Dipotassium dihydrogen ethylenediaminetetraacetate

Endpoint summary

Key value for chemical safety assessment

Skin sensitisation

Link to relevant study records

Reference

Reference 1

Endpoint:

skin sensitisation: in vivo (non-LLNA)

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP guideline study (OECD 406)

Justification for type of information:

Read across is based on a structurally similar material. An analogue justification is attached in section 13 of dataset.

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

OECD Guideline 406 (Skin Sensitisation)

Deviations:

yes

Remarks:

Total amount of TS applied is not stated

GLP compliance:

yes

Type of study:

guinea pig maximisation test

Justification for non-LLNA method:

Chelating agents are generally very poorly dermally absorbed (<0.001%), use of the guinea pig maximisation test which employs intradermal injection during the induction phase would maximise potential exposure of test material to the immune system..

Species:

guinea pig

Strain:

Hartley

Sex:

female

Details on test animals and environmental conditions:

TEST ANIMALS
- Source: Charles River France, 76410 Saint-Aubin-16s-Elbeuf, France
- Age at study initiation: 3 months
- Weight at study initiation: 374 ± 22 g
- Housing: individually in polycarbonate cages with stainless steel lid
- Diet: "106 pelleted diet" ad libitum
- Water: ad libitum
- Acclimation period: at least 5 days


ENVIRONMENTAL CONDITIONS
- Temperature : 21 ± 2°C
- Relative humidity : 30 to 70%
- Light/dark cycle: 12 h/12 h
- Ventilation: approximately 12 cycles/hour of filtered, non-recycled air

Route:

intradermal and epicutaneous

Vehicle:

corn oil

Concentration / amount:

intradermal injections: TRILON BD at the concentration of 0.5% (w/w) in corn oil,
topical application: TRILON BD at the concentration of 30% (w/w) in corn oil

Route:

epicutaneous, occlusive

Vehicle:

corn oil

Concentration / amount:

intradermal injections: TRILON BD at the concentration of 0.5% (w/w) in corn oil,
topical application: TRILON BD at the concentration of 30% (w/w) in corn oil

No. of animals per dose:

5 females/control group
10 females/treated group

Details on study design:

RANGE FINDING TESTS:
In order to determine the concentration of the TS in the main study one range finding test were performed on two animals (1 male and 1 female).
By intradermal route (tested concentrations: 1 % and 0.1 % (w/w):
24 hours before treatment, the dorsal region of the animals was clipped, intradermal injections of the dosage form preparations (0.1 ml) were performed in the interscapular region , cutaneous reactions were evaluated approximately 24, 48 hours and 6 days after the injections.
By cutaneous route (tested concentrations: 30% and 10% (w/w):
24 hours before treatment, both flank regions of the animals were clipped, the filter paper of a chamber was fully-loaded with one dosage form preparation. The chamber was then applied to the clipped area of the skin (one concentration per flank) . The chamber was held in place by means of an occlusive dressing for 24 hours, cutaneous reactions were evaluated approximately 24 and 48 hours after removal of the dressings.

MAIN STUDY
A. INDUCTION EXPOSURE
- No. of exposures: once intradermal (day 1) and additionally once cutanenously (day 8)
- Test groups: 1
- Control group: 2

INTRADERMAL EXPOSURE
- Site: six injections as pairs in the interscapular area
- Treatment:
Test group: A) front row: Freund's complete adjuvant (FCA) at 50% (v/v) in 0.9% NaCl; B) middle row: 0.1 ml TS at 0.5% (w/w) in corn oil; C) test substance at 0.5% (w/w) in the mixture FCA/0.9% NaCl (50/50)
Control groups: The animals were given the same injections (A, B, C) but without test substance, only with the formulating agent.

CUTANEOUS EXPOSURE
- Site: interscapular area
- Treatment:
Test group: a pad of filter paper (approximately 8 cm2) was fully-loaded with the test substance at the concentration of 30% (w/w) and was then applied to the interscapular region of the animals. The pad was held in place for 48 hours by means of an adhesive hypoallergenic dressing and an adhesive anallergenic waterproof plaster
Control groups: received an application of the vehicle alone under the same experimental conditions.

B. CHALLENGE EXPOSURE
Challenge:
- Day of challenge: day 22 (21 days after intradermal induction)
- Exposure period: 24 h
- Site: interscapular
- Treatment:
Test groups: received an application of the test substance and vehicle. The filter paper of a chamber (Finn Chambero) was fully-loaded with the test substance at the concentration of 30% (w/w) and was then applied to a clipped area of the skin of the posterior right flank of all animals. The vehicle was applied under the same experimental conditions to the skin of the posterior left flank. The chambers were held in contact with the skin for 24 hours by means of an adhesive an allergenic waterproof plaster. On removal of the dressing, any residual test substance was removed by means of a moistened
gauze pad.
Control groups: Control group one was treated like the treatment group; control group 2 remained untreated
- Evaluation (hr after challenge): 24, 48 h

Rechallenge:
- Day of challenge: day 29
- Exposure period: 24 h
- Treatment: the animals of all groups received an application of the test substance at the concentration of 30% (w/w) to the anterior left flank and the vehicle to the anterior right flank, under the same experimental conditions as for the first challenge application.
- Evaluation (hr after challenge): 24, 48 h

Challenge controls:

Yes, 2 groups

Positive control substance(s):

yes

Remarks:

Mercaptobenzothiazole

Reading:

1st reading

Hours after challenge:

24

Group:

test chemical

Dose level:

30%

No. with + reactions:

3

Total no. in group:

10

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: test group. Dose level: 30%. No with. + reactions: 3.0. Total no. in groups: 10.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

test chemical

Dose level:

30%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: test group. Dose level: 30%. No with. + reactions: 0.0. Total no. in groups: 10.0.

Reading:

1st reading

Hours after challenge:

24

Group:

other: Negative control group 1

Dose level:

0%

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: other: Negative control group 1. Dose level: 0%. No with. + reactions: 0.0. Total no. in groups: 5.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

other: Negative control group 1

Dose level:

0%

No. with + reactions:

0

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: other: Negative control group 1. Dose level: 0%. No with. + reactions: 0.0. Total no. in groups: 10.0.

Reading:

rechallenge

Hours after challenge:

24

Group:

test chemical

Dose level:

30%

No. with + reactions:

1

Total no. in group:

10

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 24.0. Group: test group. Dose level: 30%. No with. + reactions: 1.0. Total no. in groups: 10.0.

Reading:

rechallenge

Hours after challenge:

48

Group:

test chemical

Dose level:

30%

No. with + reactions:

0

Total no. in group:

10

Clinical observations:

Dryness of skin in 3/10 animals

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 48.0. Group: test group. Dose level: 30%. No with. + reactions: 0.0. Total no. in groups: 10.0. Clinical observations: Dryness of skin in 3/10 animals.

Reading:

rechallenge

Hours after challenge:

24

Group:

other: Negative control group 1

Dose level:

30%

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 24.0. Group: other: Negative control group 1. Dose level: 30%. No with. + reactions: 0.0. Total no. in groups: 5.0.

Reading:

rechallenge

Hours after challenge:

48

Group:

other: 1egative control group 1

Dose level:

30%

No. with + reactions:

0

Total no. in group:

5

Clinical observations:

Dryness of skin was observed in 1/5 animals

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 48.0. Group: other: 1egative control group 1. Dose level: 30%. No with. + reactions: 0.0. Total no. in groups: 5.0. Clinical observations: Dryness of skin was observed in 1/5 animals.

Reading:

rechallenge

Hours after challenge:

24

Group:

other: Negative control group 2

Dose level:

30%

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 24.0. Group: other: Negative control group 2. Dose level: 30%. No with. + reactions: 0.0. Total no. in groups: 5.0.

Reading:

rechallenge

Hours after challenge:

48

Group:

other: negative control group 2

Dose level:

30%

No. with + reactions:

0

Total no. in group:

5

Remarks on result:

other: Reading: rechallenge. . Hours after challenge: 48.0. Group: other: negative control group 2. Dose level: 30%. No with. + reactions: 0.0. Total no. in groups: 5.0.

Reading:

1st reading

Hours after challenge:

24

Group:

positive control

Dose level:

20%

No. with + reactions:

7

Total no. in group:

10

Remarks on result:

other: Reading: 1st reading. . Hours after challenge: 24.0. Group: positive control. Dose level: 20%. No with. + reactions: 7.0. Total no. in groups: 10.0.

Reading:

2nd reading

Hours after challenge:

48

Group:

positive control

Dose level:

20%

No. with + reactions:

7

Total no. in group:

10

Remarks on result:

other: Reading: 2nd reading. . Hours after challenge: 48.0. Group: positive control. Dose level: 20%. No with. + reactions: 7.0. Total no. in groups: 10.0.

One animal of the control group 1 was found dead on day 13. Hypoactivity and dyspnea were observed prior to death. The authors stated that such spontaneous clinical signs and mortality are sometimes observed in this species.

Interpretation of results:

not sensitising

Remarks:

Migrated information

Reference 2

Endpoint:

skin sensitisation: in vitro

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

The Magnusson Kligman Test according to OECD Test Guideline 406 using Na2EDTA (purity 91%) was chosen as key study. This test was performed under GLP by BASF (2000). 10 test animals and 5 control animals were used. A 0.5% substance concentration in corn oil was used for intradermal induction and a 30% test concentration for topical induction. Control animals were treated with corn oil as vehicle control. The challenge was conducted with 30% Na2EDTA in corn oil. 3/10 test animals showed a discrete patchy erythema 24 h after patch removal, after 48 h 0/10 showed a patchy erythema. 7 days later a rechallenge was conducted using 30% substance in corn oil. 1/10 test animals exhibited a discrete patchy erythema after 24 h, which was reversible within 48 h. Control animals did not exhibit skin reaction after challenge or rechallenge. The positive control group using 20% mercaptobenzodiazol induced positive skin sensitisation reactions in 7/10 animals at the 24 and 48 h reading.

With Na3EDTA a Repeated Insult Patch Test gave a negative result (0/10 animals) (Henck, 1980). Within 10 days the animals received 4 topical treatments (0.1 ml) of 10% Na3EDTA in dipropyleneglycolmethylether; at the third treatment Freud's adjuvants was injected additionally. 2 weeks after the last treatment the challenge was conducted using 10% Na3EDTA in dipropyleneglykolmethylether. Within the same test Henck et al also tested for cross-sensitisation between the known skin sensitizer ethylenediamine (EDA) and Na3EDTA. Animals were sensitized with EDA and challenged topically with Na3EDTA on the one flank and EDA on the other. None of the animals reacted positive after the challenge with Na3EDTA, but all of the animals which were challenge with EDA showed a slight to marked erythema and slight edema. Therefore it was concluded that Na3EDTA does not cross-sensitize with EDA.

Human data:

Several reports on human skin sensitisation are available: 2/529 dermatitis patients reacted positive to EDTA (Angelini, 1985). In another study after patch testing with 1% EDTA 0.9% of 215 subjects reacted positive towards EDTA (Rudner, 1977). In further patch tests using Na2EDTA 13/743 or 10/345 patients reacted positive (Penvy1980, 1981). However, in the studies of Penvy a 10% solution was used not a 1% solution as usual. Therefore these result may only suggest irritation rather than contact allergy. In studies of Fisher (1986) hundreds of patients were tested without a single positive response.

Migrated from Short description of key information:
No skin sensitisation studies on edetic acid are available. However, except for acute toxic and local effects H4EDTA shows similar properties as Na salts of EDTA, therefore studies using Na2EDTA and Na3EDTA have been used for read across.
In the OECD 406 guideline study with Na2EDTA 3/10 guinea pigs showed a patch erythema after the first challenge and 1/10 after the second challenge. The reports on humans are conflicting and in case of the positive results it can not be ruled out that the reactions reflected irritation rather than sensitisation. However, overall these results do not warrant a labeling according to EU or GHS critieria, which was also confirmed by the independent evaluation of the MAK Commission for the Investigation of Health Hazards of Chemical Compounds in the work area (MAK, 46. Lieferung, 2009).

Respiratory sensitisation

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (not sensitising)

Additional information:

6/22 patients with stable asthma developed bronchoconstriction after inhaling 4 ml of a nebuliser containing EDTA (0.5 g/l) and benzalkonium bromide (0.25 g/l) as preservatives together with the bronchodilatator ipratropium bromide. When these six subjects inhaled 4 ml EDTA and benzalkonium bromide free ipratropium bromide solution all subjects showed bronchodilatation. Inhalation of EDTA and benzalkonium bromide administered separately (EDTA solutions containing 0.25-10 g/l) produced dose related bronchoconstriction which persisted for longer than 60 minutes. The cumulative geometric mean (range) of a 20% fall in FEV, (forced expiratory volume) was 2.40 g/l (1.2-12.8) for edetic acid. Although the mechanism by which edetic acid causes bronchoconstriction is uncertain, it probably relates to its action as a chelator of calcium ions (Beasley, 1987). However in a study by the same group (Beasley, 1989) those results could not be confirmed. EDTA (0.5 g/l) did not influence the bronchodilator effect of a single dose inhaled Duovent with fenoterol (0.31 g/l) and ipratropium bromide (0.13 g/l). However, the airway effects of repeated inhalations of EDTA were not investigated

In letters from industry (BASF-Letter, 2001) no adverse acute or chronic respiratory health effects from exposure to EDTA or Na4EDTA have been observed in workers.

Justification for classification or non-classification

Based on the results obtained in the toxicity studies and taking into account the provisions laid down in Regulation EC1272/2008, requirements for classification are not fulfilled with respect to dermal or respiratory sensitisation.