Boric acid (H3BO3), solid soln. with strontium oxide, europium-doped

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Report date:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Boric acid (H3BO3), solid soln. with strontium oxide, europium-doped

EC Number:

310-028-8

EC Name:

Boric acid (H3BO3), solid soln. with strontium oxide, europium-doped

Cas Number:

102110-29-2

Molecular formula:

SrB4O7:Eu

IUPAC Name:

Boric acid (H3BO3), solid soln. with strontium oxide, europium-doped

Test material form:

solid

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Zhejiang Jingneng Phosphor Material Co., Ltd; 170825
- Expiration date of the lot/batch: 24 August 2019
- Purity: 99%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: room temperature, protected from light

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) and therefore represents in vitro the target organ of the species of interest and closely mimics the biochemical and physiological properties of the upper parts of the human skin, i.e. the epidermis

Vehicle:

other: Dulbecco’s phosphate buffered saline (DPBS)

Details on test system:

RECONSTRUCTED HUMAN EPIDERMIS (RHE) TISSUE
- Model used: in vitro EpiDerm™ Skin Irritation Test (EPI-200-SIT), MatTek
- Tissue batch number(s): 28646

TEMPERATURE USED FOR TEST SYSTEM
- Temperature used during treatment / exposure: 35 ± 1 min at 37 ± 1 °C and 25 ± 1 min at room temperature
- Temperature of post-treatment incubation (if applicable): 42 ± 2 h at 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, this process also occurred sequentially, e.g. in one-minute intervals. 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: 5 mg/mL
- Incubation time: 3 h ± 5 min
- Wavelength: 570 nm
- Filter and bandwidth: filter band pass of maximum ± 30 nm

NUMBER OF REPLICATE TISSUES: 3

CONTROL TISSUES USED IN CASE OF MTT DIRECT INTERFERENCE
To check the non-specific MTT-reducing capability of the test item 25 mg of the test item were mixed per 1 mL MTT medium and incubated for 60 min at 37 ± 1 °C in the incubator. 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%.

Colour Interference
To check the colouring potential of the test item 25 mg of the test item were mixed per 300 µL aqua dest. and per 300 µL isopropanol each in a transparent recipient and incubated at 37 ± 1°C for 60 min. No colouring was detectable by unaided eye-assessment. Therefore, NSC equalled 0%.

PREDICTION MODEL / DECISION CRITERIA (choose relevant statement)
The test item is considered to be irritant to skin in accordance with Regulation EC 1272/2008, if the tissue viability after exposure and post-incubation is less or equal to 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 + 25 µL DPBS

VEHICLE :
Dulbecco’s phosphate buffered saline (DPBS; Gibco, Cat. No. 14040-091, Lot No.: 1838067).

NEGATIVE CONTROL
- Amount(s) applied (volume or weight): 30 µL DPBS


POSITIVE CONTROL
- 30 µL 5% sodium dodecyl sulfate (TC-SDS-5%; MatTek, CAS No.: 151-21-3, Lot No.: 081418ALA).

Duration of treatment / exposure:

35 ± 1 min at 37 ± 1 °C and 25 ± 1 min at room temperature

Duration of post-treatment incubation (if applicable):

42 ± 2 h at 37 ± 1 °C

Number of replicates:

3

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

% tissue viability

Run / experiment:

Test substance

Value:

93.6

Vehicle controls validity:

not examined

Negative controls validity:

valid

Positive controls validity:

valid

Other effects / acceptance of results:

- OTHER EFFECTS:
- Visible damage on test system: None specified
- Direct-MTT reduction: 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%.
- Colour interference with MTT: No colouring was detectable by unaided eye-assessment. Therefore, NSC equalled 0%.

ACCEPTANCE OF RESULTS:
- Acceptance criteria met for negative control: The mean absolute OD570 of the three negative control tissues was  0.8 and ≤ 2.8 (1.789).
- Acceptance criteria met for positive control: The mean relative tissue viability (% negative control) of the positive control was ≤ 20% (4.7%).
- Acceptance criteria met for variability between replicate measurements: Standard deviation of viability of replicate tissues of all dose groups was ≤ 18% (0.3% – 4.3%).
- Range of historical values if different from the ones specified in the test guideline: Historical data were generated from 2015 to 2018 (n=40).

Applicant's summary and conclusion

Interpretation of results:

GHS criteria not met

Conclusions:

In an in vitro skin irritation test using a human epidermal model (EPI-200), the substance is not irritating.

Executive summary:

In an in vitro skin irritation assay in a human epidermal model EPI-200 (185424), reconstructed human epidermis tissue was exposed to 25 mg of boric acid (H3BO3), solid soln. with strontium oxide, europium-doped (99%) + 25 µL DPBS for 35 ± 1 min at 37 ± 1 °C and 25 ± 1 min  at room temperature. DPBS was used for the negative control and 5% SDS was used for the positive control. After removal of the test substance, tissues were post-incubated for approximately 42 hours. Three hours incubation with MTT and a 2-hour isopropyl alcohol extraction period followed. The OD570 of isopropyl alcohol extracts was measured on a spectrophotometer. Relative cell viability was calculated for each tissue as % of the mean viability of the negative control tissues.

The colour of the test substance did not interfere with the endpoint. The test substance does not directly reduce MTT. The average viability of tissues treated by the test substance was 93.6% compared to the negative control i.e. viability was > 50 %. The average viability of tissues treated by the positive control was 4.7% of negative control average value. According to these results, the test substance is not irritating.