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EC number: 911-811-2 | CAS number: -
- Life Cycle description
- Uses advised against
- Endpoint summary
- Appearance / physical state / colour
- Melting point / freezing point
- Boiling point
- Density
- Particle size distribution (Granulometry)
- Vapour pressure
- Partition coefficient
- Water solubility
- Solubility in organic solvents / fat solubility
- Surface tension
- Flash point
- Auto flammability
- Flammability
- Explosiveness
- Oxidising properties
- Oxidation reduction potential
- Stability in organic solvents and identity of relevant degradation products
- Storage stability and reactivity towards container material
- Stability: thermal, sunlight, metals
- pH
- Dissociation constant
- Viscosity
- Additional physico-chemical information
- Additional physico-chemical properties of nanomaterials
- Nanomaterial agglomeration / aggregation
- Nanomaterial crystalline phase
- Nanomaterial crystallite and grain size
- Nanomaterial aspect ratio / shape
- Nanomaterial specific surface area
- Nanomaterial Zeta potential
- Nanomaterial surface chemistry
- Nanomaterial dustiness
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- Endpoint summary
- Stability
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- Environmental data
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- Ecotoxicological Summary
- Aquatic toxicity
- Endpoint summary
- Short-term toxicity to fish
- Long-term toxicity to fish
- Short-term toxicity to aquatic invertebrates
- Long-term toxicity to aquatic invertebrates
- Toxicity to aquatic algae and cyanobacteria
- Toxicity to aquatic plants other than algae
- Toxicity to microorganisms
- Endocrine disrupter testing in aquatic vertebrates – in vivo
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- Sediment toxicity
- Terrestrial toxicity
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- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
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- Repeated dose toxicity
- Genetic toxicity
- Carcinogenicity
- Toxicity to reproduction
- Specific investigations
- Exposure related observations in humans
- Toxic effects on livestock and pets
- Additional toxicological data

Skin irritation / corrosion
Administrative data
- Endpoint:
- skin corrosion: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 2014-08-19 to 2014-08-28
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- other: The study was conducted to the appropriate OECD test guideline and in compliance with GLP
Data source
Reference
- Reference Type:
- study report
- Title:
- Unnamed
- Year:
- 2 015
- Report date:
- 2015
Materials and methods
Test guidelineopen allclose all
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)
- Version / remarks:
- 2008
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.4 (Acute Toxicity: Dermal Irritation / Corrosion)
- Version / remarks:
- 2008
- GLP compliance:
- yes (incl. QA statement)
Test material
- Reference substance name:
- Reaction Mass of [[(2-hydroxyethyl)imino]dimethylene]bisphosphonic acid and [(2-hydroxy-2-oxo-1,4,2-oxazaphosphinan-4-yl)methyl]phosphonic acid
- EC Number:
- 911-811-2
- Molecular formula:
- C4H13NO7P2 and C4H11NO6P2
- IUPAC Name:
- Reaction Mass of [[(2-hydroxyethyl)imino]dimethylene]bisphosphonic acid and [(2-hydroxy-2-oxo-1,4,2-oxazaphosphinan-4-yl)methyl]phosphonic acid
- Test material form:
- other: liquid at room temperature
Constituent 1
Test animals
- Species:
- human
- Strain:
- other: not relevant
- Details on test animals or test system and environmental conditions:
- - Source: Episkin, Lyon, France
Test system
- Type of coverage:
- other: not relevant
- Preparation of test site:
- other: not relevant
- Vehicle:
- unchanged (no vehicle)
- Controls:
- other: not relevant
- Amount / concentration applied:
- TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 50 µL - Duration of treatment / exposure:
- not relevant
- Observation period:
- not relevant
- Number of animals:
- not relevant
- Details on study design:
- Preliminary tests:
1. Test for direct MTT reduction with the test item:
To identify any test item interference, 50 µl of test item was added to 2 ml of a 0.3 mg/ml freshly prepared MTT solution. The mixture was incubated in darkness at ±37°C for 3 hours (±5 minutes) under continuous stirring.
A negative control was tested concurrently by adding 50 µl of water in place of the test solution, in the same manner.
The colour of both solutions was observed.
2. Test for the detection of the colouring potential of the test item:
10 µl of test item was added to 90 µl of water for injection in a transparent recipient. After 1 hour of mixing, the colouration was checked.
Main test:
On 12-well plate was used for each of the 3 exposure times (3 minutes, 1 hour and 4 hours) for test item-treated tissues.
Positive (glacial acetic acid) and negative (0.9% NaCl) controls were placed on separate plates.
Test item, and negative and positive controls were applied on duplicate tissues.
1. Pre-incubation of the tissues on day 0:
2 ml of 37°C maintenance medium was added to 2 wells per plate as follows:
- duplicate wells for each test item and negative control exposure time and for the 4-hour positive control exposure time.
One Episkin tissue was transferred into each maintenance medium pre-filled well.
All 12-well plates were incubated at 37°C, 5% CO2, >95% humidity for 1-48 h pre-incubation.
2. Treatment of tissues:
2 ml of 37°C assay medium was added to 2 wells per 12-well plate as follows:
- duplicate wells for each test item and negative control exposure time and for the 4-hour positive control exposure time.
The tissues were removed from the incubator and one tissue was transferred in to each assay medium pre-filled well.
The test item and controls were applied on each designated tissue.
The lids were replaced on each plate before incubation at room temperature as follows:
- positive control for 4 hours (±10 minutes)
-test item and negative control for 3 minutes (±5 seconds), 1 hours (±5 minutes) and 4 hours (±10 minutes)
3. Rinsing of tissues:
All treated tissues were rinsed with D-PBS at the end of the designated incubation period
4. MTT viability assay:
Two empty wells were filled with 2 ml MTT solution (0.3 mg/ml), and the corresponding tissues were placed in these wells. Each plate was protected from light and incubated for 3 hours ±15 minutes at 37°C, 5% CO2 in a humidified incubator.
After incubation, the underside of each tissue culture insert was blotted. The tissues were removed using a biopsy punch. Any tissue discolouration was evaluated with the naked eye.
For each tissue, the epidermis was separated from the collagen matrix. Both parts were put in a stoppered plastic tube and 0.5ml acidified isopropanol were added. After vortexing, each tube was protected from light and left at room temperature overnight to extract the formazan (reduced MTT).
5. Optical density measurements:
After the extraction period, each tube was vortexed, and centrifuged if needed. Each tube was used to fill 2 consecutive wells of a 96-well plate with 200 µl if extract per well. A separate 96-well plate was used for the test item dose formulation. For each plate, the OD value of 4 wells containing 200 µl of acidified isoproponal only was used as the blank.
The OD was measured at a wavelength of 570 nm.
Results and discussion
In vitro
Resultsopen allclose all
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 4 hours exposure
- Value:
- 82
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 1 hour exposure
- Value:
- 84
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Irritation / corrosion parameter:
- % tissue viability
- Run / experiment:
- 3 minutes exposure
- Value:
- 104
- Vehicle controls validity:
- valid
- Negative controls validity:
- valid
- Positive controls validity:
- valid
Any other information on results incl. tables
The mixture of 50 µL test item per 2 mL MTT solution showed no reduction of MTT compared to the negative control (0.9% NaCl). The mixture did not turn blue/purple.
The mixture of 10 µL of the test item per 90 µL water showed no colouring and the test item was presumed to not stain the tissue.
Following the 3 minute exposure period, a blue discolouration of the test item-treated tissues was noted. The discolouration was representative of viable tissues. Following the 1 and 4 hour exposure periods, a blue discolouration was noted on one tissue and a blue/white discolouration on the second tissue. This discolouration was representative of viable or semi-viable tissues, respectively. The controls confirmed the validity of the study.
The relative mean viabilities of the test item-treated tissues were: - 104% for the 3 minutes exposure, - 84 % for the 1 hour exposure, - 82% for the 4 hour exposure. All mean viabilities were ≥35%. Table 1: Mean tissue viability, standard deviations and difference (%) between replicate tissues for the test item, the negative and positive controls
Group |
Exposure duration |
cOD570 nm |
Viability (%) |
|||
Mean |
SD |
Mean |
SD |
Difference (%) |
||
Negative control |
3 min |
1.068 |
0.023 |
100 |
2 |
3 |
1 h |
1.057 |
0.015 |
100 |
1 |
2 |
|
4 h |
1.138 |
0.051 |
100 |
4 |
6 |
|
Positive control |
4 h |
0.027 |
0.006 |
2 |
1 |
n.c. |
Test item |
3 min |
1.112 |
0.011 |
104 |
1 |
1 |
1 h |
0.884 |
0.293 |
84 |
28 |
39 |
|
4 h |
0.935 |
0.129 |
82 |
11 |
16 |
cOD = blank corrected optical density
n.c. = not calculated
SD = standard deviation
Applicant's summary and conclusion
- Interpretation of results:
- GHS criteria not met
- Conclusions:
- In an in vitro Human Skin Model Test (EPISKIN-SM™) conducted to OECD TG 431 and in compliance with GLP, the mean relative tissue viability (% negative control) was ≥35% after 4-hour exposure for [[(2-hydroxyethyl)imino]bis(methylene)]bisphosphonic acid. This result is negative for skin corrosivity potential.
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