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Toxicological information

Skin irritation / corrosion

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Administrative data

Endpoint:
skin irritation: in vitro / ex vivo
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
28 Jan - 31 Jan 2010
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Study of a draft guideline with acceptable restrictions (limited documentation).

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2010
Report Date:
2010

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
other: OECD guideline for the testing of chemicals. Draft proposal for a new guideline. In vitro skin irritation: human skin model test, December, 2007
Deviations:
yes
Remarks:
- Interleukin-1 alpha was not determined
Principles of method if other than guideline:
The skin irritancy potenial of silicic acid, aluminium salt was tested by using the human skin model EpiDerm and measurement of cell viability by dehydrogenase conversion of MTT into a blue formazan salt.
GLP compliance:
yes (incl. certificate)
Remarks:
Landesamt für Umwelt, Wasserwirtschaft und Gewerbeaufsicht, Kaiser-Friedrich-Strasse 7, 55116 Mainz, Germany

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): silicic acid, aluminium salt
- Physical state: white powder
- Analytical purity: > 99%
- Batch No.: 1000168802
- Expiration date of the batch: Apr 2010
- Storage condition of test material: at room temperature, 20 ± 5 °C

Test animals

Species:
other: in vitro test: human skin
Strain:
other: in vitro test: human skin
Details on test animals and environmental conditions:
not applicable

Test system

Type of coverage:
other: in vitro test: human skin model test
Preparation of test site:
other: in vitro test: human skin model test
Vehicle:
other: DPBS buffer
Controls:
other: not applicable
Amount / concentration applied:
TEST MATERIAL
- Amount(s) applied (weight): 25 mg of the neat test item were applied to each of triplicate tissues and wetted with 25 µL DPBS (Dulbecco's Phophate Buffered Saline) buffer.

VEHICLE
- Amount(s) applied (volume): 25 µL DPBS buffer
Duration of treatment / exposure:
60 min
Observation period:
incubation time before determination of cell viability: 42 h
Number of animals:
not applicable
Details on study design:
The Epi-200 -Kit was purchased from MatTek Corporation (Ashland, USA). The tissue consists of normal, human-derived epidermal keratinocytes which have been cultured to form a multilayered, highly differentiated model of the human epidermis. The EpiDermTM tissues are cultured on specially prepared cell culture inserts.
EpiDermTM tissues reached LAUS GmbH on September 23, 2009. On the day of receipt, EpiDermTM tissues were transferred to 12-well plates with maintenance medium.
After 18.5 hours incubation of the EpiDermTM tissues, three tissues of the human skin model EpiDermTM were treated with either the test item, the negative (Dulbecco's Phosphate Buffered Saline (DPBS buffer)) or the positive control (5% sodium lauryl sulfate in deionised water) for a total of 60 min. Approx. 25 mg of the test item were applied to each tissue and wetted with 25 µL DPBS buffer. 30 µL of either the negative or the positive control were applied to each tissue. The 6-well plates were placed into the incubator for 35 min at 37 ± 1 °C, 5 ± 0.5% CO2.
After the end of the treatment interval the inserts were removed from the plates using sterile forceps.The tissues were rinsed immediately and transferred into a new 6 -well plate with fresh assay medium. All inserts were dried with sterile cotton tips. The tissues were set in the incubator for 42 hours.

Cell viability was measured by dehydrogenase conversion of MTT [(3,4,5 -dimethyl thiazole 2 -yl) 2,5 -diphenyltetrazoliumbromide], present in cell mitochondria, into a blue formazan salt that is quantitatively measured after extraction from tissues. The percent reduction of cell viability in comparison to untreated negative controls is used to predict skin irriation potential.
The tissues were placed in MTT solution of 1 mg/mL for 3 hours (37 ± 1 °C, 5 ± 0.5% CO2). The precipitated blue formazan product is then extracted using isopropanol. The concentration of formazan was measured by determining the Optical Density (OD) at 570 nm in a plate spectral photometer.

Evaluation of the results
The mean OD of the three negative control tissues was calculated. This value corresponds to 100% tissue viability in the current test. For each individual tissue treated with the test item or the positive control the individual relative tissue viability was calculated according to the following formula:
Relative viability (%)= (OD(test item) x 100) / OD (negative control)

Acceptability of the assay
The absolute OD 570 nm of the negative control tissues in the MTT test is an indicator of tissue viability obtained after the shipping and storing procedure and under specific conditions of the assay. Tissue viability is meeting the acceptance criterion if the mean OD of the three tissues is between 1.0 and 2.5. An assay is meeting the acceptance criterion if mean relative tissue viability of the positive control is ≤ 20% of the negative control and the variation within replicates is below 18%.

Results and discussion

In vivo

Irritant / corrosive response data:
After treatment with the negative control the absorbance values were well within the required acceptability criterion thus showing the quality of the tissues. The positive control induced a decrease in the relative absorbance as compared to the negative control to 9.4% thus ensuring the validity of the test system. After tissue incubation with silicic acid, aluminium salt, the relative absorbance values were increased to 103.4%. This value is well above the threshold for irritancy of 50%. Therefore, the test item is considered to have no skin irritation potential. For detailed results see table 1 in "any other information on results incl. tables".

Any other information on results incl. tables

Table 1:

Dose group

Absorbance 570 nm, tissue 1*

Absorbance 570 nm, tissue 2*

Absorbance 570 nm, tissue 3*

Mean absorbance of 3 tissues

Mean formazan production [%]

Negative control

2.242

2.027

1.760

2.010

 100%

Positive control

0.183

0.192

0.192

0.189

 9.4%

Test item

2.185

2.011

2.041

2.079

103.4%

*Mean of two replicate wells after blank correction

** relative absorbance [rounded values]: (100 x (absorbance test item)) / (absorbance negative control)

 

Applicant's summary and conclusion