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Administrative data

Description of key information

- Skin irritation/corrosion study: based on the results of in vitro skin irritation study (OECD 431, reliability: 1), the Reaction Mass of "Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate" is considered as corrosive.
- Eye irritation: based on the corrosivity of submitted substance, a waiving was proposed.
- Respiratory irritation: no study was available

Key value for chemical safety assessment

Skin irritation / corrosion

Link to relevant study records
Reference
Endpoint:
skin corrosion: in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
The study was performed between 06 March 2012 and 08 March 2012.
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study
Qualifier:
according to
Guideline:
other: OECD guideline for the testing of chemicals 431
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.40 (In Vitro Skin Corrosion: Transcutaneous Electrical Resistance Test (TER))
Deviations:
no
Principles of method if other than guideline:
not applicable
GLP compliance:
yes (incl. certificate)
Remarks:
2011-08-31
Test system:
human skin model
Source species:
human
Cell type:
other: reconstruted Human Epidermis
Cell source:
other: Culture
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 item was applied neat.

- Amount(s) applied (volume or weight with unit):
50 µl of the test item was applied topically to the corresponding tissues ensuring uniform coverage of the tissues.

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

VEHICLE
No vehicle used
Duration of treatment / exposure:
3, 60 or 240 minutes
Observation period:
Not applicable
Number of animals:
Not applicable
Details on study design:
TEST SITE
- Area of exposure:
50 µl of the test item was applied topically to the corresponding tissues ensuring uniform coverage of the tissues.

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

REMOVAL OF TEST ITEM
- Washing (if done):
At the end of each exposure period, each tissue was removed from the well using forceps and rinsed using a wash bottle containing Phosphate Buffered Saline Dulbeccos (PBS) 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 item.

- Time after start of exposure: 3, 60 or 240 minutes

SCORING SYSTEM:
Quantitative MTT Assessment (percentage tissue viability)
The corrosivity potential of the test item was predicted from the relative mean tissue viabilities obtained after the 3, 60 and 240-minute treatments, compared to the mean of the negative control tissues. 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)
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
3 minutes
Value:
103.7
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
60 minutes
Value:
11.1
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritation / corrosion parameter:
% tissue viability
Run / experiment:
240 minutes
Value:
4.1
Vehicle controls validity:
valid
Negative controls validity:
valid
Positive controls validity:
valid
Irritant / corrosive response data:
The relative mean viability of the test material treated tissues was as follows:
240 minutes exposure: 4.1%
60 minutes exposure: 11.1%
3 minutes exposure: 103.7%
Other effects:
No

Direct MTT Reduction:

The MTT solution containing the test item did not turn blue. This was taken to indicate the test item did not reduce MTT.

Quality Criteria:

The relative mean tissue viability for the positive control treated tissues was 8.3% relative to the negative control treated tissues following the 240-minute exposure period. The positive control acceptance criterion was therefore satisfied. The mean OD540for the negative control treated tissues was 0.217. The negative control acceptance criterion was therefore satisfied.

Table 7.3.1/1:Mean OD540Values and Viabilities for the Negative Control Item, Positive Control Item and Test Item

Item

Exposure Period

Mean OD540of duplicate tissues

Relative mean viability (%)

Negative Control Item

240 Minutes

0.217

100*

Positive Control Item

240 Minutes

0.018

8.3

Test Item

240 Minutes

0.009

4.1

60 Minutes

0.024

11.1

3 Minutes

0.225

103.7

*= The mean viability of the negative control tissues is set at 100% Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate (IBAP)

Interpretation of results:
Category 1B (corrosive) based on GHS criteria
Remarks:
Migrated information Criteria used for interpretation of results: EU
Conclusions:
As viability of cells is higher than 35% after 3 minutes exposure but lower than 35% after 1 hour exposure the test item is classified as corrosive to the skin, Skin Corr. 1B (H314) according to the Regulation (EC) 1272/2008 (CLP) and as C, R34 according to the Directive 67/548/EEC under the test conditions of this study.
Executive summary:

In an in vitro skin corrosion study performed according to the OECD guideline 431 and in compliance with the GLP, Reaction Mass of "Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate" (purity 96%) was applied undiluted to Reconstructed Human Epidermis (RHE) model (Episkin TM model kit 0.38cm²).

Duplicate tissues were treated with the test item for exposure periods of 3, 60 and 240 minutes. At the end of the exposure period each tissue was rinsed from the test item before each tissue was taken for MTT-loading. After MTT loading a total biopsy of each epidermis was made and placed into micro tubes containing acidified isopropanol for extraction of formazan crystals out of the MTT-loaded tissues.  At the end of the formazan extraction period each tube was mixed thoroughly and duplicate 200 µl samples were transferred to the appropriate wells of a pre‑labelled 96‑well plate. The optical density (OD) was measured at 540 nm (OD540). 0.9 % (w/v) Sodium Chloride Solution was used as the negative control. Glacial Acetic Acid was used as the positive control.

The relative mean tissue viability for the positive control treated tissues was 8.3% relative to the negative control treated tissues following the 240-minute exposure period. The positive control acceptance criterion was therefore satisfied. The mean OD540for the negative control treated tissues was 0.217. The negative control acceptance criterion was therefore satisfied.

Percentage of cell viability (MTT reduction in the test item treated tissues relative to negative control tissues) was determined for each exposure period. The relative mean viability of the test item treated tissues was 103.7; 11.1 and 4.1% for exposure period of 3, 60 and 240 min, respectively.

As viability of cells is higher than 35% after 3 minutes exposure but lower than 35% after 1 hour exposure the Reaction Mass of "Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate" is classified as corrosive to the skin, Skin Corr. 1B (H314) according to the Regulation (EC) 1272/2008 (CLP) and as C, R34 according to the Directive 67/548/EEC.

This study is considered as acceptable as it satisfies the main criteria of OECD guideline No. 431.

Endpoint conclusion
Endpoint conclusion:
adverse effect observed (corrosive)

Eye irritation

Link to relevant study records
Reference
Endpoint:
eye irritation: in vitro / ex vivo
Data waiving:
study scientifically not necessary / other information available
Justification for data waiving:
the study need not be conducted because the available information indicates that the criteria are met for classification as corrosive to the skin or irritating to eyes
Endpoint conclusion
Endpoint conclusion:
no study available

Respiratory irritation

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

- Skin irritation/corrosion study:

One study was available and was considered as the key study (OECD 431, reliability: 1).

In this study, the Reaction Mass of "Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate" (purity 96%) was applied undiluted to Reconstructed Human Epidermis (RHE) model (Episkin TM model kit0.38cm²). Duplicate tissues were treated with the test item for exposure periods of 3, 60 and 240 minutes. At the end of the exposure period each tissue was rinsed from the test item before each tissue was taken for MTT-loading. After MTT loading a total biopsy of each epidermis was made and placed into micro tubes containing acidified isopropanol for extraction of formazan crystals out of the MTT-loaded tissues.  At the end of the formazan extraction period each tube was mixed thoroughly and duplicate 200 µl samples were transferred to the appropriate wells of a pre‑labelled 96‑well plate. The optical density (OD) was measured at 540 nm (OD540). 0.9 % (w/v) Sodium Chloride Solution was used as the negative control. Glacial Acetic Acid was used as the positive control.

The relative mean tissue viability for the positive control treated tissues was 8.3% relative to the negative control treated tissues following the 240-minute exposure period. The positive control acceptance criterion was therefore satisfied. The mean OD540 for the negative control treated tissues was 0.217. The negative control acceptance criterion was therefore satisfied.

Percentage of cell viability (MTT reduction in the test item treated tissues relative to negative control tissues) was determined for each exposure period. The relative mean viability of the test item treated tissues was 103.7; 11.1 and 4.1% for exposure period of 3, 60 and 240 min, respectively.

As viability of cells is higher than 35% after 3 minutes exposure but lower than 35% after 1 hour exposure the Reaction Mass of "Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate" is classified as corrosive to the skin into category 1B, H314.

- Eye irritation study:

No data was available. However, as the Reaction mass of “Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate" is classified as corrosive, no further test has to be conducted (Guidance on the application of Regulation (EC) No 1272/2008). A skin corrosive substance is considered to also cause serious eye damage which is indicated in the hazard statement for skin corrosion (H314: causes severe skin burns and eye damage).

 

- Respiratory irritation study:

No data was available.


Justification for selection of skin irritation / corrosion endpoint:
only one study available

Justification for selection of eye irritation endpoint:
As the Reaction mass of “ Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate" is classified as corrosive, no further test has to be conducted (Guidance on the application of Regulation (EC) No 1272/2008). A skin corrosive substance is considered to also cause serious eye damage which is indicated in the hazard statement for skin corrosion (H314: causes severe skin burns and eye damage).

Effects on skin irritation/corrosion: corrosive

Justification for classification or non-classification

Skin Irritation/Corrosion:

Based on the above data, the Reaction Mass of "Diisobutyl hydrogen phosphate and Isobutyl dihydrogen phosphate" is classified as corrosive into category 1B (H314; causes severe skin burns and eye damage) according to the criteria of the Regulation (EC) 1272/2008 (CLP) and as skin corrosive (C; R34 Causes burns) according to the criteria of the annex VI of the Directive 67/548/EEC.

Eye irritation:

The classification as Skin Corr. 1B (H314) or C, R34 implies that the substance is corrosive to the skin and also to the eyes. Therefore an additional classification for the eyes irritation is not necessary.

Respiratory irritation:

No study is available; therefore no classification is possible due to lack of data.