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Description of key information

In vitro skin and eye irritation studies did not show irritating properties of EDTA-Mn(NH4)2. In addition, in vitro studies with EDTA-MnNa2 and in vivo studies with EDTA-FeNa, a structurally related compound, did not show irritating properties (see also the read across document in section 13). 

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records
Reference
Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
17-Jun-2014 to 23-Jun-2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.
Qualifier:
according to
Guideline:
other: EU method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Deviations:
no
Qualifier:
according to
Guideline:
other: OECD Guideline no. 439: In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method
Deviations:
no
GLP compliance:
yes (incl. certificate)
Vehicle:
unchanged (no vehicle)
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 µl

NEGATIVE CONTOL:
- Amount(s) applied (volume or weight with unit): 10 µl Phosphate buffered saline

POSITIVE CONTROL
- Amount(s) applied (volume or weight with unit): 10 µl
- Concentration (if solution): 5% (aq) Sodium dodecyl sulphate
Duration of treatment / exposure:
Exposure:15 minutes
Post incubation period: 42 hours
Details on study design:
TEST SITE
- Area of exposure: human epidermis model
- % coverage: 0.38 cm2

REMOVAL OF TEST SUBSTANCE
- Washing (if done): phosphate buffered saline
- Time after start of exposure: 15 minutes

POST INCUBATION PERIOD
- 42 hours

SCORING SYSTEM:
- After a 42 hour incubation period, determination of the cytotoxic (irritancy) effect was performed. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT at the end of the treatment. Cell viability was calculated for each tissue as a percentage of the mean of the negative control tissues.
Irritation / corrosion parameter:
other:
Value:
82
Remarks on result:
other:
Remarks:
Basis: other: percentage of control. Time point: 15 minutes. (migrated information)

Skin irritation is expressed as the remaining cell viability after exposure to the test substance. The relative mean tissue viability obtained after 15 minutes treatment with EDTA-Mn(NH4)2 compared to the negative control tissues was 82%. Since the mean relative tissue viability for EDTA-Mn(NH4)2 was below 50% after 15 minutes treatment it is considered to be irritant.

 

The positive and negative controls were valid as they met the criteria, for results see the tables in the attached background material.

Interpretation of results:
not irritating
Remarks:
Migrated information
Conclusions:
It is concluded that EDTA-Mn(NH4)2 is non-irritant in the in vitro skin irritation test.

Executive summary:

Skin irritation is expressed as the remaining cell viability after exposure to the test substance. The relative mean tissue viability obtained after 15 minutes treatment with EDTA-Mn(NH4)2compared to the negative control tissues was 82%. Since the mean relative tissue viability for EDTA-Mn(NH4)2was above 50% EDTA-Mn(NH4)2is considered to be non-irritant.

The positive control had a mean cell viability of 35% after 15 minutes exposure. The absolute mean OD570(optical density at 570 nm) of the negative control tissues was within the laboratory historical control data range. The standard deviation value of the percentage viability of three tissues treated identically was less than 8%, indicating that the test system functioned properly.

Finally, it is concluded that this test is valid and that EDTA-Mn(NH4)2is non-irritant in thein vitroskin irritation test under the experimental conditions described in this report.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
28-May-2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study has been performed according to OECD and/or EC guidelines and according to GLP principles.
Qualifier:
according to
Guideline:
other: OECD guideline 437 “Bovine Corneal Opacity and Permeability Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage”
Deviations:
no
Qualifier:
according to
Guideline:
other: EU Method B.47 “Bovine corneal opacity and permeability method for identifying ocular corrosives and severe irritants
Deviations:
no
GLP compliance:
yes (incl. certificate)
Vehicle:
unchanged (no vehicle)
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 750 µl per cornea

NEGATIVE CONTROL
- Amount(s) applied (volume or weight with unit): 750 µl of physiological saline per cornea

POSITIVE CONTROL
Amount(s) applied (volume or weight with unit): 750 µl per cornea
Concentration (if solution): 10% (w/v) Benzalkonium Chloride


Duration of treatment / exposure:
10 minutes
Details on study design:
TEST SITE
- Isolated bovine cornea

REMOVAL OF TEST SUBSTANCE
- Washing (if done): yes
- Time after start of exposure: 10 minutes

SCORING SYSTEM:
- After exposure the cornea is thoroughly rinsed to remove the test substance followed by immediate opacity measurement and permeability evaluation of the cornea.

- The mean opacity and mean permeability values (OD490) were used for each treatment group to calculate an in vitro score:

In vitro irritancy score (IVIS) = mean opacity value + (15 x mean OD490 value).


TOOL USED TO ASSESS SCORE:
- opacitymeter and microplate reader

DATA EVALUATION:
In vitro score range UN GHS
≤ 3 No Category
> 3; ≤ 55 No prediction can be made
>55 Category 1

Irritation parameter:
other: In vitro irritancy score (IVIS)
Basis:
mean
Time point:
other: 10 minutes
Score:
0.9
Other effects:
The corneas were clear after the 10 minutes of treatment with EDTA-Mn(NH4)2.
A pH effect of the test substance was observed on the rinsing medium.
Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
EDTA-Mn(NH4)2 is not irritant or corrosive in the Bovine Corneal Opacity and Permeability test.

Executive summary:

The negative control responses of the opacity and permeability values were less than the upper limits of the laboratory historical range indicating that the negative control did not induce irritancy on the corneas.The mean in vitro irritancy score of the positive control (10% (w/v) Benzalkonium Chloride) was 126 and within the historical positive control data range. It was therefore concluded that the test conditions were adequate and that the test system functioned properly.

EDTA-Mn(NH4)2 did not induce ocular irritation through both endpoints, resulting in a mean in vitro irritancy score of 0.9 after 10 minutes of treatment.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Additional information

In vitro skin and eye irritation studies did not show irritating properties of EDTA-Mn(NH4)2; in vitro studies were carried out to to confirm the absence of skin and eye irritation as already had been found with a structurally related analag EDTA-FeNa and several other chelates (see read across document in section 13). In addition, it is not expected that EDTA-Mn(NH4)2 will irritate the respiratory tract in case of inhalation exposure of droplets.


Justification for selection of skin irritation / corrosion endpoint:
The results of this in vitro study were confirmed by in vivo skin irritation studies with other chelating agents (see also read across document in section 13)

Justification for selection of eye irritation endpoint:
The results of this in vitro study were confirmed by in vivo skin irritation studies with other chelating agents (see also read across document in section 13)

Justification for classification or non-classification

Because EDTA-Mn(NH4)2 did not show skin or eye irritation, no classification is needed.