[cyclohexane-1,2-diylbis[nitrilobis(methylene)]]tetrakisphosphonic acid, sodium salt

Endpoint summary

Description of key information

[Cyclohexane-1,2-diylbis[nitrilobis(methylene)]]tetrakisphosphonic acid, sodium salt was found not sensitising to guinea pig skin as reported in a Guinea pig maximisation study conducted according to OECD test guideline 406 and in compliance with GLP (Safepharm, 1992)

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

experimental study

Adequacy of study:

key study

Study period:

28 July 1982 - 4 September 1992

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

GLP compliance:

yes (incl. QA statement)

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

At the time of testing (1992), the LLNA method was not yet adopted as OECD Guideline and not yet identified as the standard information requirement for in-vivo sensitization.
For this reason the use of the guinea pig maximisation test (GPMT) was considered te be acceptable.

Species:

guinea pig

Strain:

Dunkin-Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: David Hall Ltd., Burton-on Trent, Stratfodshire, UK
- Age at study initiation: 8-12 weeks
- Weight at study initiation: 370-450g
- Housing: The animals were housed in groups of up to three in solid-floor polypropylene cages furnished with softwood shavings.
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: minimum of 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-23
- Humidity (%): 46-72
- Air changes (per hr): ca. 15
- Photoperiod (hrs dark / hrs light): 12/12

Route:

intradermal and epicutaneous

Vehicle:

water

Concentration / amount:

Intradermal induction: 1% (w/v) in distilled water
Topical induction: undiluted as supplied
Topical challenge: undiluted as supplied and 75% (v/v) in distilled water

Route:

epicutaneous, occlusive

Vehicle:

water

Concentration / amount:

Intradermal induction: 1% (w/v) in distilled water
Topical induction: undiluted as supplied
Topical challenge: undiluted as supplied and 75% (v/v) in distilled water

No. of animals per dose:

20 test and 10 control animals were used for the main study

Details on study design:

RANGE FINDING TESTS:
Selection of concentrations for intradermal induction:
Two animals were intradermally injected with preparations of test material (1% or 5% w/v in distilled water). The highest concentration that did not cause local necrosis, ulceration or systemic toxixicty, was selected for the intradermal induction stage of the main study.

Selection of concentration for topical induction:
Two guinea pigs (intradermally injected with Freund's complete adjuvant twenty one days earlier) were treated with the undiluted test material and three preparations of the test material (75%, 50% and 25% v/v in distilled water). The highest concentration producing only mild to moderate dermal irritation after the 48 hour occlusive exposure, was selected for the topical induction stage of the maiin study.

Selection of concentration for topical challenge:
The undiluted test material and three preparations of the test material (75%, 50% and 25% v/v 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 non-irritant 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
On day 0 a row of three injections (0.1ml) was made on each side of the midline. The injections were: (1) Freund's complete adjuvant plus distilled water in the ratio 1:1, (2) a 1% (w/v) dilution of test material in distilled water, (3) a 1% (w/v) dilution of test material in a 1:1 preparation of Freund's complete adjuvant plus distilled water. One week later (Day 7), the same area on the shoulder region used previously for intradermal injections was clipped again and treated with a topical application of the undiluted test material, which was applied on filter paper which was held in place occlusively for 48 hours. Erythemaeous reactions were quantified one and twenty-four hours following removal of the patches. In the case of the induction of the control animals, intradermal injections were administered using an identical procedure to that used for the test animals, except that test material was omitted and substituted with distilled water. The topical applications followed the same procedure as for the test animals except that nothign was applied to the filter paper.

B. CHALLENGE EXPOSURE
On Day 21, a quantity of the undiluted test material was applied to the shorn right flank of each animal on a square of filter paper which was held in place. To ensure that the maximum non-irritant concentration was used at challenge, the test material at a concentration of 75% (v/v) 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, and torso wrapped. After 24 hours the dressing was removed, and their position identified with marker-pen. Approximately 24 and 48 hours after challenge dressing removal, erythematous reactions were quantified using the four point scale shown overleaf.

Positive control substance(s):

yes

Remarks:

DNCB (89% sensitisation rate)

Positive control results:

DNCB produced an incidence on 16/18 sensitising reactions.

Reading:

rechallenge

Hours after challenge:

24

Group:

test chemical

Dose level:

Undiluted and 75% v/v in distilled water

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 24.0. Group: test group. Dose level: Undiluted and 75% v/v in distilled water. No with. + reactions: 0.0. Total no. in groups: 20.0.

Reading:

rechallenge

Hours after challenge:

48

Group:

test chemical

Dose level:

Undiluted and 75% v/v in distilled water

No. with + reactions:

0

Total no. in group:

20

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: Undiluted and 75% v/v in distilled water. No with. + reactions: 0.0. Total no. in groups: 20.0.

Reading:

rechallenge

Hours after challenge:

24

Group:

negative control

Dose level:

adjuvant and distilled water in the ratio of 1:1

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 24.0. Group: negative control. Dose level: adjuvant and distilled water in the ratio of 1:1. No with. + reactions: 0.0. Total no. in groups: 10.0.

Reading:

rechallenge

Hours after challenge:

48

Group:

negative control

Dose level:

adjuvant and distilled water in the ratio of 1:1

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 48.0. Group: negative control. Dose level: adjuvant and distilled water in the ratio of 1:1. No with. + reactions: 0.0. Total no. in groups: 10.0.

Interpretation of results:

GHS criteria not met

Conclusions:

ALBRITE CIX(N) was found not sensitising to guinea pig skin as reported in a Guinea pig maximisation study conducted according to OECD test guideline 406 and in compliance with GLP.

Executive summary:

In a GLP-compliant OECD Guideline 406 study, skin sensitisation properties of ALBRITE CIX(N) were studied in guinea pig.

Test substance concentrations selected for the main study were based on the results of a preliminary study. In the main study, twenty experimental animals were intradermally injected with a 1% concentration and epidermally exposed to the undiluted test substance. Ten control animals were similarly treated, but with the vehicle (distilled water) only. Two weeks after epidermal application all animals were challenged with a 75% test substance concentration an the vehicle. No evidence was obtained that the test substance had caused skin hypersensitivity in the guinea pig, since no responses were observed in the experimental animals in the challenge phase. This result indicates a sensitization rate of 0%.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

On Day 0 of a Guinea pig Maximisation test conducted to OECD test guideline 406 and GLP, 20 female Dunkin-Hartley guinea pigs were given a series of three intradermal induction injections. These injections were either 1% (w/v) test substance plus distilled water, 1% (w/v) test substance plus Freund's Complete Adjuvant, or distilled water with Freund's Complete Adjuvant. On Day 7 an occlusive 48 hour topical induction of the undiluted test substance was applied to the same site on the shoulder. Ten negative control animals received the same induction treatment, but the test substance was replaced by distilled water in the intradermal injections and an untreated dressing was applied in the topical induction. Skin reactions were recorded 1 and 24 hours after removal of these induction patches. On Day 21 the topical challenge phase was initiated. Undiluted test substance or 75% (v/v) test substance in distilled water was applied to different sites on the right flank of each animal (test and negative control). Distilled water was applied to the left flank (test and negative control). The positive control group was treated the same throughout, but the test substance was replaced by DNCB. For the challenge phase occlusive patches were left in place for 24 hours. Irritation reactions were graded 24 and 48 hours after dressing removal. The positive control group had a positive response in 16/18 animals. There were no positive skin reactions or adverse effects in the test or negative control groups. It was concluded that the test substance is not a skin sensitiser.

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no study available

Justification for classification or non-classification

Based on the available skin sensitisation study, [cyclohexane-1,2-diylbis[nitrilobis(methylene)]]tetrakisphosphonic acid, sodium salt is not classified for skin sensitisation under Regulation (EC) 1272/2008.