Fatty acids, C18-unsatd., dimers, oligomeric reaction products with fatty acids, C16-18 and C18-unsatd., branched and linear, tetraethylenepentamine and triethylenetetramine

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation / corrosion

Remarks:

in vitro

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Conducted in accordance with GLP and appropriate testing guidelines.

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Human Skin Model Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

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

Test material

Test material information

Constituent 1

Reference substance name:

Fatty acids, C18-unsatd., dimers, oligomeric reaction products with fatty acids, C16-18 and C18-unsatd., branched and linear, tetraethylenepentamine and triethylenetetramine

EC Number:

500-382-3

EC Name:

Fatty acids, C18-unsatd., dimers, oligomeric reaction products with fatty acids, C16-18 and C18-unsatd., branched and linear, tetraethylenepentamine and triethylenetetramine

Cas Number:

157707-73-8

IUPAC Name:

Reaction product of fatty acids, C18-unsatd., dimers, and trimers, with Amines, polyethylenepoly-, triethylenetetramine fraction and amines, polyethylenepoly-, tetraethylenepentamine fraction

Constituent 2

Reference substance name:

MonoFA_DimerFA_TETA_TEPA_PAA

IUPAC Name:

MonoFA_DimerFA_TETA_TEPA_PAA

Constituent 3

Reference substance name:

Fatty acids, C18-unsatd., dimers, oligomeric reaction products with fatty acids, C16-18 and C-18 unsatd., branched and linear, tetraethylenepentamine and triethylenetetramine

IUPAC Name:

Fatty acids, C18-unsatd., dimers, oligomeric reaction products with fatty acids, C16-18 and C-18 unsatd., branched and linear, tetraethylenepentamine and triethylenetetramine

Test material form:

other: liquid

Details on test material:

- Name of test material (as cited in study report): MonoFA_DimerFA_TETA_TEPA_PAA- Physical state: Yellow liquid with a brown hue. - Analytical purity: 100%- Lot/batch No.: BB001148V1- Expiration date of the lot/batch: 30th June 2013- Storage condition of test material: When not in use the test article was stored in a sealed container, at room temperature in the dark.

Test animals

Species:

other: Not applicable - in vitro study

Strain:

other: Not applicable - in vitro study

Details on test animals or test system and environmental conditions:

Not applicable - in vitro study.

Test system

Type of coverage:

other: Not applicable

Preparation of test site:

other: Not applicable

Vehicle:

unchanged (no vehicle)

Controls:

not required

Amount / concentration applied:

TEST MATERIALIn the preliminary MTT test, 40 μL of test article was added to 0.3 mL of 1.0 mg/mL MTT.In the Epiderm test, an average of approximately 50 mg of test article was applied to each tissue.Further tissues were concurrently treated with 40 μL distilled water (negative control) and with 40 μL 8N potassium hydroxide (positive control).

Duration of treatment / exposure:

Not applicable.

Observation period:

Not applicable.

Number of animals:

Not applicable.

Details on study design:

In order to assess the potential non-specific reduction of the test article, 40 μL of test article was added to 0.3 mL of 1.0 mg/mL MTT and the colour change was assessed after three hours.An increase in optical density was noted over the control (untreated MTT solution) therefore additional controls were included to enable assessment of the potential impact of the non-specific reduction of MTT by residual test article following the wash-off procedure.To evaluate whether residual test article was binding to the tissue, a functional check (using freeze-killed control tissue) was performed. Freeze-killed tissues were prepared by placing untreated EpiDerm constructs in the -20°C freezer overnight. The freeze-killed tissues were allowed to thaw once at room temperature and placed back in the freezer overnight.On the day of receipt EpiDermTM tissues were placed in a refrigerator. Ninety minutes before starting the assay, the tissues were transferred to 6-well plates containing the assay medium. The test was performed on a total of four tissues per test article, negative and positive control, out of which two were used for a 3 minute application and two tissues were used for a 1 hour application. Sufficient test article was applied to cover the apical surface. The cryovials containing the test article were weighed both prior to and post treatment and an average of approximately 50 mg of test article was applied to each tissue (an average of 34 mg was applied to the freeze-killed tissues).Further tissues were concurrently treated with 40 μL distilled water (negative control) and with 40 μL 8N potassium hydroxide (positive control). After the 3-minute or 1-hour contact periods, the tissues were washed with phosphate buffered saline (PBS) to remove residual material. The rinsed tissues were kept in 24-well plates (holding plates) until all tissues were dosed and rinsed.Once all tissues had been rinsed, they were transferred to wells containing 300 μL of 1 mg/mL MTT-medium and were incubated for 3 hours (37°C, 5% CO2). After incubation, the tissues were washed with PBS and any resultant colour was extracted with 2 mL isopropanol overnight. The optical density of each resultant extract was determined spectrophotometrically at 570 nm and cell viability was calculated for each tissue as a percentage of the mean of the negative control tissue. Skin corrosivity potential of the test article was classified according to the remaining cell viability obtained after test article treatment with either of the two treatment times. Also, to evaluate whether residual test article was binding to the tissue, a functional check (using freeze-killed control tissue) was performed. Freeze-killed tissues were prepared by placing untreated EpiDerm constructs in the -20°C freezer overnight. The killed tissues were allowed to thaw once at room temperature and were placed back at -20°C overnight. Once killed, the tissues were stored in the freezer until required.

Results and discussion

In vivo

Results

Results 1

Irritation parameter:

other: Skin viability

Basis:

mean

Time point:

other: 3 minute exposure

Remarks on result:

other: 57% skin viability

Results 2

Irritation parameter:

other: Skin viability

Basis:

mean

Time point:

other: 1 hour exposure

Remarks on result:

other: 56% skin viability

Irritant / corrosive response data:

Skin viability after a three minute exposure was 57%.Skin viability after a one hour exposure was 56%.Skin viability in the positive control after a three minute exposure was 23%.Skin viability in the positive control after a one hour exposure was 16%.

Other effects:

No other effects reported.

Any other information on results incl. tables

Three minute exposure period

Test substance	OD570			Mean	Tissue Mean	Adjusted mean value	% variability	% survival
Negative	1.862	1.902	1.921	1.895	1.771	N/A/	-15.1	100
Negative	1.603	1.622	1.714	1.646
Test article	1.437	1.457	1.430	1.441	1.398	1.006	-6.4	57
Test article	1.343	1.340	1.381	1.355
Test article#	0.453	0.477	0.490	0.474	0.392		34.4
Test article#	0.313	0.307	0.312	0.311
Positive	0.406	0.436	0.439	0.427	0.413	N/A/	6.4	23
Positive	0.394	0.399	0.406	0.399

# Freeze-killed tissues

One hour exposure period

Test substance	OD570			Mean	Tissue Mean	Adjusted mean value	% variability	% survival
Negative	1.584	1.587	1.676	1.616	1.640	M/A	2.9	100
Negative	1.688	1.650	1.654	1.664
Test article	1.311	1.306	1.274	1.297	0.328	0.926	4.5	56
Test article	1.368	1.348	0.359	1.359
Test article#	0.370	0.393	0.400	0.388	0.402		-7.1
Test article#	0.410	0.421	0.415	0.415
Positive	0.251	0.270	0.261	0.261	0.268	N/A/	-5.7	16
Positive	0.281	0.273	0.272	0.275

# Freeze-killed tissues

Applicant's summary and conclusion

Interpretation of results:

not classified

Remarks:

Migrated informationCriteria used for interpretation of results: EU

Conclusions:

Under the conditions of this study, the test article, MonoFA_DimerFA_TETA_TEPA_PAA, was not corrosive to skin in the in vitro skin model EpiDerm.

Executive summary:

The potential of MonoFA_DimerFA_TETA_TEPA_PAA to cause skin corrosion was evaluated using the in vitro skin model, EpiDerm^TM, in accordance with GLP and OECD Test Guideline 431 and EU Method B.40.

Duplicate EpiDerm^TM inserts were treated with the test article, distilled water (negative control) and 8N potassium hydroxide (positive control) for 3 minutes and 1 hour. At the end of the treatment period, the tissues were washed with phosphate buffered saline (PBS) and cell viability was assessed using the MTT assay. The skin corrosivity potential was assessed according to the remaining cell viability obtained after test material treatment with either of the two treatment times.

Skin viability after a three minute or one hour exposure to the test article was 57% and 56%, respectively. Skin viability after a three minute or one hour exposure to the positive control article was 23% and 16%, respectively, demonstrating appropriate performance of the assay.

Based on these results and under the conditions of this test, the test article, MonoFA_DimerFA_TETA_TEPA_PAA, was not corrosive to skin in the in vitro skin model EpiDerm^TM.