9-Octadecenoic acid (Z)-, monoester with 1,2,3-propanetriol ester with boric acid (H3BO3)

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

12 November 1981 to 11 January 1982

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guidelines and the study was conducted under GLP conditions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1982

Report date:

1982

Materials and methods

Test guideline

Test guideline 1

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

no

Test guideline 2

Qualifier:

equivalent or similar to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Existing study

Test material

Test material information

Constituent 1

Reference substance name:

9-Octadecenoic acid (Z)-, monoester with 1,2,3-propanetriol ester with boric acid (H3BO3)

EC Number:

264-092-6

EC Name:

9-Octadecenoic acid (Z)-, monoester with 1,2,3-propanetriol ester with boric acid (H3BO3)

Cas Number:

63310-16-7

Molecular formula:

C21H39O5B (based on the representative structure below)

IUPAC Name:

2-hydroxy-3-{[hydroxy({2-hydroxy-3-[(9Z)-octadec-9-enoyloxy]propoxy})boranyl]oxy}propyl (9Z)-octadec-9-enoate; 3-{[bis({2-hydroxy-3-[(9Z)-octadec-9-enoyloxy]propoxy})boranyl]oxy}-2-hydroxypropyl (9Z)-octadec-9-enoate; {2-hydroxy-3-[(9Z)-octadec-9-enoyloxy]propoxy}boronic acid

Test material form:

not specified

Details on test material:

- Physical state: liquid

In vivo test system

Test animals

Species:

guinea pig

Strain:

Hartley

Sex:

male

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Labs, Wilmington, Massachusetts, USA
- Age at study initiation: 56 days
- Weight at study initiation: 434 - 649 g
- Housing: individually in wire-bottom cages
- Diet: Purina Guinea Pig Chow 5025® ad libitum. Fresh lettuce provided weekly
- Water: purified water ad libitum
- Acclimation period: 28 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): approximately 22 ºC
- Humidity (%): 48 - 77 %
- Photoperiod (hrs dark / hrs light): 12 hours dark / 12 hours light

IN-LIFE DATES: From 12 November 1981 to 11 January 1982

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal and epicutaneous

Vehicle:

other: See "Details on study design" for information

Induction 2

Concentration / amount:

See "Details on study design" for information

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

other: See "Details on study design" for information

Challenge 2

Concentration / amount:

See "Details on study design" for information

No. of animals per dose:

20 animals per treatment group

Details on study design:

A. INTRADERMAL INDUCTION EXPOSURE
A 4 x 6 cm area on the back, behind the shoulder girdle, was clipped one day prior to treatment. Pre-test screening on surplus animals indicated that 1% test material (w/v in peanut oil) was the maximum concentration not causing local necrosis or ulceration upon intradermal injection. In the following anterior to posterior order, three pairs of intradermal injections were made along the back within a 2 x 4 cm area corresponding to the size of a patch applied one week later:
(a) 0.1 mL 1% test material w/v in peanut oil
(b) 0.1 mL Freund's Complete Adjuvent (emulsion prepared by blending the commercial adjuvent with equal volume of sterile saline)
(c) 0.1 mL 1% test material w/v in Freund's Complete Adjuvent (prepared by first dissolving/mixing oil-soluble test material in the commercial adjuvent before adding the appropriate volume of sterile saline)
The first two pairs of injections were given approximately 0.3 cm apart, the final pair were located 0.6 cm caudally from the second pair.

The concurrent control group received injections as follows:
(a) 0.1 mL peanut oil alone
(b) 0.1 mL Freund's Complete Adjuvent (commercial adjuvent and sterile saline in a 1:1 volume ratio)
(c) 0.1 mL peanut oil incorporated in Freund's Complete Adjuvent

The ethylenediamine (EDA) group received the following injections:
(a) 0.1 mL 0.1% EDA (w/v) in sterile saline
(b) 0.1 mL Freund's Complete Adjuvent
(c) 0.1 mL 0.1% EDA (w/v) in Freund's Complete Adjuvent

The concurrent control group received the following injections:
(a) 0.1 mL sterile saline
(b) 0.1 mL Freund's Complete Adjuvent
(c) 0.1 mL saline incorporated in Freund's Complete Adjuvent

B. TOPICAL INDUCTION EXPOSURE
One week after the intradermal injections, the same area of the animal's back was reclipped. Pre-test screening on surplus animals indicated 5% test material (w/v in peanut oil) as the appropriate concentration for producing mild-moderate skin irritation upon topical administration. 0.2 mL 5% test material (w/v in peanut oil) was mixed into 0.5 g petrolatum pre-weighed and spread over a 2 x 4 cm patch of Whatman 3 mm filter paper. The patch, with an overlying 5 cm square of polyethylene, was placed on the clipped area and was held in place for 48 hours by a cohesive bandage wrapped around the animal's torso. The concurrent control group was similarly treated with 0.2 mL peanut oil incorporated into 0.5 g of petrolatum. The EDA group was treated with 0.2 mL 5% EDA (w/v in saline) mixed in 0.5 g petrolatum, while its concurrent control group was treated with 0.2 mL saline mixed in 0.5 g petrolatum.

C. CHALLENGE EXPOSURE
Pre-test screening on surplus animals, previously treated with an intradermal injection of Freund's Complete Adjuvent, indicated 1% test material (w/v in peanut oil) as the appropriate non-irritating concentration for topical challenge. Fourteen days after the topical induction, challenge treatment was carried out on both flanks (clipped the previous day) as follows: 0.2 mL 1% test material (w/v in peanut oil) was mixed into 0.5 g petrolatum pre-weighed and spread on a 2 x 2 cm filter paper patch placed on the left flank while 0.2 mL peanut oil alone was mixed into 0.5 g petrolatum pre-weighed and spread on a patch placed on the right flank. A polyethylene square covered each of the patches and was held in places by cohesive wrap for 24 hours. Both the test material induced group and the concurrent control group received identical challenge treatment. Both the EDA-induced animals and the concurrent controls received topical administration of 0.2 mL 10% EDA (w/v in saline), mixed into 0.5 g petrolatum, on the left flank, and 0.2 mL saline (mixed into 0.5 g petrolatum) on the right flank.

Positive control substance(s):

yes

Remarks:

ethylenediamine (EDA)

Results and discussion

In vivo (non-LLNA)

Results

Results 1

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

1% (intradermal), 5% (topical); 1% (challenge)

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 1% (intradermal), 5% (topical); 1% (challenge). No with. + reactions: 0.0. Total no. in groups: 20.0.

Results 2

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

1% (intradermal), 5% (topical); 1% (challenge)

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 1% (intradermal), 5% (topical); 1% (challenge). No with. + reactions: 0.0. Total no. in groups: 20.0.

Results 3

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

0.1% (intradermal), 5% (topical); 10% (challenge)

No. with + reactions:

16

Total no. in group:

18

Clinical observations:

Acanthosis abd hyperkeratosis observed.

Remarks on result:

other: see Remark

Remarks:

Reading: 1st reading. . Hours after challenge: 24.0. Group: positive control. Dose level: 0.1% (intradermal), 5% (topical); 10% (challenge). No with. + reactions: 16.0. Total no. in groups: 18.0. Clinical observations: Acanthosis abd hyperkeratosis observed..

Results 4

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

0.1% (intradermal), 5% (topical); 10% (challenge)

No. with + reactions:

17

Total no. in group:

18

Clinical observations:

Acanthosis abd hyperkeratosis observed.

Remarks on result:

other: see Remark

Remarks:

Reading: 2nd reading. . Hours after challenge: 48.0. Group: positive control. Dose level: 0.1% (intradermal), 5% (topical); 10% (challenge). No with. + reactions: 17.0. Total no. in groups: 18.0. Clinical observations: Acanthosis abd hyperkeratosis observed..

Any other information on results incl. tables

Bodyweights

No significant differences between treated and control animals were observed.

Mortality

One test material control animal, two EDA-induced animals, and four EDA control animals died within 48 hours after the topical induction treatment. The cause of death for the animals was undetermined.

Histopathology

Pathologic changes in the skin were fairly uniform, consisting of acanthosis and hyperkeratosis. Pulmonary congestion and hepatopathy were observed in both induced and control animals, which died during the study.

Table 1: Skin Irritation Data

Treatment	24 hour reading		48 hour reading		Treatment	24 hour reading		48 hour reading
	left flank	right flank	left flank	right flank		left flank	right flank	left flank	right flank
Test material induced	0	0	0	0	Test material control	0	0	0	0
	0	0	0	0		0	0	1	1†
	0	0	0	0		0	0	2	1†
	0	0	0	0		1	0	2	2†
	0	0	0	0		0	0	1	3†
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0		0	0	0	0
	0	0	0	0
EDA induced	3	0	3	0	EDA control	0	0	0	0
	0	0	1	0		0	0	0	0
	3	0	3	0		0	0	0	0
	0	0	2	0		0	0	0	0
	3	0	3	0		0	0	0	0
	3	0	3	0		0	0	1	0
	2	0	3	0		3	0	1	2†
	2	0	2	0		0	0	1	0
	3	0	3	0		0	0	0	0
	3	0	3	0		0	0	0	0
	3	0	3	1		0	0	0	0
	3	0	3	0		2	2	2	2†
	2	0	2	0		0	0	1	1†
	3	0	3	3†		0	0	0	0
	3	0	3	0		3	0	3	0
	3	0	3	0		2	0	3	0
	3	0	3	0
	3	0	3	0

† excluded from positive response count; irritation observed on both the treated and control flanks

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

Neither the test material induced animals nor the test material control animals responded to challenge treatment. Therefore, no positive sensitisation reactions were observed for the test material and so the test material was considered to be unlikely to be a skin sensitiser. The study is considered to be reliable, relevant and adequate for risk assessment and classification and labelling purposes.

Executive summary:

The skin sensitisation potential of the test material was investigated following a methodology similar to standardised guidelines OECD 406 and EU Method B.6 using the guinea pig maximisation test. The procedure consisted of two induction treatments (an intradermal injection conducted on day 0 and a topical application scheduled on day 7) followed 14 days later by a challenge treatment (topical application). A concurrent control group was submitted to a similar regimen: induction treatments with the diluent followed by challenge treatment with the test material. A positive control group was also included to confirm validity of the test method employed. Under the conditions of the study neither the test material induced animals nor the test material control animals responded to challenge treatment. Therefore, no positive sensitisation reactions were observed for the test material and so the test material was considered to be unlikely to be a skin sensitiser. The positive control was shown to have the capacity to cause skin sensitisation confirming the validity of the protocol used for this study.