2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

for read across substance

Justification for type of information:

Data for the target chemical is summarized based on the structurally similar read across chemicals

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

other: as mentioned below

Principles of method if other than guideline:

WoE report is based on two toxicity study of aquatic algae and cyanobacteria for the test chemical :
1)To evaluate short term toxicity of test material on aquatic algae and cyanobacteria
2)Aim of this study was to evaluate the nature of chemical when comes in contact with the test organism. Test was conducted according to the OECD guideline 201.

GLP compliance:

not specified

Specific details on test material used for the study:

IUPAC: 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one
Molecular Formula: C16H8O2S2
Molecular Weight: 296.369 g/mol
Smiles:O=C1c2c(S\C1=C1/Sc3c(cccc3)C1=O)cccc2
InChI:1S/C16H8O2S2/c17-13-9-5-1-3-7-11(9)19-15(13)16-14(18)10-6-2-4-8-12(10)20-16/h1-8H/b16-15-

Analytical monitoring:

not specified

Vehicle:

not specified

Details on test solutions:

2) The solution 200.0 mg.l'1 was prepared by dissolving blue powder in OECD growth medium. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture. The substance was declared as light sensitive.

Test organisms (species):

other: 1) Raphidocelis subcapitata 2) Desmodesmus subspicatus

Details on test organisms:

1) -Method of cultivation : Algae were routinely cultured at 25±1 °C with light in Cyanobacteria BG-11 Freshwater Solution. The culture was split twice a week, adding fresh medium.


2)TEST ORGANISM
- Common name:
- Strain: 86.81 SAG
- Source (laboratory, culture collection): Institute of botany of the ASCR
- Initial biomass concentration: 5x10(3) cells /ml
- Method of cultivation: No data available

ACCLIMATION - No data available

Test type:

static

Water media type:

freshwater

Total exposure duration:

72 h

Test temperature:

1) 25±1 °C 2) 23±2°C

pH:

2) the sample at concentration 200.0 mg/l: pH = 7.9 changed to pH=7.8 during the test,
control: pH = 8.2 changed to pH = 7.8 during the test

Nominal and measured concentrations:

2) 200 mg/l

Details on test conditions:

1) - Test vessel: sterile 24-well plates
- fill volume: 2 ml
- Determination of cell concentrations: Algal density at the beginning and end of treatment was measured with a TC20™ Automated Cell Counter.
-orbital shaker (90 rpm) in a thermostatic chamber.
Light was provided by a 2 W LED unit for each plate.
-Replicates: 3

2) TEST SYSTEM
- Test vessel: 50ml Glass vessel
- Type : closed
- Sample volume: 7.4ml
- Initial cells density: 5x10(3) cells/ml
- No. of vessels per concentration (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Adjustment of pH: No
- Photoperiod: Continuous
- Light intensity and quality: 6000-8000 lx

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: microscope with counting chamber Cyrus I or electronic particle counter.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

72 h

Dose descriptor:

other: ErC50

Effect conc.:

142 mg/L

Nominal / measured:

not specified

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: CI : 126 – 164 mg/l

Duration:

72 h

Dose descriptor:

other: ErC50

Effect conc.:

159 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95 % CI 129.1 - 197.9

Results with reference substance (positive control):

1)potassium dichromate
ErC50 72h CI : 126 – 164 mg/l
2) - Results with reference substance valid
- EC50: 0.69 mg/L (24 hours)

Reported statistics and error estimates:

2) EC50 was calculated using non linear regression by the software Prism 4.0 The test chemical Ammonium iodide is not likely to be toxic to fish atleast in the dose range of 100-405.0
mg/l

Conclusions:

The test chemical 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one is not likely to be toxic to fish atleast in the concentration range of 142 - 159 mg/l

Executive summary:

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the toxicity of aquatic algae and cyanobacteria of the test chemical 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one (522 -75 -8).The studies are as mentioned below:

1) The 72 h algal growth inhibition test with Raphidocelis subcapitata was done according to OECD 201 (2011) and ISO 8692 (2012) guidelines with slight modifications. Algae were routinely cultured at 25±1 °C with light in Cyanobacteria BG-11 Freshwater Solution. The

culture was split twice a week, adding fresh medium. Algae were exposed to the treatment in sterile 24-well plates with 2 mL of each treatment solution to the respective microplate well, for three days.

Untreated ISO formulation freshwater and potassium dichromate (0.10 – 1.8 mg/L) were used as negative and positive controls respectively. Each treatment was replicated three times and randomized within the plate. Algal stock solution containing 1.01E6 cells/mL was added (20 μL) to each well to obtain a starting algal density of 1E4 cells/mL. Plates were incubated for 72 h at 25±1 °C on an orbital shaker (90 rpm) in a thermostatic chamber. Light was provided by a 2 W LED unit for each plate. Algal density at the beginning and end of treatment was measured with a TC20™ Automated Cell Counter (Bio-Rad Laboratories, Inc) . All statistical analyses were done using Prism5 (GraphPad Software, Inc.). One-way ANOVA was used for statistical comparisons with Bonferroni's, Dunnett's and Tukey's post hoc tests for multiple comparisons. Significance was set at p<0.05. REGTOX macro Excel™ for dose-response was used to obtain EC50 values by a non-linear regression analysis with Hill's model. The effect concentration on test material for the aquatic algae was observed to be 142 mg/l. Based on the above value it can be considered that the test material is not toxic to aquatic algae and can be considered to be not classified as per CLP criterion.

2) Aim of this study was to evaluate the nature of chemical test chemical when comes in contact with the test organism Desmodesmus subspicatus (previous name: Scenedesmus subspicatus). Test was conducted according to the OECD guideline 201.

The solution 200.0 mg.l'1 was prepared by dissolving blue powder in OECD growth medium. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture. The substance was declared as light sensitive. With the test substance one positive control Potassium dichromate (K2Cr2O7) was also run simultaneously. After the exposure of chemical, effect concentration EC50 was calculated using nonlinear regression by the software Prism 4.0. Effect on the growth of algae was determine after an exposure period of 72 hrs.

 

The median effective concentration (ErC50) for the test substance, in algae was determined to be 159.8 mg/L on the basis of growth rate inhibition effects in a 72 hour study. Based on the EC50 value, indicates that the substance is likely to be non-hazardous to aquatic algae and cannot be classified as aquatic as per the CLP criteria.

Thus, based on the above summarised studies, 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one and it’s structurally and functionally similar read across substance, it can be concluded that effect concentration value is greater than 1000 mg/L. Thus, comparing this value with the criteria of CLP regulation, 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one cannot be classified

for toxicity for aquatic algae and cyanobacteria .Hence, based on the data available for the structurally and functionally similar

read across, test chemical 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one is not likely to be toxic atleast in the range concentration range of 159 mg/L .

Description of key information

Toxicity to aquatic algae and cyanobacteria:

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the toxicity of aquatic algae and cyanobacteria of the test chemical 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one (522 -75 -8).The studies are as mentioned below:

1) The 72 h algal growth inhibition test with Raphidocelis subcapitata was done according to OECD 201 (2011) and ISO 8692 (2012) guidelines with slight modifications. Algae were routinely cultured at 25±1 °C with light in Cyanobacteria BG-11 Freshwater Solution. The

culture was split twice a week, adding fresh medium. Algae were exposed to the treatment in sterile 24-well plates with 2 mL of each treatment solution to the respective microplate well, for three days.

Untreated ISO formulation freshwater and potassium dichromate (0.10 – 1.8 mg/L) were used as negative and positive controls respectively. Each treatment was replicated three times and randomized within the plate. Algal stock solution containing 1.01E6 cells/mL was added (20 μL) to each well to obtain a starting algal density of 1E4 cells/mL. Plates were incubated for 72 h at 25±1 °C on an orbital shaker (90 rpm) in a thermostatic chamber. Light was provided by a 2 W LED unit for each plate. Algal density at the beginning and end of treatment was measured with a TC20™ Automated Cell Counter (Bio-Rad Laboratories, Inc) . All statistical analyses were done using Prism5 (GraphPad Software, Inc.). One-way ANOVA was used for statistical comparisons with Bonferroni's, Dunnett's and Tukey's post hoc tests for multiple comparisons. Significance was set at p<0.05. REGTOX macro Excel™ for dose-response was used to obtain EC50 values by a non-linear regression analysis with Hill's model. The effect concentration on test material for the aquatic algae was observed to be 142 mg/l. Based on the above value it can be considered that the test material is not toxic to aquatic algae and can be considered to be not classified as per CLP criterion.

2) Aim of this study was to evaluate the nature of chemical test chemical when comes in contact with the test organism Desmodesmus subspicatus (previous name: Scenedesmus subspicatus). Test was conducted according to the OECD guideline 201.

The solution 200.0 mg.l'1 was prepared by dissolving blue powder in OECD growth medium. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture. The substance was declared as light sensitive. With the test substance one positive control Potassium dichromate (K2Cr2O7) was also run simultaneously. After the exposure of chemical, effect concentration EC50 was calculated using nonlinear regression by the software Prism 4.0. Effect on the growth of algae was determine after an exposure period of 72 hrs.

 

The median effective concentration (ErC50) for the test substance, in algae was determined to be 159.8 mg/L on the basis of growth rate inhibition effects in a 72 hour study. Based on the EC50 value, indicates that the substance is likely to be non-hazardous to aquatic algae and cannot be classified as aquatic as per the CLP criteria.

Thus, based on the above summarised studies, 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one and it’s structurally and functionally similar read across substance, it can be concluded that effect concentration value is greater than 1000 mg/L. Thus, comparing this value with the criteria of CLP regulation, 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one cannot be classified

for toxicity for aquatic algae and cyanobacteria .Hence, based on the data available for the structurally and functionally similar

read across, test chemical 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one is not likely to be toxic atleast in the range concentration range of 159 mg/L .

Key value for chemical safety assessment

EC50 for freshwater algae:

159 mg/L

Additional information

Toxicity to aquatic algae and cyanobacteria:

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the toxicity of aquatic algae and cyanobacteria of the test chemical 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one (522 -75 -8).The studies are as mentioned below:

1) The 72 h algal growth inhibition test with Raphidocelis subcapitata was done according to OECD 201 (2011) and ISO 8692 (2012) guidelines with slight modifications. Algae were routinely cultured at 25±1 °C with light in Cyanobacteria BG-11 Freshwater Solution. The

culture was split twice a week, adding fresh medium. Algae were exposed to the treatment in sterile 24-well plates with 2 mL of each treatment solution to the respective microplate well, for three days.

Untreated ISO formulation freshwater and potassium dichromate (0.10 – 1.8 mg/L) were used as negative and positive controls respectively. Each treatment was replicated three times and randomized within the plate. Algal stock solution containing 1.01E6 cells/mL was added (20 μL) to each well to obtain a starting algal density of 1E4 cells/mL. Plates were incubated for 72 h at 25±1 °C on an orbital shaker (90 rpm) in a thermostatic chamber. Light was provided by a 2 W LED unit for each plate. Algal density at the beginning and end of treatment was measured with a TC20™ Automated Cell Counter (Bio-Rad Laboratories, Inc) . All statistical analyses were done using Prism5 (GraphPad Software, Inc.). One-way ANOVA was used for statistical comparisons with Bonferroni's, Dunnett's and Tukey's post hoc tests for multiple comparisons. Significance was set at p<0.05. REGTOX macro Excel™ for dose-response was used to obtain EC50 values by a non-linear regression analysis with Hill's model. The effect concentration on test material for the aquatic algae was observed to be 142 mg/l. Based on the above value it can be considered that the test material is not toxic to aquatic algae and can be considered to be not classified as per CLP criterion.

2) Aim of this study was to evaluate the nature of chemical test chemical when comes in contact with the test organism Desmodesmus subspicatus (previous name: Scenedesmus subspicatus). Test was conducted according to the OECD guideline 201.

The solution 200.0 mg.l'1 was prepared by dissolving blue powder in OECD growth medium. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture. The substance was declared as light sensitive. With the test substance one positive control Potassium dichromate (K2Cr2O7) was also run simultaneously. After the exposure of chemical, effect concentration EC50 was calculated using nonlinear regression by the software Prism 4.0. Effect on the growth of algae was determine after an exposure period of 72 hrs.

 

The median effective concentration (ErC50) for the test substance, in algae was determined to be 159.8 mg/L on the basis of growth rate inhibition effects in a 72 hour study. Based on the EC50 value, indicates that the substance is likely to be non-hazardous to aquatic algae and cannot be classified as aquatic as per the CLP criteria.

Thus, based on the above summarised studies, 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one and it’s structurally and functionally similar read across substance, it can be concluded that effect concentration value is greater than 1000 mg/L. Thus, comparing this value with the criteria of CLP regulation, 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one cannot be classified

for toxicity for aquatic algae and cyanobacteria .Hence, based on the data available for the structurally and functionally similar

read across, test chemical 2-(3-oxobenzo[b]thien-2(3H)-ylidene)benzo[b]thiophene-3(2H)-one is not likely to be toxic atleast in the range concentration range of 159 mg/L .