Reaction mass of 3,4,5-trichlorophthalic acid, strontium salt and 3,4,6-trichlorophthalic acid, strontium salt and 3,4-dichlorophthalic acid, strontium salt and 3,5-dichlorophthalic acid, strontium salt and 3,6-dichlorophthalic acid, strontium salt and 3-chlorophthalic acid, strontium salt and 4,5-dichlorophthalic acid, strontium salt and 4-chlorophthalic acid, strontium salt

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2019-06-24 to 2019-09-10

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

adopted on 18th June 2019

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Strontium dichlorophthalate

EC Number:

304-292-3

EC Name:

Strontium dichlorophthalate

Cas Number:

94248-20-1

Molecular formula:

C8H4Cl2O4.Sr

Test material form:

solid

Details on test material:

Strontium phthalate (HPLC): 0.51 w/w %
Strontium 3-chlorophthalate (HPLC): 0.73 w/w %
Strontium 3,6-dichlorophthalate (HPLC) / Strontium 4-chlorophthalate (HPLC): 10.59 w/w %
Strontium 3,4-dichlorophthalate (HPLC): 20.56 w/w %
Strontium 3,5-dichlorophthalate (HPLC): 0.66 w/w %
Strontium 3,4,6-trichlorophthalate (HPLC): 0.18 w/w %
Strontium 4,5-dichlorophthalate (HPLC): 60.59 w/w %
Strontium 3,4,5-trichlorophthalate (HPLC): 4.39 w/w %
Strontium tetrachlorophthalate (HPLC): 0 w/w %

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch number of test material:Dynamit Nobel GmbH Explosivstoff- und Systemtechnik, WE50352354
- Expiration date of the lot/batch: 28 November 2019
- Purity test date: 2 May 2019

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: 5 °C – 40 °C, protected from light, separated from oxidizers, strong acids and alkalis; do not heat
- Stability under storage conditions: stable under normal conditions

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
- Treatment of test material prior to testing: The test item (25 mg) was directly applied atop the tissue

In vitro test system

Test system:

human skin model

Source species:

human

Cell type:

non-transformed keratinocytes

Justification for test system used:

This test uses the EpiDerm™ reconstructed human epidermis model (MatTek) which consists of normal human epidermal keratinocytes (NHEK) which have been cultured to form a multi-layered, highly differentiated model of the human epidermis and therefore the EpiDerm™ epidermis model exhibits in vivo-like morphological and growth characteristics which are uniform and highly reproducible.

Vehicle:

unchanged (no vehicle)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: EpiDerm™ Standard Model (EPI-200-SIT, MatTek)
- Tissue batch number(s): Lot No.: 30807
- Date of initiation of testing: 24th June 2019
TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 37 ± 1 °C
- Temperature of post-treatment incubation (if applicable): 37 ± 1 °C
REMOVAL OF TEST MATERIAL AND CONTROLS
- Volume and number of washing steps: the tissues were washed by filling and emptying the inserts 15 times with DPBS using a constant stream in about 1.5 cm distance from the tissue surface. Subsequently, the inserts were completely submerged three times in 150 mL DPBS and shaken to remove rests of the test item. Finally, the inserts were rinsed once from the inside and the outside with sterile DPBS. Excess DPBS was removed by blotting the bottom with blotting paper.
MTT DYE USED TO MEASURE TISSUE VIABILITY AFTER TREATMENT / EXPOSURE
- MTT concentration: 1 mg/mL
- Incubation time: 3 h ± 5 min
- Wavelength: 570 nm
- Filter bandwidth: ± 30 nm
NUMBER OF REPLICATE TISSUES: 3
PREDICTION MODEL / DECISION CRITERIA
- The test substance is considered to be irritant to skin if the tissue viability after exposure and post-incubation is less than or equal to 50%
- The test substance is considered to be non-irritant to skin if the viability after exposure and post-incubation is greater than 50%

Control samples:

yes, concurrent negative control

yes, concurrent positive control

Amount/concentration applied:

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 25 mg
NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 μL DPBS

POSITIVE CONTROL
- Amount(s) applied (volume or weight): 30 μL 5% SDS solution

Duration of treatment / exposure:

60 ± 1 min

Duration of post-treatment incubation (if applicable):

42 h post-incubation

Number of replicates:

3 per dose group

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

mean of three tissues

Value:

86.3

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Remarks on result:

no indication of irritation

Other effects / acceptance of results:

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: Yes, the mean absolute OD570 of the three negative control tissues was ≥ 0.8 and ≤ 2.8 (1.646)
- Acceptance criteria met for positive control: Yes, the mean relative tissue viability of the positive control was ≤ 20% (7.4%)
- Acceptance criteria met for variability between replicate measurements: Yes, the standard deviation of viability of replicate tissues of all dose groups was ≤ 18% (1.1%).

For detailed results see Table 1 in box "Any other information on results incl. tables".

Any other information on results incl. tables

Results of the Pre-Experiments:

The mixture of 25 mg 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 equalled 0%.

The mixture of 25 mg of the test item per 300 μL aqua dest. and per 300 μL isopropanol showed no colouring detectable by unaided eye-assessment. Therefore, NSC equalled 0%.

The test item showed no non-specific reduction of MTT and no relevant colouring potential after mixture with aqua dest. and with isopropanol. Therefore, no additional controls for correction of possible false-negative results were necessary.

 

Results of the main experiment:

 

Name	Negative Control			Positive Control			Test Item
Replicate Tissue	1	2	3	1	2	3	1	2	3
Absolute OD570	1.716	1.750	1.419	0.138	0.171	0.173	1.368	1.380	1.487
	1.737	1.803	1.453	0.160	0.152	0.193	1.375	1.469	1.482
Mean Absolute OD570	1.646****			0.164			1.427
OD570(Blank Corrected)	1.669	1.704	1.372	0.091	0.125	0.126	1.321	1.333	1.441
	1.691	1.757	1.406	0.114	0.105	0.147	1.328	1.423	1.435
Mean OD570of the Duplicates (Blank Corrected)	1.680	1.730	1.389	0.102	0.115	0.136	1.325	1.378	1.438
Total Mean OD570of the 3 Replicate Tissues (Blank Corrected)	1.600*			0.118			1.380
SD of Mean OD570of the 3 Replicate Tissues (Blank Corrected)	0.184			0.017			0.057
Relative Tissue Viability [%]	105.0	108.2	86.8	6.4	7.2	8.5	82.8	86.1	89.9
Mean Relative Tissue Viability [%]	100.0			7.4**			86.3
SD of Relative Tissue Viability [%]***	11.5			1.1			3.5
CV [% Viabilities]	11.5			14.6			4.1

 * Blank-corrected mean OD₅₇₀ 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%.

**** The mean absolute OD₅₇₀ of the negative control i s≥ 0.8 and ≤ 2.8.

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In conclusion, in this in vitro skin irritation study (OECD 439), strontium dichlorophthalate is considered to no be irritant to the skin (UN GHS “No category”).

Executive summary:

In a primary dermal irritation study conducted according to OECD guideline 439, the EpiDerm™-Model (EPI-200-SIT) was topically exposed to strontium dichlorophthalate for 60 min and 42 h post incubation period. Irritant potential of the test item was not predicted from the relative mean tissue viabilities obtained compared to the corresponding negative control tissues concurrently treated with DPBS. The mean relative tissue viability (% negative control) was > 50% (86.3 %) after 60 min treatment and 42 h post-incubation. Therefore, the test item is not considered to be irritating to the skin in accordance with UN GHS "No Category”.