Trisodium bis[3-hydroxy-4-[(2-hydroxy-1-naphthyl)azo]-7-nitronaphthalene-1-sulphonato(3-)]chromate(3-)

Specific investigations: other studies

Administrative data

Endpoint:

cytotoxicity

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: ISO 10993, EN 7405: 1997

Principles of method if other than guideline:

The XTT test is based on the cleavage of the yellow tetrazolium salt XTT [= (sodium-3'-(1phenylaminocarbonyl)-3,4-tetrazolium)-bis-(4-metoxy-6-nitro)-benzenesulfonic acid hydrate)] to form an orange water soluble formazan dye by dehydrogenase activity in active mitochondria. This method was first described 1988 by SCUDIERO et al. (1,2) and improved in subsequent years by several other investigators.

GLP compliance:

yes (incl. QA statement)

Type of method:

in vitro

Test material

Test material information

Constituent 1

Reference substance name:

Trisodium bis[3-hydroxy-4-[(2-hydroxy-1-naphthyl)azo]-7-nitronaphthalene-1-sulphonato(3-)]chromate(3-)

EC Number:

260-906-9

EC Name:

Trisodium bis[3-hydroxy-4-[(2-hydroxy-1-naphthyl)azo]-7-nitronaphthalene-1-sulphonato(3-)]chromate(3-)

Cas Number:

57693-14-8

Molecular formula:

C40H20CrN6O14S2.3Na

IUPAC Name:

trisodium bis[3-hydroxy-4-[(2-hydroxy-1-naphthyl)azo]-7-nitronaphthalene-1-sulphonato(3-)]chromate(3-)

Test material form:

solid: particulate/powder

Test animals

Strain:

other: ATCC, CCl 1 NCTC clone 929 (clone of strain l, mouse connective tissue) cellline

Administration / exposure

Route of administration:

other: in vitro culture

Vehicle:

not specified

Duration of treatment / exposure:

24 hours

Frequency of treatment:

once

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

39.1 other: µg/ml

Doses / concentrations 2

Dose / conc.:

78.1 other: µg/ml

Doses / concentrations 3

Dose / conc.:

156.3 other: µg/ml

Doses / concentrations 4

Dose / conc.:

312.5 other: µg/ml

Doses / concentrations 5

Dose / conc.:

625 other: µg/ml

Doses / concentrations 6

Dose / conc.:

1 250 other: µg/ml

Doses / concentrations 7

Dose / conc.:

2 500 other: µg/ml

Doses / concentrations 8

Dose / conc.:

5 000 other: µg/ml

Examinations

Examinations:

Photometric determination of formazan dye from the cleavage of the yellow tetrazolium salt XTT [= (sodium-3'-(1phenylaminocarbonyl)-3,4-tetrazolium)-bis-(4-metoxy-6-nitro)-benzenesulfonic acid hydrate)]

Positive control:

1. Solvent control for positive control: RPMI 1640 + 10 % (v/v) FCS + 10.0 % (v/v) deion. water
2. Solvent control for test item: RPMI + 10 % (v/v) FCS
3. Negative control: RPMI + 10 % (v/v) FCS
4. Positive control: SDS:
3.125 µg/ml; 6.25 µg/ml; 12.5 µg/ml; 25 µg/ml; 50 µg/ml; 100 µg/ml; 125 µg/ml; 250 µg/ml;

Results and discussion

Details on results:

Toxic effects were observed following incubation with test substance from 312.5 µg/ml up to the highest tested concentration (5000 µg/ml). The calculated XTT so value ís 464.4 µg/ml. Even after stringent washing not all of the test ítem could be removed from the cells in the higher concentrations leading to high chemical blank values and there with freak high viability values.

Applicant's summary and conclusion

Conclusions:

The test item possesses a cytotoxic potential.

Executive summary:

Method

This in vitro study was performed to assess the cytotoxic potential of the test item by means of the XTT test using the mouse cell line L929. The following concentrations of the test item were tested: 39.1, 78.1, 156.3, 312.5, 625, 1250, 2500, 5000 µg/ml.

Complete medium (RPMI containing 10 % (v/v) FCS) was used as negative control.

The solvent control for the positive control was also RPMI medium containing 10 % (v/v) FCS and 10.0 % (v/v) deion. water. SDS was used as positive control.

The following concentrations were applied: 3.125, 6.25, 12.5, 25, 50, 100, 125, 250 µg/ml. The incubation time was 24 hours at 37 ± 1.5 °C.

Results

Negatíve control and solvent control showed no reduction in cell viability. The posítíve control (SDS) induced a distinct dose-related reduction in cell viabilíty.

Toxic effects were observed following incubation with the Test item from 312.5 µg/ml up to the highest tested concentration (5000 µg/ml). The calculated XTT50 value ís 464.4 µg/ml. Even after stringent washing not all of the test ítem could be removed from the cells in the higher concentrations leading to high chemical blank values and therewith freak high viability values. In conclusion, ít can be stated that in this study and under the experimental conditions reported, test item possesses a cytotoxic potential.