Potassium 3,5,5-trimethylhexanoate

Skin irritation / corrosion

Administrative data

Endpoint:

skin corrosion: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2016

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Title:

Unnamed

Year:

2016

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 431 (In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Potassium 3,5,5-trimethylhexanoate

EC Number:

299-890-3

EC Name:

Potassium 3,5,5-trimethylhexanoate

Cas Number:

93918-10-6

Molecular formula:

C9H18O2.K

IUPAC Name:

potassium 3,5,5-trimethylhexanoate

Test material form:

solid: crystalline

Details on test material:

Name: Potassium 3,5,5-trimethylhexanoate
CAS No.: 93918-10-6
Batch No.: PURS151015
Purity: 99.4%
Physical State: solid
Colour: white
Molecular Weight: 196.33 g/mol
Storage Conditions: room temperature
Re-certification Date: 14 October 2016

Specific details on test material used for the study:

Name: Potassium 3,5,5-trimethylhexanoate
CAS No.: 93918-10-6
Batch No.: PURS151015
Purity: 99.4%
Physical State: solid
Colour: white
Molecular Weight: 196.33 g/mol
Storage Conditions: room temperature
Re-certification Date: 14 October 2016

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

skin obtained from plastic surgery from multiple donors

Source strain:

other: adult donors

Vehicle:

physiological saline

Details on test system:

The test was carried out with the reconstituted three-dimensional human skin model EPISKIN-SM™ (SkinEthic). This skin model consists of normal human epidermal keratinocytes (NHEK) which have been cultured to form a multilayered, highly differentiated model of the human epidermis. The NHEK are cultured on chemically modified, collagen-coated cell culture inserts. A highly differentiated and stratified epidermis model is obtained after 13-day culture period comprising the main basal, supra basal, spinous and granular layers and a functional stratum corneum.

Morphology:
Histology scoring (HES stained vertical paraffin sections, n = 6):
specification ≥ 19.5
result: 21.8 ± 0.3, CV = 1.2%
Well-differentiated epidermis consisting of a basal layer, several spinous and granular layers and a
thick stratum corneum.
Barrier function:
IC50 determination (SDS concentration, MTT test, n = 14):
specification ≥ 1.5 mg/mL
result: 2.8 mg/mL

The mixture of 20 mg test item per 2 mL MTT medium showed no reduction of MTT as compared to the solvent. The mixture did not turn blue/purple. Therefore, NSMTT equaled 0%. The mixture of 10 mg test item per 300 µL Aqua. dest. and per 90 µL isopropanol showed no colouring as compared to the solvent. Therefore NSC equaled 0%.

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

20 ± 2 mg (52.6 mg/cm2) of the test item was applied directly atop the EPISKIN-SM™ tissue. To ensure good contact with the epidermis surface, the test item was moistened with 100 ± 5 µL 0.9% NaCl solution. The test item was spread to match size of the tissue.

Duration of treatment / exposure:

In the present study Potassium 3,5,5-trimethylhexanoate was applied topically to the EPISKIN-SM™ tissue for 3 min, 60 min and 4 h followed by immediate determination of cytotoxic effects via MTT reduction assay.

Number of replicates:

The test was performed on a total of 6 tissues for each test item and for the negative control, 2 replicates for each treatment period (3 min, 60 min and 4 h exposure time). For the positive control the test was performed with 2 replicates for the 4 h exposure time.

Results and discussion

In vitro

Results

Results 1

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

1 - 3 min experiment

Value:

104.3

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not applicable

Remarks on result:

no indication of irritation

Results 2

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

2 - 3 min experiment

Value:

101.8

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not applicable

Remarks on result:

no indication of irritation

Results 3

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

3 - 60 min experiment

Value:

20

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not applicable

Remarks on result:

positive indication of irritation

Results 4

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

4 - 60 min experiment

Value:

23

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

not applicable

Remarks on result:

positive indication of irritation

Results 5

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

5 - 4h experiment

Value:

16.1

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks:

glacial acetic acid

Remarks on result:

positive indication of irritation

Results 6

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

6 - 4h experiment

Value:

12.9

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks:

glacial acetic acid

Remarks on result:

positive indication of irritation

Other effects / acceptance of results:

The controls confirmed the validity of the study. The mean OD570 of the two negative control tissues was between 0.6 and 1.5 for each exposure period. The mean relative tissue viability (% negative control) of the positive control was ≤ 20% (3%) after 4 h treatment. Inter tissue viability difference of replicate tissues of all dose groups was ≤ 30% (0.2% - 9.4%).

Any other information on results incl. tables

The potential of the test item to induce skin corrosion was analysed by using the three-dimensional human skin model EPISKIN-SM™, comprising a reconstructed epidermis with a functional stratum corneum. In the present study Potassium 3,5,5-trimethylhexanoate was applied topically to the EPISKIN-SM™ tissue for 3 min, 60 min and 4 h followed by immediate determination of cytotoxic effects via MTT reduction assay. Corrosivity potential of the test item was predicted from the relative mean tissue viabilities compared to the corresponding negative control tissues concurrently treated with 0.9% NaCl. The test item showed no non-specific MTT-reducing or water-colouring potential. The test item showed corrosive effects. The mean relative tissue viability (% negative control) was reduced below 35% (20%) after 60 min treatment and to not more than 35% (103%) after 3 min treatment. Relative mean tissue viability was reduced to 15% after 4 h treatment. The controls confirmed the validity of the study. The mean OD570 of the two negative control tissues was between 0.6 and 1.5 for each exposure period. The mean relative tissue viability (% negative control) of the positive control was ≤ 20% (3%) after 4 h treatment. Inter tissue viability difference of replicate tissues of all dose groups was ≤ 30% (0.2% - 9.4%).

Applicant's summary and conclusion

Interpretation of results:

Category 1 (corrosive) based on GHS criteria

Conclusions:

In this study under the given conditions the test item showed corrosive effects. The relative mean tissue viability after 60 min was reduced to less than 35% but not more than 35% after 3 min treatment. The test item is therefore classified as corrosive in accordance with a combination of
optional sub-categories 1B and 1C.

Executive summary:

In the present study the skin corrosivity potential of Potassium 3,5,5-trimethylhexanoate was analysed. Since corrosive chemicals are cytotoxic after a short time exposure to the stratum corneum of the epidermis the cytotoxic effects of the test item on EPISKIN-SM™, a reconstituted three-dimensional human epidermis model, were determined. Hereby, the test item was applied topically. Cytotoxicity is expressed as the reduction of mitochondrial dehydrogenase activity

measured by formazan production from MTT after a 3 min, 60 min and 4 h exposure period and compared to those of the concurrent negative controls. In this study under the given conditions the test item showed corrosive effects. The relative mean tissue viability after 60 min was reduced to less than 35% but not more than 35% after 3 min treatment. The test item is therefore classified as corrosive in accordance with a combination of optional sub-categories 1B and 1C.