4',5'-dibromo-3',6'-dihydroxyspiro[isobenzofuran-1(3H),9'-[9H]xanthene]-3-one

Toxicity to microorganisms

Administrative data

Endpoint:

toxicity to microorganisms

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Data is from peer reviewed journal

Data source

Reference

Reference Type:

publication

Title:

In Vitro Antibacterial Activities of Phloxine B and Other Halogenated Fluoresceins against Methicillin-Resistant Staphylococcus aureus.

Author:

Avraham Rasooly and Adrian Weisz

Year:

2002

Bibliographic source:

ANTIMICROBIAL AGENTS AND CHEMOTHERAPY, Nov. 2002, p. 3650–3653

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: refer below principle

Principles of method if other than guideline:

Toxicity to micro-organisms study was conducted on Staphylococcus aureus of enterotoxigenic strain ATCC 13565.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

Fluorescein sodium

IUPAC Name:

Fluorescein sodium

Constituent 2

Reference substance name:

Disodium 2-(3-oxo-6-oxidoxanthen-9-yl)benzoate

EC Number:

208-253-0

EC Name:

Disodium 2-(3-oxo-6-oxidoxanthen-9-yl)benzoate

Cas Number:

518-47-8

Molecular formula:

C20H12O5.2Na

IUPAC Name:

disodium 3-oxo-3H-spiro[2-benzofuran-1,9'-xanthene]-3',6'-diolate

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

- Name of test material (as cited in study report): Fluorescein sodium - Molecular formula (if other than submission substance): C20-H10-O5.2Na- Molecular weight (if other than submission substance): 376.274 g/mol- Smiles notation (if other than submission substance): c1(c2c(oc3c1ccc(c3)[O-])cc(=O)cc2)c1c(cccc1)C(=O)[O-].[Na+].[Na+]- InChl (if other than submission substance): 1S/C20H12O5.2Na/c21-11-5-7-15-17(9-11)25-18-10-12(22)6-8-16(18)19(15)13-3-1-2-4-14(13)20(23)24;;/h1-10,21H,(H,23,24);;/q;2*+1/p-2- Substance type: Organic - Physical state: Solid

Sampling and analysis

Analytical monitoring:

not specified

Details on sampling:

- Concentrations: 100 µg/ml (0.1 mg/ml)- Sampling method: For the preparation of dye solution, a test portion (approximately 10 mg) was placed in a 10-ml volumetric flask, which was wrapped in aluminum foil, and dissolved by adding aqueous sodium hydroxide (approx. 0.4% NaOH) (1 ml) and water (9 ml) (high-pressure liquid chromatography grade). The solution obtained by this procedure were found to be stable for at least 6 months in refrigerator at 4ᵒC.- Sample storage conditions before analysis: No data available

Test solutions

Vehicle:

yes

Test organisms

Test organisms (species):

other: Staphylococcus aureus (S. aureus) ATCC 13565

Details on inoculum:

- Laboratory culture: The test organism S. aureus ATCC 13565 was obtained from American Type Culture Collection, Manassas, Va.- Method of cultivation: For the study, a 300-µl aliquot of dye solution (1 mg/ml) was mixed with 0.7 ml of BHI and then added to a tube of diluted culture. The tubes were aerated vigorously, and the OD was measured every 45 min to check culture growth. The survival rate was measured by plating serial dilutions of a sample taken during the experiment on BHI plates.- Preparation of inoculum for exposure: For inoculum preparation, brain heart infusion broth (BHI) was used. Cultures were incubated shaking (200 rpm) at 37°C. All the experiments were conducted under conditions of standard room illumination (fluorescent ceiling light, Sylvania Octron 4100K) in tubes containing 3 ml of medium. Growth was analyzed by measuring turbidity (absorbance at a wavelength of 660 to 680 nm) with aspectrophotometer (Spectronic 21D). Experiments were initiated with ̴0.2 ml of inoculum from a fresh culture grown aerobically at an optical density (OD) of ̴1. Colony counts were measured on BHI plates.

Study design

Test type:

not specified

Water media type:

freshwater

Limit test:

no

Total exposure duration:

250 min

Post exposure observation period:

OD was measured every 45 mins to check culture growth.

Test conditions

Hardness:

No data available

Test temperature:

No data available

pH:

No data available

Dissolved oxygen:

No data available

Salinity:

No data available

Nominal and measured concentrations:

Nominal concentrations

Details on test conditions:

TEST SYSTEM- Test vessel: Test tubes- Aeration: The tubes containing the test solutions and cultures were aerated vigorously.TEST CONCENTRATIONS- Test concentrations: 100 µg/ml (0.1 mg/ml)

Reference substance (positive control):

not specified

Results and discussion

Effect concentrations

Duration:

250 min

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: The test substance do not have any activity against S. aureus i.e, no difference was observed between the treated test tubes and the control tubes.

Details on results:

Test result reported in mg/ml i.e 0.1 mg/ml when converted to mg/l it becomes 100 mg/l

Results with reference substance (positive control):

No data available

Reported statistics and error estimates:

No data available

Applicant's summary and conclusion

Validity criteria fulfilled:

not specified

Conclusions:

In Toxicity to micro-organisms study as the test substance Fluorescein sodium do not have any activity against S. aureus i.e, no difference was observed between the treated test tubes and the control tubes, the NOEC value was found to be 100 mg/l.

Executive summary:

Toxicity to micro-organisms study was conducted on Staphylococcus aureusof enterotoxigenic strain ATCC 13565.

 

For the preparation of dye solution, a test portion (approximately 10 mg) was placed in a 10-ml volumetric flask, which was wrapped in aluminum foil, and dissolved by adding aqueous sodium hydroxide (approx. 0.4% NaOH) (1 ml) and water (9 ml).

 

Inoculum was prepared inbrain heart infusion broth (BHI). Cultures were incubated shaking (200 rpm) at 37°C. All the experiments were conducted under conditions of standard room illumination (fluorescent ceiling light, Sylvania Octron 4100K) in tubes containing        3 ml of medium. Growth was analyzed by measuring turbidity (absorbance at a wavelength of 660 to 680 nm) with a spectrophotometer (Spectronic 21D). Experiments were initiated with ̴0.2 ml of inoculum from a fresh culture grown aerobically at an optical density (OD) of ̴1. Colony counts were measured on BHI plates.

 

For the study,a 300-µl aliquot of dye solution (1 mg/ml) was mixed with 0.7 ml of BHI and then added to a tube of diluted culture. The tubes were aerated vigorously, and the OD was measured every 45 min to check culture growth. The survival rate was measured by plating serial dilutions of a sample taken during the experiment on BHI plates.

 

During the study period of 250 mins, OD was measured every 45 mins to check culture growth.

 

Experiment was performed in triplicate on separate days to ascertain reproducibility.

 

As the test substance do not have any activity againstS. aureusi.e, no difference was observed between the treated test tubes and the control tubes, the NOEC value was found to be 100 mg/l.