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EC number: 229-856-5 | CAS number: 6789-88-4
- 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
- Nanomaterial porosity
- Nanomaterial pour density
- Nanomaterial photocatalytic activity
- Nanomaterial radical formation potential
- Nanomaterial catalytic activity
- Endpoint summary
- Stability
- Biodegradation
- Bioaccumulation
- Transport and distribution
- Environmental data
- Additional information on environmental fate and behaviour
- 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
- Toxicity to other aquatic organisms
- Sediment toxicity
- Terrestrial toxicity
- Biological effects monitoring
- Biotransformation and kinetics
- Additional ecotoxological information
- Toxicological Summary
- Toxicokinetics, metabolism and distribution
- Acute Toxicity
- Irritation / corrosion
- Sensitisation
- 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
Endpoint summary
Administrative data
Description of key information
Skin irritation (OECD TG 439): Not irritating
Eye irritation (OECD TG 438): Not irritating
Key value for chemical safety assessment
Skin irritation / corrosion
Link to relevant study records
- Endpoint:
- skin irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 07 August 2017 - 14 August 2017
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)
- Version / remarks:
- 28 July 2015
- Deviations:
- no
- Qualifier:
- according to guideline
- Guideline:
- EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)
- Version / remarks:
- 20 July 2012
- Deviations:
- no
- GLP compliance:
- yes
- Test system:
- human skin model
- Source species:
- human
- Cell type:
- other: epidermal keratinocytes
- Cell source:
- other: SkinEthic Laboratories, Lyon, France.
- Source strain:
- other: Not applicable
- Justification for test system used:
- In the interest of sound science and animal welfare, a sequential testing strategy is recommended to minimise the need of in vivo testing. One of the validated in vitro skin irritation tests is the EPISKIN test, which is recommended in international guidelines (e.g. OECD and EC).
- Vehicle:
- unchanged (no vehicle)
- Details on test system:
- TEST SYSTEM
- EPISKIN Small Model (TM) (EPISKIN-SM (TM), 0.38 cm^2, Lot no.: 17-EKIN-032); a three-dimensional human epidermis model, which consists of adult human-derived epidermal keratinocytes which have been seeded in 12-well plates on a dermal substitute consisting of a collagen type I matrix coated with type IV collagen and cultured for 13 days.
SKIN DISC PREPARATION
- Procedure used: On the day of receipt the tissues were transferred to 12-well plates and preincubated with prewarmed Maintenance Medium for 2.5 hours at 37°C
ENVIRONMENTAL CONDITIONS
- Temperature used during treatment / exposure: room temperature
- Temperature of post-treatment incubation: 36.6-37.5 °C
- Humidity (%): 74-90
REMOVAL OF TEST MATERIAL AND CONTROLS
- Number of washing steps: 1, with phosphate buffered saline
- Observable damage in the tissue due to washing: no
DYE BINDING METHOD
- Dye used in the dye-binding assay: MTT
- Incubation time: 3 hours
- Spectrophotometer: TECAN Infinite® M200 Pro Plate Reader.
- Wavelength: 570 nm
- Interaction with MTT was tested before the study by adding 10 mg of the test item to 2 mL of MTT solution. The mixture was incubated for 3 hours at 37°C. At the end of the incubation period a color check was performed.
PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritant to skin if the relative mean tissue viability of three individual tissues after 15 minutes of exposure to the test item and 42 hours of post incubation is ≤ 50% of the mean viability of the negative controls.
- The test substance is considered to be non-irritant to skin if the relative mean tissue viability of three individual tissues after 15 minutes of exposure to the test item and 42 hours of post incubation is > 50% of the mean viability of the negative controls.
SCORING SYSTEM:
- Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity measured by formazan production from MTT at the end of the treatment.
- Cell viability: The corrected OD (ODc) for each sample or control was calculated by subtracting the value of the blank mean (ODbl) from each reading (ODraw): ODc = ODraw – ODbl
The OD value representing 100% cell viability is the average OD of the negative controls (ODlt_u+MTT).
The % viability for each sample and the positive control is calculated as follows: %Viability = (ODc/mean ODlt_u+MTT) * 100
- Skin irritation is expressed as the remaining cell viability after exposure to the test item.
ACCEPTABILITY CRITERIA:
a) The absolute mean OD570 (optical density at 570 nm) of the three tissues of the negative control should reasonably be within the laboratory historical control data range and the Standard Deviation value (SD) of the % viability should be ≤18.
b) The mean relative tissue viability of the positive control should be ≤40% relative to the negative control and the Standard Deviation value (SD) of the % viability should be ≤18.
c) The SD calculated from individual % tissue viabilities of the three identically treated replicates should be ≤18. - Control samples:
- yes, concurrent negative control
- yes, concurrent positive control
- Amount/concentration applied:
- TEST ITEM:
25 μL directly on top of the tissue
NEGATIVE CONTOL:
- Amount applied: 25 µl Phosphate buffered saline
POSITIVE CONTROL
- Amount applied: 25 µl
- Concentration: 5% (aq) Sodium dodecyl sulphate
- Re-spread after 7 minutes contact time - Duration of treatment / exposure:
- 15 ±0.5 minutes
- Duration of post-treatment incubation (if applicable):
- 42 hours and 3 hours with MTT
- Number of replicates:
- 3 for the test item, the negative and the positive control, each.
- Irritation / corrosion parameter:
- % tissue viability
- Value:
- 94
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks:
- Cell viability: 4.8%
- Other effects / acceptance of results:
- - OTHER EFFECTS:
- Visible damage on test system: no
- Direct-MTT reduction: no
- Colour interference with MTT: no
ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: yes, the absolute mean OD570 of the three tissues of the negative control were within the laboratory historical control data range (i.e., a mean of 1.234 ± 0.016) and the SD of the % viability was <18% (i.e., 1.3%)
- Acceptance criteria met for positive control: yes, the mean relative tissue viability of the positive control was <50% (i.e., 4.8%) and the SD of the % viability was <18% (i.e., 2.3%).
- Acceptance criteria met for variability between replicate measurements: yes, the SD calculated from individual % tissue viabilities of the three identically treated replicates was <18% (i.e., 2.9%).
- Since the mean relative tissue viability for the substance was above 50%, it is considered to be non-irritant. - Interpretation of results:
- other: Not a skin irritant
- Remarks:
- According to Regulation (EC) No. 1272/2008 and its amendments.
- Conclusions:
- In the in vitro skin irritation test conducted according to OECD 439 guideline and GLP principles the substance was not irritating.
- Executive summary:
- The possible skin irritation potential of the substance was tested in vitro using a human skin model through topical application for 15 minutes. The study procedures described in this report were according to OECD TG 439 guideline and GLP principles. Skin tissue was treated by topical application of 25 μL undiluted test substance. After 42 hours 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 (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) MTT at the end of treatment. Skin irritation is expressed as the remaining cell viability after exposure to the test substance. Reliable negative and positive controls were included. All validity criteria were met and the study was considered to be valid. The positive control had a mean cell viability of 4.8% after 15 minutes exposure. The standard deviation value of the percentage viability of three tissues treated identically was less than 2.9%, indicating that the test system functioned properly. The relative mean tissue viability obtained after 15 minutes treatment with the substance compared to the negative control tissue was 94%. Since the mean relative tissue viability for the substance was above 50% after 15 minutes treatment the substance is considered to be non-irritant to the skin.
- Endpoint:
- skin corrosion: in vitro / ex vivo
- Data waiving:
- study scientifically not necessary / other information available
- Justification for data waiving:
- other:
Referenceopen allclose all
Table 2 Individual OD measurements (570 nm)
|
A (OD570) |
B (OD570) |
C (OD570) |
Negative control OD570 measurement 1 OD570 measurement 2 |
1.2719 1.2577 |
1.3013 1.2371 |
1.3201 1.2682 |
Test item OD570 measurement 1 OD570 measurement 2 |
1.1583 1.1649 |
1.2045 1.1867 |
1.3262 1.1400 |
Positive control OD570 measurement 1 OD570 measurement 2 |
0.0730 0.0751 |
0.1377 0.1216 |
0.1040 0.0990 |
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (not irritating)
Eye irritation
Link to relevant study records
- Endpoint:
- eye irritation: in vitro / ex vivo
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 04 Aug 2017 to 18 Aug 2017
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 438 (Isolated Chicken Eye Test Method for Identifying i) Chemicals Inducing Serious Eye Damage and ii) Chemicals Not Requiring Classification for Eye Irritation or Serious Eye Damage)
- Version / remarks:
- 26 July 2013
- GLP compliance:
- yes (incl. QA statement)
- Remarks:
- Triskelion B.V., Utrechtseweg 48, 3700 AV, Zeist
- Species:
- other: Eyes of male or female chickens (ROSS, spring chickens)
- Details on test animals or tissues and environmental conditions:
- SOURCE OF COLLECTED EYES
- Source: Slaughterhouse v.d. Bor, Nijkerkerveen, The Netherlands
- Characteristics of donor animals: Approximately 7 weeks old, male or female chickens, body weight range approximately 1.5-2.5 kg, were used as eye donors.
- Storage, temperature and transport conditions of ocular tissue: Heads of the animals were cut off immediately after sedation of the animals by electric shock and incision of the neck for bleeding, and before they reached the next station on the process line. The heads were placed in small plastic boxes on a bedding of paper tissues moistened with isotonic saline. Next, they were transported to the testing facility. During transportation, the heads were kept at ambient temperature.
- Time interval prior to initiating testing: Within 2 hours after kill, eyes were carefully dissected and placed in a superfusion apparatus.
- Indication of any existing defects or lesions in ocular tissue samples: No
- Indication of any antibiotics used: No - Vehicle:
- unchanged (no vehicle)
- Controls:
- yes, concurrent positive control
- yes, concurrent negative control
- Amount / concentration applied:
- 30 μL neat substance
- Duration of treatment / exposure:
- 10 seconds
- Duration of post- treatment incubation (in vitro):
- 240 minutes
- Number of animals or in vitro replicates:
- Test group and positive control: triplicates
Negative control: Singlo - Details on study design:
- SELECTION AND PREPARATION OF ISOLATED EYES
Within 2 hours after kill, eyes were carefully dissected and placed in a superfusion apparatus using the following procedure: First the eye-lids were carefully removed without damaging the cornea and a small drop of Fluorescein sodium 2.0% w/v was applied to the corneal surface for a few seconds and subsequently rinsed off with isotonic saline at ambient temperature. Next, the head with the fluorescein-treated cornea was examined with a slit-lamp microscope (Slit-lamp 900 BP, Haag-Streit AG, Liebefeld-Bern, Switzerland) to ensure that the cornea was not damaged. If undamaged (e.g., fluorescein retention and corneal opacity scores of ≤ 0.5), the eye was further dissected from the head without damaging the eye or cornea. Care was taken to remove the eye-ball from the orbit without cutting off the optical nerve too short. The enucleated eye was placed in a stainless steel clamp with the cornea positioned vertically and transferred to a chamber of the superfusion apparatus. The clamp holding the eye was positioned in such a way that the entire cornea was supplied with isotonic saline from a bent, stainless steel tube, at a target rate of 0.10-0.15 mL/min. The chambers of the superfusion apparatus as well as the saline were temperature controlled at approximately 32 °C (water pump set at 36.4 °C). After placing in the superfusion apparatus, the eyes were examined again with the slit-lamp microscope to ensure that they were not damaged. An accurate measurement was taken at the corneal apex of each eye. Eyes with a corneal thickness deviating more than 10% of the average corneal thickness of the eyes, eyes showing opacity (score higher than 0.5), or were unacceptably stained with fluorescein (score higher than 0.5) indicating the cornea to be permeable, or eyes that showed any other signs of damage, were rejected as test eyes and replaced.
EQUILIBRATION AND BASELINE RECORDINGS
Each eye provided its own baseline values for corneal swelling, corneal opacity and fluorescein retention. For that purpose, after an equilibration period of 45-60 minutes, the corneal thickness of the eyes was measured again to determine the zero reference value for corneal swelling calculations.
REMOVAL OF TEST SUBSTANCE
- Volume and washing procedure after exposure period: 20 mL saline. After rinsing, each eye in the holder was returned to its chamber.
- Indicate any deviation from test procedure in the Guideline: none
METHODS FOR MEASURED ENDPOINTS:
- Corneal opacity: Slit-lamp microscope examination
- Damage to epithelium based on fluorescein retention: Slit-lamp microscope examination
- Swelling: measured with optical pachymeter on a slit-lamp microscope; slit-width setting: set at 0.095 mm
- Others: After the final examination, the test substance treated eyes, the negative and positive control eyes were preserved in a neutral aqueous phosphate-buffered 4% solution of formaldehyde. The corneas were embedded in paraffin wax, sectioned at ca 4 μm and stained with PAS (Periodic Acid-Schiff). The microscopic slides were subjected to histopathological examination.
SCORING SYSTEM:
Defined scoring scales were used for each parameter to define the severity of effects into four categories (I-IV).
- Mean corneal swelling (%): According to OECD 438 guideline. Examination of the eyes after 0, 30, 75, 120, 180, and 240 minutes
- Mean maximum opacity score: According to OECD 438 guideline. Examination of the eyes after 0, 30, 75, 120, 180, and 240 minutes
- Mean fluorescein retention score at 30 minutes post-treatment: According to OECD 438 guideline.
DECISION CRITERIA: According to OECD 438 guideline - Irritation parameter:
- percent corneal swelling
- Run / experiment:
- slit-lamp examination
- Value:
- 0
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: maximum mean values
- Irritation parameter:
- cornea opacity score
- Run / experiment:
- slit-lamp examination
- Value:
- 0
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: maximum mean values
- Irritation parameter:
- fluorescein retention score
- Run / experiment:
- slit-lamp examination
- Value:
- 0
- Vehicle controls validity:
- not applicable
- Negative controls validity:
- valid
- Positive controls validity:
- valid
- Remarks on result:
- other: maximum mean values
- Other effects / acceptance of results:
- Slit-lamp examination: The test substance did not cause any corneal effects. The negative control eye did not show any corneal effect and demonstrated that the general conditions during the tests were adequate. The positive control BAC 5% caused severe corneal effects and demonstrated the ICE test valid to detect severe eye irritants.
Microscopic examination of the corneas revealed very slight erosion of the epithelium in all three corneas. Microscopic examination of the cornea treated with the negative control (saline) did not reveal any abnormalities as well. Microscopic examination of the corneas treated with the positive control BAC 5% generally revealed severe erosion and very slight vacuolation of the epithelium. - Interpretation of results:
- other: Not an eye irritant.
- Remarks:
- According to Regulation (EC) No. 1272/2008 and its amendments.
- Conclusions:
- Under the test conditions (OECD 438 and GLP) the test substance is considered not considered to be an eye irritant.
- Executive summary:
The test substance was examined for its in vitro eye irritating potential using the Isolated Chicken Eye (ICE) test, in accordance with OECD guideline 438 and GLP. In the ICE test, 3 eyes were exposed to 30 µL test substance for 10 seconds. In addition, one negative control eye (30 µL saline) and three positive control eyes (30 µL 5% Benzalkonium Chloride (BAC)) were tested. After the exposure, the eyes were rinsed with 20 mL saline and were examined at approximately 0, 30, 75, 120, 180, and 240 minutes after treatment. Mean fluorescein retention score was determined at 30 minutes post-treatment. The test substance did not cause any corneal effects. The negative control eye did not show any corneal effect and demonstrated that the general conditions during the tests were adequate. The positive control BAC 5% caused severe corneal effects and demonstrated the ICE test valid to detect severe eye irritants. Microscopic examination of the corneas treated with the test substance revealed very slight erosion of the epithelium in all three corneas. Microscopic examination of the cornea treated with the negative control (saline) did not reveal any abnormalities as well. Microscopic examination of the corneas treated with the positive control BAC 5% generally revealed severe erosion and very slight vacuolation of the epithelium. Based on these results, the substance is not considered to be irritating to the eyes.
Reference
Endpoint conclusion
- Endpoint conclusion:
- no adverse effect observed (not irritating)
Additional information
In vitro skin irritation:
The possible skin irritation potential of the substance was tested in vitro using a human skin model through topical application for 15 minutes. The study procedures described in this report were according to OECD TG 439 guideline and GLP principles. Skin tissue was treated by topical application of 25 μL undiluted test substance. After 42 hours 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 (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide) MTT at the end of treatment. Skin irritation is expressed as the remaining cell viability after exposure to the test substance. Reliable negative and positive controls were included. All validity criteria were met and the study was considered to be valid. The positive control had a mean cell viability of 4.8% after 15 minutes exposure. The standard deviation value of the percentage viability of three tissues treated identically was less than 2.9%, indicating that the test system functioned properly. The relative mean tissue viability obtained after 15 minutes treatment with the substance compared to the negative control tissue was 94%. Since the mean relative tissue viability for the substance was above 50% after 15 minutes treatment the substance is considered to be non-irritant to the skin.
In vitro eye irritation:
The test substance was examined for its in vitro eye irritating potential using the Isolated Chicken Eye (ICE) test, in accordance with OECD guideline 438 and GLP. In the ICE test, 3 eyes were exposed to 30 µL test substance for 10 seconds. In addition, one negative control eye (30 µL saline) and three positive control eyes (30 µL 5% Benzalkonium Chloride (BAC)) were tested. After the exposure, the eyes were rinsed with 20 mL saline and were examined at approximately 0, 30, 75, 120, 180, and 240 minutes after treatment. Mean fluorescein retention score was determined at 30 minutes post-treatment. The test substance did not cause any corneal effects. The negative control eye did not show any corneal effect and demonstrated that the general conditions during the tests were adequate. The positive control BAC 5% caused severe corneal effects and demonstrated the ICE test valid to detect severe eye irritants. Microscopic examination of the corneas treated with the test substance revealed very slight erosion of the epithelium in all three corneas. Microscopic examination of the cornea treated with the negative control (saline) did not reveal any abnormalities as well. Microscopic examination of the corneas treated with the positive control BAC 5% generally revealed severe erosion and very slight vacuolation of the epithelium. Based on these results, the substance is not considered to be irritating to the eyes.
Skin corrosion: the substance is not corrosive in absence of skin and eye irritation.
Justification for classification or non-classification
Based on the results, the substance does not need to be classified for skin irritation/corrosion and eye irritation according to Regulation (EC) No. 1272/2008 and its amendments.
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