Registration Dossier

Administrative data

Description of key information

Skin irritation: In accordance with the testing strategy detailed in Annex VIII, column 1 of Regulation (EC) No. 1907/2006 the assessment of the endpoint ‘skin irritation or skin corrosion’  has been performed following the consecutive steps detailed in the Regulation. As such an in vitro skin corrosion study has been performed. This study is not considered as the key study because it is not sufficient for classification and labelling in accordance with Regulation (EC) No. 1272/2008 (EU CLP) and is therefore submitted as supporting data.  The key study (Warren N, 2012) is conducted according to an appropriate validated in vitro guideline and under the conditions of GLP and therefore the study is considered to be acceptable and to adequately satisfy both the guideline requirement and the regulatory requirement as a key study for this endpoint. In addition, the data is considered to be adequate and reliable for classification and labelling in accordance with Regulation (EC) No. 1272/2008 (EU CLP). 
Eye irritation: In accordance with the testing strategy detailed in Annex VIII, column 1 of Regulation (EC) No. 1907/2006 an ex vivo study has been performed prior to conducting an in vivo study. This study is not considered as the key study because it is not sufficient for classification and labelling in accordance with Regulation (EC) No. 1272/2008 (EU CLP). However, the study does support the conclusion that tetrairon tris(pyrophosphate) is not corrosive to the eyes and the data can therefore be used to support the conclusions made in the key study. The key study (Bradshaw J, 2012) is conducted according to an appropriate guideline and under the conditions of GLP and therefore the study is considered to be acceptable and to adequately satisfy the regulatory requirement as a key study for this endpoint.

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:
The study was performed between 12 June 2012 and 18 June 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. This study is conducted according to an appropriate validated in vitro guideline and under the conditions of GLP and therefore the study is considered to be acceptable and to adequately satisfy both the guideline requirement and the regulatory requirement as a key study for this endpoint. In addition, the data is considered to be adequate and reliable for classification and labelling in accordance with Regulation (EC) No. 1272/2008 (EU CLP).
Reference:
Composition 0
Qualifier:
according to
Guideline:
EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Date of inspectection: 19-21 July 2011 Date of Signature: 31 August 2011
Test material information:
Composition 1
Species:
other: reconstituted human epidermis model
Strain:
other: reconstituted human epidermis model
Details on test animals and environmental conditions:
Not applicable
Type of coverage:
other: Topical
Preparation of test site:
other: Not applicable
Vehicle:
other: No vehicle used
Controls:
no
Amount / concentration applied:
TEST MATERIAL

- The test Material was applied neat.

- Amount(s) applied (volume or weight with unit):
Approximately 10 mg of the test item was applied to the epidermis surface. The epidermis surface had previously been moistened with 5 μl of sterile distilled water to improve contact between the solid test item and the epidermis.

- Concentration (if solution):
The test material was used as supplied.

VEHICLE
No vehicle used
Duration of treatment / exposure:
15 minute exposure & 42 hour post-exposure incubation
Observation period:
Not applicable
Number of animals:
Not applicable
Details on study design:
TEST SITE
- Area of exposure:
Approximately 10 mg of the test item was applied to the epidermis surface. The epidermis surface had previously been moistened with 5 μl of sterile distilled water to improve contact between the solid test item and the epidermis.

- % coverage:
The test material was applied topically to the corresponding tissues ensuring uniform covering.

- Type of wrap if used:
None used

REMOVAL OF TEST SUBSTANCE
- Washing (if done):
At the end of the exposure period, each tissue was removed from the well using forceps and rinsed using a wash bottle containing DPBS with Ca++ and Mg++. Rinsing was achieved by filling and emptying each tissue insert for approximately 40 seconds using a constant soft stream of PBS to gently remove any residual test material. The rinsed tissues were transferred to the second column of 3 wells containing 2 ml of maintenance medium in each well. The rinsed tissues were incubated at 37ºC, 5% CO2 in air for 42 hours.

- Time after start of exposure:
15 minutes post-exposure

SCORING SYSTEM:
Quantitative MTT Assessment (percentage tissue viability)
For the test material the relative mean tissue viabilities obtained after the 15 minute treatment followed by the 42 hour post-exposure incubation period were compared to the mean of the negative control treated tissues (n=3). The relative mean viabilities were calculated in the following way:

mean OD540 of test material / mean OD540 of negative control x 100 = Relative mean tissue viability (percentage of negative control)

Classification of irritation potential is based upon relative tissue viability following the 15 minute exposure period followed by the 42 hour post-exposure incubation period according to the following:

Mean tissue viability is ≤50% : Irritant (I) R38

Mean tissue viability is >50% : Non-Irritant (NI)
Irritation / corrosion parameter:
other:
Value:
110.7
Remarks on result:
other:
Remarks:
Basis: mean Viability of cells (%). Time point: Day 6. Max. score: 100.0. Reversibility: other: Not applicable. Remarks: See relative mean viability below.. (migrated information)
Irritant / corrosive response data:
The relative mean viability of the test material treated tissues was 110.7% after a 15-minute exposure.
Other effects:
No

RESULTS

Direct MTT Reduction

The MTT solution containing the test material did not turn blue/purple which indicated that the test material did not directly reduce MTT.

Test Material, Positive Control Material and Negative Control Material

The individual and mean OD540 values, standard deviations and tissue viabilities for the test material, negative control material and positive control material are given in Table 1. The mean viabilities and standard deviations of the test material and positive control, relative to the negative control are also given in Table 1.

The relative mean viability of the test material treated tissues was 100.0% after a 15 minute exposure.

The MTT solution containing the test item did not turn blue which indicated that the test item did not directly reduce MTT.

Quality Criteria

The relative mean tissue viability for the positive control treated tissues was 8.6% relative to the negative control treated tissues and the standard deviation value of the percentage viability was 1.0%. The positive control acceptance criterion was therefore satisfied.

The mean OD540 for the negative control treated tissues was 0.659 and the standard deviation value of the percentage viability was 6.2%. The negative control acceptance criterion was therefore satisfied.

The standard deviation calculated from individual percentage tissue viabilities of the three identically treated tissues was 10.5%. The test item acceptance criterion was therefore satisfied.

Table1 : Mean OD540 Values and Percentage Viabilities for the Negative Control Material, Positive Control Material and Test Material

Material

OD540of tissues

Mean OD540of triplicate tissues

±SDof OD540

Relative individual tissue viability (%)

Relative mean viability (%)

± SD of Relative mean viability (%)

Negative Control Material¤

0.666

0.659

0.041

101.1

100*

6.2

0.696

105.6

0.615

93.3

Positive Control Material¤

0.059

0.056

0.006

9.0

8.6

1.0

0.061

9.3

0.049

7.4

Test Material

0.769

0.730

0.069

116.7

110.7

10.5

0.650

98.6

0.770

116.8


SD=    Standard deviation

*=     The mean viability of the negative control tissues is set at 100%

¤ = Control group shared with Harlan Laboratories Ltd, Project numbers 41200853, 41200860, 41200861, 41200866, 41200871, 41200880 and 41200884


Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test material was considered to be Non-Irritant (NI).
This study is conducted according to an appropriate validated in vitro guideline and under the conditions of GLP and therefore the study is considered to be acceptable and to adequately satisfy both the guideline requirement and the regulatory requirement as a key study for this endpoint. In addition, the data is considered to be adequate and reliable for classification and labelling in accordance with Regulation (EC) No. 1272/2008 (EU CLP). Tetrairon tris(pyrophosphate) is not considered to be classified in accordance with Regulation (EC) No. 1272/2008 (EU CLP).
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
The study was performed between 30 July 2012 and 09 August 2012.
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no deviations from standard test guidelines and no minor methodological deficiencies. This study is conducted according to an appropriate guideline and under the conditions of GLP and therefore the study is considered to be acceptable and to adequately satisfy both the guideline requirement and the regulatory requirement (Regulation (EC) No. 1907/2006; REACH) as a key study for this endpoint. In addition, this study is considered to be acceptable for classification and labelling in accordance with Regulation (EC) No. 1272/2008 (EU CLP).
Reference:
Composition 0
Qualifier:
according to
Guideline:
EU Method B.5 (Acute Toxicity: Eye Irritation / Corrosion)
Deviations:
no
Qualifier:
according to
Guideline:
OECD Guideline 405 (Acute Eye Irritation / Corrosion)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
Date of GLP inspection: 19-21 July 2011 Date of Signature on GLP certificate: 31 August 2011
Test material information:
Composition 1
Species:
rabbit
Strain:
New Zealand White
Details on test animals or tissues and environmental conditions:
TEST ANIMALS
- Source: Harlan Laboratories UK Ltd., Leicestershire, UK.

- Age at study initiation: Twelve to twenty weeks old

- Weight at study initiation: 2.18 or 2.25 kg

- Housing: The animals were individually housed in suspended cages. The animals were provided with environmental enrichment items which were considered not to contain any contaminant of a level that might have affected the purpose or integrity of the study.

- Diet (e.g. ad libitum): ad libitum (2930 Teklad Global Rabbit diet supplied by Harlan Laboratories UK Ltd., Oxon, UK)

- Water (e.g. ad libitum): ad libitum.

- Acclimation period: At least five days


ENVIRONMENTAL CONDITIONS

- Temperature (°C): 17 to 23°C

- Humidity (%): 30 to 70%

- Air changes (per hr): At least fifteen changes per hour

- Photoperiod (hrs dark / hrs light): Twelve hours continuous light (06:00 to 18:00) and twelve hours darkness


IN-LIFE DATES:
From: day 1 To:day 3
Vehicle:
unchanged (no vehicle)
Controls:
other: The left eye remained untreated and was used for control purposes.
Amount / concentration applied:
TEST MATERIAL

- Amount(s) applied (volume or weight with unit): A volume of 0.1 ml of the test item, which was found to weigh approximately 98 mg (as measured by gently compacting the required volume into an adapted syringe) was placed into the conjunctival sac of the right eye, formed by gently pulling the lower lid away from the eyeball.

- Concentration (if solution): Undiluted and used as supplied

VEHICLE

- Amount(s) applied (volume or weight with unit):
Not applicable

Duration of treatment / exposure:
72 hours
Observation period (in vivo):
Approximately 1 hour and 24, 48 and 72 hours following treatment.
Number of animals or in vitro replicates:
2 animals were tested in total. (After consideration of the ocular responses produced in the first treated animal, one additional animal was treated. )

Details on study design:
REMOVAL OF TEST SUBSTANCE

- Washing (if done):
Not applicable

- Time after start of exposure:
Not applicable


SCORING SYSTEM:
Assessment of ocular damage/irritation was made approximately 1 hour and 24, 48 and 72 hours following treatment, according to the numerical evaluation given in Appendix 2, (from Draize J H (1977) "Dermal and Eye Toxicity Tests" In: Principles and Procedures for Evaluating the Toxicity of Household Substances, National Academy of Sciences, Washington DC p.48 to 49).


TOOL USED TO ASSESS SCORE:

Examination of the eye was facilitated by the use of the light source from a standard ophthalmoscope.
Irritation parameter:
cornea opacity score
Basis:
animal: 72298 Male
Time point:
other: Mean 24, 48 and 72 hours
Score:
0
Max. score:
4
Reversibility:
other: No effects observed
Remarks on result:
other: Initial pain reaction = 2
Irritation parameter:
cornea opacity score
Basis:
animal: 72344 Male
Time point:
other: Mean 24, 48 and 72 hours
Score:
0
Max. score:
4
Reversibility:
other: No effects observed
Remarks on result:
other: Initial pain reaction = 2
Irritation parameter:
iris score
Basis:
animal: 72298 Male
Time point:
other: Mean 24, 48 and 72 hours
Score:
0
Max. score:
2
Reversibility:
other: No effect observed
Irritation parameter:
iris score
Basis:
animal: 72344 Male
Time point:
other: Mean 24, 48 and 72 hours
Score:
0
Max. score:
2
Reversibility:
other: No effect observed
Irritation parameter:
other: redness
Basis:
animal: 72298 Male
Time point:
other: Mean 24, 48 and 72 hours
Score:
0.33
Max. score:
3
Reversibility:
fully reversible within: 48 hours
Irritation parameter:
other: redness
Basis:
animal: 72344 Male
Time point:
other: Mean 24, 48 and 72 hours
Score:
1
Max. score:
3
Reversibility:
fully reversible within: 72 hours
Irritation parameter:
chemosis score
Basis:
animal: 72298 Male
Time point:
other: Mean 24, 48 and 72 hours
Score:
0
Max. score:
4
Reversibility:
other: No effect observed
Irritation parameter:
chemosis score
Basis:
animal: 72344 Male
Time point:
other: Mean 24, 48 and 72 hours
Score:
0.33
Max. score:
4
Reversibility:
fully reversible within: 48 hours
Irritant / corrosive response data:
Individual and group mean scores for ocular irritation are given in Table 1 and Table 2.
No corneal effects were noted during the test.
Iridial inflammation was noted in both treated eyes one hour after treatment.
Moderate conjunctival irritation was noted in both treated eyes one hour after treatment with minimal conjuctival irritation noted at the 24-hour observation. Minimal conjunctival irritation persisted in one treated eye at the 48-hour observation.
One treated eye appeared normal at the 48-hour observation and the other treated eye appeared normal at the 72-hour observation.
Other effects:
Bodyweight
Both animals showed expected gain in bodyweight during the study.

Interpretation of Results

The numerical values corresponding to each animal, tissue and observation time were recorded. The data relating to the conjunctivae were designated by the letters A (redness), B (chemosis) and C (discharge), those relating to the iris designated by the letter D and those relating to the cornea by the letters E (degree of opacity) and F (area of cornea involved). For each tissue the score was calculated as follows:

Score for conjunctivae =         (A + B + C) x 2
Score for iris                            =         D x 5
Score for cornea                      =         (E x F) x 5

Using the numerical data obtained a modified version of the system ( Modified Kay and Calandra Interpretation of Eye Irritation Test was used to classify the ocular irritancy potential of the test material. This was achieved by adding together the scores for the cornea, iris and conjunctivae for each time point for each rabbit. The group means of the total scores for each observation were calculated. The highest of these group means (the maximum group mean score) together with the persistence of the reactions enabled classification of the eye irritancy potential of the test material.

If evidence of irreversible ocular damage is noted, the test material will be classified as corrosive to the eye.

 

Table1               IndividualScores and Individual Total Scores for Ocular Irritation

Rabbit Number and Sex

72298 Male

72344 Male

IPR= 2

IPR = 2

Time After Treatment

1
Hour

24
Hours

48
Hours

72
Hours

1
Hour

24
Hours

48
Hours

72
Hours

CORNEA

 

 

 

 

 

 

 

 

E = Degree of Opacity

0

0

0

0

0

0

0

0

F = Area of Cornea Involved

0

0

0

0

0

0

0

0

Score (E x F) x 5

0

0

0

0

0

0

0

0

IRIS

 

 

 

 

 

 

 

 

D

0

0

0

0

1

0

0

0

Score (D x 5)

0

0

0

0

5

0

0

0

CONJUNCTIVAE

 

 

 

 

 

 

 

 

A = Redness

2

1

0

0

2

2

1

0

B = Chemosis

1

0

0

0

2

1

0

0

C = Discharge

1

0

0

0

2

0

0

0

Score (A + B + C) x 2

8

2

0

0

12

6

2

0

Total Score

8

2

0

0

17

6

2

0

 

IPR=  Initial pain reaction

 

Table 2               Individual Total Scores and Group Mean Scores for Ocular Irritation

 

Rabbit Number

and Sex

Individual Total Scores At:

 

1 Hour

24 Hours

48 Hours

72 Hours

 

72298 Male

8

2

0

0

 

72344 Male

17

6

2

0

 

Group Total

25

8

2

0

 

Group Mean Score

12.5

4.0

1.0

0.0

 

 

 

Interpretation of results:
not irritating
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
The test material did not meet the criteria for classification as irritant according to Regulation (EC) No. 1272/2008 (EU CLP). This study is conducted according to the appropriate guidelines (OECD 405 ) and under the conditions of GLP and therefore the study is considered to be acceptable and to adequately satisfy both the guideline requirement and the regulatory requirement as a key study for this endpoint. Study is sufficient for classification and labelling purposes, in accordance with Regulation (EC) No. 1272/2008 (EU CLP).
Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (not irritating)

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Justification for selection of skin irritation / corrosion endpoint:
This study has been selected as the key study in accordance with Regulation (EC) No. 1907/2006 (REACH) because the results are sufficient in order to derive a reliable conclusion on classification and labelling in accordance with Regulation (EC) No. 1272/2008 (EU CLP). No effects are noted.

Justification for selection of eye irritation endpoint:
This study has been selected as the key study in accordance with Regulation (EC) No. 1907/2006 (REACH) because the results are sufficient in order to derive a reliable conclusion on classification and labelling in accordance with Regulation (EC) No. 1272/2008 (EU CLP). No effects are noted.

Justification for classification or non-classification

No classification is proposed for skin or eye irritancy of tetrairon tris(pyrophosphate) in accordance with Regulation (EC) No. 1272/2008 (EU CLP). This conclusion is based on reliable (Klimisch 1) studies and adequate data and as such no further testing is anticipated.