Tetramethylammonium hydrogen phthalate

Skin sensitisation

Administrative data

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

August-September 1993

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles. Test substance only indicated by abbreviation; no information on test substance purity. No positive control included.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1993

Report date:

1993

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Version / remarks:

1981

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.6 (Skin Sensitisation)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Test material

Test material information

Constituent 1

Reference substance name:

Tetramethylammonium hydrogen phthalate

EC Number:

416-900-5

EC Name:

Tetramethylammonium hydrogen phthalate

Cas Number:

79723-02-7

Molecular formula:

C12H17NO4

IUPAC Name:

tetramethylazanium 2-carboxybenzoate

Test material form:

other: solid

Details on test material:

- Name of test material (as cited in study report): TMAP
- Physical state: white solid
- Storage condition of test material: room temperature over silica 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: David Hall Limited, Burton-on-Trent, Staffordshire, U.K.
- Age at study initiation: approximately eight to twelve weeks old
- Weight at study initiation: 328 - 408g
- Housing: in groups of up to three in solid-floor polypropylene cages furnished with softwood shavings.
- Diet: ad libitum (Guinea Pig FD1 Diet, Special Diet Services Limited, Witham, Essex, U.K.)
- Water: free access to mains tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24
- Humidity (%): 47 - 66
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12

IN-LIFE DATES: From: 2 August 1993 To: 4 September 1993

Study design: in vivo (non-LLNA)

Induction

Induction 1

Route:

intradermal

Vehicle:

water

Induction 2

Concentration / amount:

5% w/v in distilled water

Challenge

Challenge 1

Route:

epicutaneous, occlusive

Vehicle:

water

Challenge 2

Concentration / amount:

5% w/v in distilled water

No. of animals per dose:

number of animals in test group: 20
number of animals in negative control group: 10

Details on study design:

RANGE FINDING TESTS:
Four animals were intradermally injected with preparations of test material (1%, 5%, 10% or 25% w/v in distilled water). The highest concentration that did not cause local necrosis, ulceration or systemic toxicity, was selected for the intradermal induction stage of the main study. For selection of concentration for topical induction two guinea pigs (intradermally injected with Freund's Complete Adjuvant nine days earlier) were treated with four preparations of the test material (75%, 50%, 25% and 10% w/w in distilled water). The highest concentration producing only mild to moderate dermal irritation after a 48-hour occlusive exposure, was selected for the topical induction stage of the main study. For selection of concentration for topical challenge four preparations of the test material (75%, 50%, 25% and 10% w/w in distilled water) were applied occlusively to the flanks of two guinea pigs for a period of 24 hours. These guinea pigs did not form part of the main study but had been treated identically to the control animals of the main study, up to Day 14. The highest nonirritant concentration of the test material and one lower concentration were selected for the topical challenge stage of the main study.

MAIN STUDY
A. INDUCTION EXPOSURE
A row of three injections (0.1 ml each) was made on each side of the mid-line. The injections were:
i) Freund's Complete Adjuvant plus distilled water in the ratio 1:1.
ii) a 5% (w/v) dilution of test material in distilled water.
iii) a 5% (w/v) dilution of test material in a 1:1 preparation of
Freund's Complete Adjuvant plus distilled water.
Approximately 24 and 48 hours after intradermal injection the degree of erythema at the test material injection sites were evaluated according to the scale from Draize J.H. (1959).
On day 6, the nuchal region was clipped and shaved. A volume of 0.5 ml of sodium laurylsulphate (10% w/w in petrolatum) was applied in order to provoke an in flammatory response. The treatment sites remained non-occluded.

One week later, the same area on the shoulder region used previously for intradermal injections was treated with a topical application of the test material formulation (75% w/w in distilled water). The test material formulation (0.2 - 0.3 ml) was applied on filter paper (approximate size 40 mm x 20 mm) which was held in place by a strip of surgical adhesive tape (approximate size 60 mm x 25 mm) and covered with an overlapping length of aluminium foil. The patch and foil were further secured by a strip of elastic adhesive bandage (approximate size 250 mm x 35 mm) wound in a double layer around the torso of each animal . This occlusive dressing was kept in place for 48 hours. The treatment sites were decontaminated with distilled water after patch removal due to the toxicity of the test material.
Erythematous reactions were quantified one and twenty-four hours following removal of the patches using the scale from Draize J.H. (1959).
Control animals were treated in the same way except for exposure to the test material.

B. CHALLENGE EXPOSURE
On Day 21, an approximately 50 mm x70 mm area on both flanks of each animal, was clipped free of hair. A quantity of 0.1 - 0.2 ml of the test material formulation (75% w/w in distilled water) was applied to the shorn right flank of each animal on a square of filter paper (approximate size 20 mm x 20 mm) which was held in place by a strip of surgical adhesive tape (approximate size 40 mm x 50 mm). To ensure that the maximum non-irritant concentration was used at challenge, the test material at a concentration of 50% w/w in distilled water was also similarly applied to a separate skin site on the right shorn flank. The vehicle alone was similarly applied to the left shorn flank. The patches were occluded with an overlapping length of aluminium foil and secured by a strip of elastic adhesive bandage (approximate size 250 mm x 75 mm) wound in a double layer around the torso of each animal. After 24 hours, the dressing was carefully removed and discarded. The challenge sites were swabbed with cotton wool soaked in distilled water to remove residual material. The vehicle sites were similarly swabbed. Approximately 24 and 48 hours after challenge dressing removal erythematous reactions were quantified using the Draize scale.

Challenge controls:

see details on study design

Positive control substance(s):

yes

Remarks:

DNCB

Results and discussion

Positive control results:

10 test group females and 5 control females were treated with the following preparations of 2,4-Dinitrochlorobenzene (DNCB) for the induction and challenge phases: Intradermal Induction (Day 0): 0.5% (w/v) in arachis oil B.P; Topical Induction (Day 7): 0.75% (w/v) in absolute ethanol; Topical Challenge (Day 21): 0.25% (w/v) in absolute ethanol. The known contact sensitiser, 2,4-Dinitrochlorobenzene (DNCB), produced a 90% (9/10) sensitisation rate.

In vivo (non-LLNA)

Results

Results 1

Reading:

other: 1st and 2nd

Hours after challenge:

24

Group:

test chemical

Dose level:

50% and 75%

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: other: 1st and 2nd. . Hours after challenge: 24.0. Group: test group. Dose level: 50% and 75%. No with. + reactions: 0.0. Total no. in groups: 20.0.

Results 2

Reading:

other: 1st and 2nd

Hours after challenge:

24

Group:

negative control

Dose level:

50% and 75%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: other: 1st and 2nd. . Hours after challenge: 24.0. Group: negative control. Dose level: 50% and 75%. No with. + reactions: 0.0. Total no. in groups: 10.0.

Any other information on results incl. tables

Maximum concentration not causing irritating effects in the range-finding test: 75%

Based on the results of the range-finding test, the following concentrations were selected for the main study:

intradermal induction: 5% (w/v) in distilled water

topical induction: 75% (w/w) in distilled water

topical challenge : 75% and 50% (w/w) in distilled water

Main study:

Signs of irritation during induction: very slight to well-defined erythema was noted at the induction sites of all test group animals one hour after dressing removal.

Very slight erythema was noted at the induction sites of nine test group animals at 24 -hour observation. Other skin reactions noted at the induction sites of test group animals were bleeding and a hardened light brown-coloured scab.

No skin reactions were noted at the treatment sites of the control group animals at the 1- and 24 -hour observations.

Bodyweight gains of guinea pigs in the test group, between Day 0 and Day 24, were comparable to those observed in the control group animals over the same period.

Applicant's summary and conclusion

Interpretation of results:

not sensitising

Remarks:

Migrated information Criteria used for interpretation of results: EU

Conclusions:

In a guinea pig maximisation test, TMAP showed no skin reaction after intradermal and topical induction. As a result, TMAP is considered to have no skin sensitizing properties.

Executive summary:

TMAP was tested for skin sensitising properties in a guinea pig maximisation test according to OECD 406 guideline. Twenty females were used for the test substance and 10 females were included for control treatment. Intradermal induction was done by intradermal injection of 5% TMAP (w/v) in distilled water, followed by dermal application of 10% SDS after 6 days. On Day 7, treatment was followed by topical induction of 75% TMAP (w/w) in distilled water, by using an occlusive dressing for 48 hours. On Day 21, animals were challenged by dermal application of 75% and 50% in distilled water on separate skin sites (under occlusion). Skin reaction was scored according to Draize after 24h and after 48h after challenge. No effects on body weight gain was observed. After intradermal induction, very slight to well-defined erythema was noted in all test group animals after 24 hours, and very slight erythema was noted after 48 hours in 11/20 animals. After topical challenge, no skin reactions were noted at the 24h and the 48h observations. Based on these results, TMAP is considered not to be a skin sensitizer.