Dipotassium propanedioate

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

18.-20.10.2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 439 (In Vitro Skin Irritation: Reconstructed Human Epidermis Test Method)

Version / remarks:

28 July 2015

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.46 (In Vitro Skin Irritation: Reconstructed Human Epidermis Model Test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Dipotassium propanedioate

EC Number:

820-064-0

Cas Number:

13095-67-5

Molecular formula:

C3H2K2O4

IUPAC Name:

Dipotassium propanedioate

Specific details on test material used for the study:

purity: ≥ 95%

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Cell source:

other: human-derived epidermal keratinocytes

Source strain:

other: not applicable

Vehicle:

other: DPBS

Remarks:

to improve the contact between the powder and the epidermis

Details on test system:

- Source: MatTek Corporation (82105 Bratislava, Slovakia).
- The EpiDerm™ tissue: normal, human-derived epidermal keratinocytes which have been cultured to
form a multilayered, highly differentiated model of the human epidermis.
- Surface: 0.63 cm.
- Pre-incubation: 60 ± 5 minutes in the incubator (37 ± 1 °C, 5% CO2) in the upper
wells. Then transferred from upper wells into the lower wells for about 19 hours (37 ± 1 °C, 5 ± 0.5% CO2).

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

25 mg + 25 μL DPBS

Duration of treatment / exposure:

60 ± 1 minutes

Duration of post-treatment incubation (if applicable):

approx. 42 hours

Number of replicates:

3

Test system

Details on study design:

Details of the test procedure used:
- EpiDerm™ tissue of human-derived epidermal keratinocytes was used (EPI-200-SIT)
- Conditions of exposure: 37 ± 1 °C, 5% CO2
- Washing: inserts gently rinsed with DPBS
- Number of tissue replicates used per test chemical and controls: 3
- MTT assay: incubation with 0.3 mL of MTT solution for 60 minutes at 37 ± 1 °C
- Data evaluation: the following was calculated: The mean OD of the three negative control tissues
was calculated after blank correction. The mean of the photometric absorbance of the negative control
is set to 100%.
For each individual tissue treated with the test item or the positive control the individual relative
tissue viability is calculated according to the following formula: Relative viability(%) = (mean OD test i
tem / positive control / mean OD negative control) x 100. For the test item and the positive control the
mean relative viability ± rel. standard deviation of the three individual tissues was calculated

Description of evaluation criteria:
- GHS Cat 2 according to UN GHS is recommended if the mean relative tissue viability of three individual tissues is reduced ≤ 50% of the negative control
- GHS “No Category” if the tissue viability after exposure and post-treatment incubation is more than 50%
- Historical data positive control: Mean Viability: 3.9%; Rel. Standard Deviation: 4.4%
- Historical data negative control: Mean Absorption: 1.831; Rel. Standard Deviation: 0.375;

The test meets acceptance criteria if:
- mean absolute OD570 nm of the three negative control tissues is ≥ 0.8 and ≤ 2.8
- mean relative tissue viability of the three positive control tissues is ≤ 20%
- standard deviation (SD) of relative tissue viability obtained from each three concurrently tested
tissues is ≤ 18%.

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Single test with three tissues

Value:

94.8

Vehicle controls validity:

not applicable

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

The controls confirmed the validity of the study:
- the mean absolute OD570 of the three negative control tissues was ≥ 0.8 and ≤ 2.8.
- the mean relative tissue viability (% negative control) of the positive control was ≤ 20% (5.0%).
- standard deviation of viability of replicate tissues of all dose groups was ≤ 18% (0.4% - 10.7%).

Any other information on results incl. tables

Result of the Test Item Dipotassium malonate

Name	NK			PC			TM
Tissue	1	2	3	1	2	3	1	2	3
absolute OD570	1.518	1.533	1.619	0.123	0.119	0.112	1.594	1.569	1.270
	1.554	1.471	1.578	0.124	0.121	0.111	1.458	1.609	1.300
OD570(blank-corrected)	1.475	1.4489	1.576	0.080	0.075	0.068	1.550	1.526	1.227
	1.510	1.428	1.535	0.080	0.078	0..067	1.414	1.566	1.257
mean OD570of the duplicates (blank-corrected)	1.493	1.459	1.555	0.080	0.076	0.068	1.482	1.546	1.242
total mean OD570of 3 replicate tissues (blank-corrected)	1.502*			0.075			1.423
SD OD570	0.049			0.006			0.160
relative tissue viability [%]	99.4	97.1	103.5	5.3	5.1	4.5	98.7	102.9	82.7
mean relative tissue viability [%]	100.0			5.0**			94.8
SD tissue viability [%]***	3.3			0.4			10.7
CV [% viabilities]	3.3			8.4			11.3

* Blank-corrected mean OD570 nm of the negative control corresponds to 100% absolute tissue viability.

** Mean relative tissue viability of the three positive control tissues is ≤ 20%.

*** Standard deviation (SD) obtained from the three concurrently tested tissues is ≤ 18%

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In this study under the given conditions the test item showed no irritant effects.
The test item is therefore classified as “non-irritant” in accordance with UN GHS “No Category”.

Executive summary:

The potential of the test item Dipotassium malonate to induce skin irritation was analysed by using the three-dimensional human epidermis model EpiDerm (MatTek) comprising a reconstructed epidermis with a functional stratum corneum. The test was performed according to OECD TG 439 and in compliance to GLP.

In the present study Dipotassium malonate was applied topically to the EpiDerm tissue for 60 min followed by a 42 h post-incubation period and immediate determination of cytotoxic effects via MTT reduction assay.

Irritant potential of the test item was predicted from the relative mean tissue viabilities obtained compared to the corresponding negative control tissues concurrently treated with DPBS.

The mixture of 30 μL test item per 1 mL MTT medium showed no reduction of MTT compared to the solvent. The mixture did not turn blue/purple. Therefore, NSMTT equaled 0%. The mixture of 30 μL of the test item per 300 μl aqua dest. or per 300 μL isopropanol showed no colouring detectable by unaided eye-assessment. Therefore, NSC equaled 0%.

The test item showed no irritant effects. The mean relative tissue viability (% negative control) was > 50% (94.8%) after 60 min treatment and 42 h post-incubation.

The controls confirmed the validity of the study. The mean absolute OD570 of the three negative control tissues was ≥ 0.8 and ≤ 2.8. The mean relative tissue viability (% negative control) of the positive control was ≤ 20% (5.0%). Standard deviation of viability of replicate tissues of all dose groups was ≤ 18% (0.4% - 10.7%).

In this study under the given conditions the test item showed no irritant effects. The test item is therefore classified as “non-irritant” in accordance with UN GHS “No Category”.