Quino[2,3-b]acridine-6,7,13,14(5H,12H)-tetrone

Skin irritation / corrosion

Administrative data

Endpoint:

skin irritation: in vitro / ex vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: well documented, according to OECD guideline and under GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2014

Report date:

2014

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Quino[2,3-b]acridine-6,7,13,14(5H,12H)-tetrone

EC Number:

216-125-0

EC Name:

Quino[2,3-b]acridine-6,7,13,14(5H,12H)-tetrone

Cas Number:

1503-48-6

Molecular formula:

C20H10N2O4

IUPAC Name:

quino[2,3-b]acridine-6,7,13,14(5H,12H)-tetrone

Details on test material:

- Analytical purity:97.59%

Test animals

Species:

other: Epi-200

Strain:

other: normal, human-derived epidermal keratinocytes , cultured to form a model of the human epidermis, consists of organized basal, spinous and granular layers, and a multi-layered stratum corneum

Details on test animals or test system and environmental conditions:

MATERIAL and EQUIPMENT
EpiDerm TM 200 kit: MatTek ln Vitro Life Science Laboratories, Bratislava, Slovakia containing: 24 Epi-200 tissues (reconstructed epidermis): surface 0.6 cm2 cultured in Millicells® 0 1 cm
Tissue for MTTreduction control: Epi-200 tissue that is killed by freezing at -20°C
Assay medium: Dulbecco's modified eagle's medium (DMEM); for the assay and for diluting MTT
Wash buffer: Dulbecco's phosphate buffered saline (P8S), w/o Ca2 + , Mg2 +
Extracting agent: lsopropanol p.a.
Detection agent: 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), 1.0 mg I ml assay medium

Test system

Type of coverage:

other: not applicable

Preparation of test site:

other: not applicable

Vehicle:

other: not applicable

Controls:

other: not applicable

Amount / concentration applied:

25 μL de-ionized water was applied first. Thereafter, a bulk volume of 25 μL of the solid test
material was applied with a sharp spoon and homogeneously distributed with the water

Duration of treatment / exposure:

1h

Observation period:

not applicable

Number of animals:

not applicable

Details on study design:

Pretest for direct MTT reduction:
To determine whether the test substance is able to reduce MTT directly, the test substance will be incubated with the substrate. Approximately 50 ul is added to 0.9 ml of the MTT solution. The mixture is incubated in the dark at about 37 oc for 55 to 65 minutes. When the color of the mixture turns blue/purple, it is assumed that the test substance can directly reduce MTT.
Preincubation of the tissues:
Corrosion test: At least 1 hour but not more than 1.5 hours before test-substance application, tissues are transferred to 6-well plates with 0.9 mL assay medium and preconditioned in the incubator at 37C°.
Irritation test: on the day of arrival in the laboratory, the tissues will be transferred to sterile 6-well plates with 0.9 mL assay medium and preconditioned in the incubator at 37°C. After 1 hour the pre-incubation medium is replaced with fresh medium and preconditioning continues for 18 ± 3 hours.
MTT incubation:
After the incubation / postincubation period, the assay medium is replaced by 0.3 ml MTT solution and the tissues are incubated in the incubator for 3 hours. After incubation, the tissues are washed with PBS to stop the MTT-incubation.
Detection of MTT metabolism:
The formazan that is metabolically produced by the tissues will be extracted by incubation of the tissues in 2 ml isopropanol at room temperature overnight (corrosion test) or for at least 2 hours on a plate shaker (ca. 120 rpm) (irritation test). After shaking the isopropanol extract and piercing the tissues, 2 aliquots of each extract per tissue will be transferred to a 96-well microtiter plate. The optical density will be determined
spectrophotometrically using a filter with a wavelength of 570 nm

Results and discussion

In vitro

Results

Irritation / corrosion parameter:

other: other: Viability (%)

Value:

94

Remarks on result:

other:

Remarks:

Basis: mean. Time point: 1h. Max. score: 100.0. Reversibility: no data. (migrated information)

Applicant's summary and conclusion

Interpretation of results:

not irritating

Remarks:

Migrated information Criteria used for interpretation of results: EU