Registration Dossier

Administrative data

Description of key information

skin sensitisation

No skin sensitisation studies on diammonium dihydrogen EDTA are available. Therefore studies using Na2EDTA and Na3EDTA have been used for read across. In the OECD 406 guideline study with Na2EDTA 3/10 animals 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 cannot 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). Additionally other salt from the ammonium category show no skin sensitizing potential.

respiratory sensitisation

No studies on respiratory sensitisation of (NH4)2EDTA are available, however two studies on dogs with airway hyperresponsiveness using Na2EDTA have been performed. In those dogs bronchoconstriction can be induced. Additionally bronchoconstriction was induced in 6/22 asthma patients after inhaling of a nebulizer containing 0.5 g/L EDTA amongst other substances. However this effect could not be reproduced in a study by the same group. Considering the fact that no adverse acute or chronic respiratory health effect was reported in workes exposed to Na4EDTA these results do not warrant a labeling according to EU or GHS criteria.

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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
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:
This study was conducted due to non-REACH regulatory requirements. With the existing data from this study not only being acceptable but of good quality (Klimisch Score 1), this study precludes the need for an additional LLNA study. In addition, a supplementary LLNA study would violate the ECHA objectives with regards to animal welfare.
Specific details on test material used for the study:
- Impurities: water 9.5% ( no explanation for the 100.5% total substance is given)
- Lot/batch No.: 41-9660
- Storage condition of test material: at room temperature
- pH (as 1% solution): 5
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 an allergenic 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
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
test chemical
Dose level:
30%
No. with + reactions:
3
Total no. in group:
10
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
test chemical
Dose level:
30%
No. with + reactions:
0
Total no. in group:
10
Key result
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
Key result
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
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
test chemical
Dose level:
30%
No. with + reactions:
1
Total no. in group:
10
Key result
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
Key result
Reading:
rechallenge
Hours after challenge:
24
Group:
other: Negative control group 1
Dose level:
30%
No. with + reactions:
0
Total no. in group:
5
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
other: Negative 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
Reading:
rechallenge
Hours after challenge:
24
Group:
other: Negative control group 2
Dose level:
30%
No. with + reactions:
0
Total no. in group:
5
Key result
Reading:
rechallenge
Hours after challenge:
48
Group:
other: Negative control group 2
Dose level:
30%
No. with + reactions:
0
Total no. in group:
5
Key result
Reading:
1st reading
Hours after challenge:
24
Group:
positive control
Dose level:
20%
No. with + reactions:
7
Total no. in group:
10
Key result
Reading:
2nd reading
Hours after challenge:
48
Group:
positive control
Dose level:
20%
No. with + reactions:
7
Total no. in group:
10

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:
GHS criteria not met
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not sensitising)
Additional information:

The Magnusson Klingman 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 animal 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.

Respiratory sensitisation

Endpoint conclusion
Additional information:

In a 5-minute inhalation challenge with 6% Na2EDTA (as aerosol) elicited bronchoconstriction in Basenji-Greyhound dogs with hyperreactive airways but not in mongrel dogs. There was an increase in pulmonary resistance (RL) (2.1 ± 0.4 cm H2O/L x s) prechallenge, 9 .0±1 .8 postchallenge) . Exposure with 6% CaNa2EDTA caused no changes in pulmonary resistance (Downes and Hirshman, 1983). In another study (Lindeman et al., 1993) the mechanism of chelator induced airway constriction was examined in anesthetised Basenji-Greyhound dogs after exposure of either 4% Na2EDTA or 4% CaNa2EDTA. Collateral resistance was significantly greater after Na2EDTA than after CaNa2EDTA exposure (ca. 1.5 versus 0.5 cm H2O/Lx s). Fluid volume recovered after bronchalveolar lavage (BAL), total cell counts and cell differentials did not differ significantly. However, a seven-fold increase in prostanoid concentration (PGD2) in the BAL fluid was found in Na2EDTA exposed dogs in comparison to CaNa2EDTA exposed dogs. There was a strong relationship between changes in collateral resistance and concentrations of prostanoids (PGD2) after Na2EDTA exposure but not after CaNa2EDTA exposure. It was concluded that calcium chelators such as Na2EDTA can produce airway constriction by stimulating release of bronchoconstricting prostanoids in dogs with airway hyperresponsiveness. In letters from industry (BASF, Dow, Akzo Nobel, CEFIC) it was reported that no adverse acute or chronic respiratory health effects from exposure to EDTA or Na4EDTA have been observed in workers (BASF-Letter, 2001).

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 (Beasly, 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.

Justification for classification or non-classification

Classification, Labelling, and Packaging Regulation (EC) No 1272/2008 

The available experimental test data are reliable and suitable for classification purposes under Regulation (EC) No 1272/2008. The substance is not considered to be classified for skin sensitisation under Regulation (EC) No 1272/2008, as amended for the ninth time in Regulation (EU) No 2016/1179. Based on the human data, classification concerning respiratory sensitisation is not warranted.