dressing.

Day(s)/duration:

7 d / 48 h

Adequacy of induction:

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

Challenge

No.:

#1

Route:

epicutaneous, occlusive

Vehicle:

paraffin oil

Concentration / amount:

2% concentration / 0.1 ml of the test preparation was spread over wound plaster (6 cm2) and applied to the test area and held in contact by an occlusive dressing

Day(s)/duration:

21 d / 24 h; assessment of skin reaction after further 24h and 48h.

Adequacy of challenge:

highest non-irritant concentration

No. of animals per dose:

20 test and 10 control animals were used.

Details on study design:

The body weight was analysed statistically using STUDENT's t-test (p ≤ 0.01).

Challenge controls:

The vehicle control animals were treated in the same way as the animals of the test group, but received the vehicle paraffin DAB 8 instead of the test item and no injection of vehicle and FCA mixture.

Positive control substance(s):

not specified

Remarks:

alpha-hexyl cinnamic aldehyde (limited details on results reported)

Results and discussion

Positive control results:

The vehicle control revealed no skin reactions.
Positive control: alpha-hexyl cinnamic aldehyde (limited details on results reported)
Animals of the same strain treated with alpha-hexyl cinnamic aldehyde exhibited a sensitising reaction in all animals in form of a moderate and confluent erythema (grade 2, 24 hours after start of exposure) and a discrete or patchy erythema (grade 1, 24 hours after start of exposure).

In vivo (non-LLNA)

Results

Results 1

Key result

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

2 mL of a 1% solution of the test item in water

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

Behaviour remained unchanged during the course of the study.

Remarks on result:

no indication of skin sensitisation

Results 2

Key result

Reading:

2nd reading

Hours after challenge:

72

Group:

test chemical

Dose level:

2 mL of a 1% solution of the test item in water

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

Behaviour remained unchanged during the course of the study.

Remarks on result:

no indication of skin sensitisation

Any other information on results incl. tables

Method

The test item was tested for its sensitization potential on the skin using a maximization method [according to Magnusson, B. and Kligman, A.M., J. Invest. Dermatol. 52, 268-276 (1969)]. In the test 36 guinea pigs from the strain: Pirbright white were used (2 x 3 animals for preexperiments, 20 »substance animals« and 10 control animals).

Preexperiment

Preexperiments showed that 1 per cent respectively 20 per cent substance was minimal irritating (intracutaneous and epicutaneous) and 2 per cent substance was the maximal non-irritating dose (epicutaneous) on the skin of guinea pigs.

Main experiment

Paraffine perliquid DAB 8 was always utilized as vehicle. After intracutanous and epicutanous (occlusive/48 h) application, using the minimal irritating concentration of the substance, the retreatment was performed by application (occlusive/24 h) of the maximum non-irritating concentration. According to the test results obtained, the substance test item can be regarded as a "very slightly sensitizer" for guinea pigs (not sufficient for classification).

None of the test animals died prematurely. No test item-related differences in body weight and body weight gain were noted between the test animals of the control group and the test animals treated with the test item.

Table 1: Main experiment - assessment of skin reactions

Animal / cage no.	hours after
	intracutaneous induction	intracutaneous induction	epicutaneous induction	epicutaneous induction	topical  challenge	topical  challenge
	1 h	24 h	1 h	24 h	24 h	48 h
1 / 1	1	2	1	0	0	0
2 / 1	1	1	1	1	0	0
3 / 1	1	2	1	1	0	0
1 / 2	1	1	1	1	0	0
2 / 2	1	1	1	1	1	1
3 / 2	1	1	1	1	0	0
1 / 3	1	1	1	1	1	0
2 / 3	1	2	1	1	1	1
3 / 3	1	2	1	1	0	0
1 / 4	1	1	1	1	0	1
2 / 4	1	1	0	1	1	0
3 / 4	1	2	1	1	0	0
1 / 5	1	2	1	1	0	0
2 / 5	1	2	1	1	0	0
3 / 5	1	2	1	1	0	0
1 / 6	1	1	2	1	0	0
2 / 6	1	2	1	1	0	0
3 / 6	1	2	1	1	0	0
1 / 7	1	1	0	1	0	0
2 / 7	1	1	1	1	0	0

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

Phosphoric acid, C8-10-branched and linear alkyl esters, compds. with diethanolamine revealed no sensitising properties in guinea pigs in a test model according to Magnusson and Kligman.

Executive summary:

The purpose of this study was to determine the potential of the test item to produce skin sensitisation reactions in guinea pigs in a test model according to the Magnusson and Kligman method. A 1% solution of the test item in water chosen for the 1st (intracutaneous) induction stage revealed a moderate and confluent erythema 24 and 48 hours after administration in all 10 test item-treated animals. 2 mL of a 25% suspension of water/animal chosen for the 2nd (topical) induction revealed a moderate and confluent erythema 24 and 48 hours after administration in all 10 test item-treated animals. The challenge with 2 mL of a 1% solution of the test item/animal revealed no skin irritation in any animal and, thus, the test item had no sensitising properties. The vehicle control revealed no skin reactions. Animals of the same strain treated with alpha-hexyl cinnamic aldehyde in sesame oil exhibited a sensitising reaction in all animals in form of a moderate and confluent erythema (grade 2, 48 hours after start of exposure) and a discrete or patchy erythema (grade 1, 72 hours after start of exposure).

The body weight gain of the animals treated with test item was within the range of the vehicle control during the experiment.

Behaviour remained unchanged during the course of the study.