Benzimidazole-2-thiol

Endpoint summary

Description of key information

Short term toxicity to fish:

Study was conducted to assess the effect of test chemical on the mortality of fish Danio rerio. Test conducted according to OECD Guideline 203 (Fish, Acute Toxicity Test).The test substance was soluble in water. Therefore, the test solution was prepared by dissolving 1 gm of the test substance in 1 liters of potable water (passed through reverse osmosis system) with 1 hour continuous stirring for achieving test concentrations of 6.25 mg/L,12.5mg/L,25mg/L,50mg/L and 100mg/L, respectively.

Bowl aquaria containing 2 liters of potable water (passed through reverse osmosis system) were loaded with 8 fishes. A static procedure was used for the study and it was conducted in compliance with the OECD guideline 203. After 96 hours of exposure to test item to various nominal test concentrations, LC50 was determine to be> 50 mg/L but < 100 mg/L.  . Based on the LC50, it can be consider that the chemical was toxic and can be consider to be classified aq aquatic chronic 3 as per the CLP classification criteria.

Short-term toxicity to aquatic invertebrates:

Based on the prediction done by EPI suite, ECOSAR version 1.1, on the basis of similarity of structure to chemicals for which the aquatic toxicity has been previously measured by structure-activity relationships (SARs) program, the LC 50 value for short term toxicity to aquatic invertebrates was predicted.

On the basis of EPI suite, ECOSAR version 1.1, the LC 50 value for short term toxicity to aquatic invertebrates was predicted to be 80.912 mg/l for test material in 48 hrs.

Based on this value it can be concluded that the substance is considered to be  toxic to aquatic environment and can be classified in aquatic chronic 3 category as per the criteria mentioned in CLP regulation.

Toxicity to aquatic algae and cyanobacteria:

The study was designed to assess the toxic effects of the test compound on the green algaPseudokirchneriella subcapitata. Test was conducted in compliance with the OECD guideline 201 (Alga, Growth Inhibition Test).

Test was carried out in 100mL conical flasks which were carefully autoclaved and sterilized. The test solution in each of these test vessels was kept constant which is 60 ml so that a sufficient amount of head space was left.The stock solution of test material was prepared by adding 300 mg of test item directly in 300 ml of OECD Media.The dose concentrations were selected on the basis solubility of test item in OECD media. To prepare 200 mg/L of stock for test, 125.554 ml from above stock and volume was makeup to 300 ml with OECD medium. The remaining test solutions were prepared by dilution from the above stock solution 200 mg/l. The recommended cell density of the culture as per OECD guideline was 5 X 103 to 1 X 104cells/ml.

For the assessment of algal growth, the test was conducted in replicates. The control flask was maintained in triplicates as recommended in the OECD guideline and the test concentration were selected in geometric series which were maintained in duplicates. To obtain a quantitative concentration-response relationship by regression analysis, a linearizing transformation of the response data into probit was performed. Using the same, effective concentration (EC) were determined.

Algal growth was calculated daily by counting the cells microscopically with the help of haemocytometer. For microscopic observations the cultures were observed daily with the help of a microscope to verify a normal and healthy appearance of the algal culture and also to observe any abnormal appearance of the algae (as may be caused by the exposure of the test item). Apart from this, the cell count of each test vessel was also noted with the help of a microscope and haemocytometer. By spectrophotometer the absorbance values of each test vessel and control vessel was noted at 680nm.The BBM was taken as blank for both control and test vessels. The absorbance value of each vessel was in line with the average specific growth rate.

As per OECD 201, the biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72 hr test period. This corresponds to a specific growth rate of 0.92 per day. Thus, the observed specific growth rate in the control cultures during the experiment was 0.358 per day. Secondly the mean coefficient of variation for section by section specific growth rates (days 0-1, 1-2 & 2-3, for 72 hr tests) in the control cultures must not exceed 35%. Thus, the observed mean coefficient of variation in the control cultures during the experiment was 33.42%. Thirdly the coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 10%. Thus, the observed coefficient of variation of average specific growth rates during the experiment in control cultures was 8.26%. Hence, the test is considered valid as per OECD guideline, 201

After 72 hours of exposure to test item to6.25mg/l,12.5mg/l,25mg/l,50mg/l,100mg/l,200mg/lnominal test concentrations, EC10 was determine to be 15.740 mg/l graphically and through probit analysis. At concentration 200 mg/l 47.44 % inhibition was observed. Based on the percentage inhibition of growth, it can be concluded that the chemical was not toxic and can be consider to be not classified as per the CLP classification criteria.

 

Toxicity to microorganisms:

The test exposes luminescent organisms in Microtox Acute Reagent to aqueous samples, and measures the increase or decrease in light output by the test organisms.Reagent contains living luminescent bacteria that have been grown under optimal conditions, harvested,and then lyophilized (freeze-dried). The lyophilized bacteria are rehydrated with Reconstitution Solution to provide a ready-to-use suspension of organisms. The test system measures the light output of the luminescent bacteria after they have been challenged by a sample and compares it to the light output of a control (reagent blank) that contains no sample. A difference in light output (between the sample and the control) is attributed to the effect of the sample on the organisms.The median effective concentration (EC50) for the test substance, in microorganism (Photobacterium phosphoreum) in a 5-30 minute study on the basis of viability and sensitivity effect was observed to be 86.4 mg/L.

Additional information

Short term toxicity to fish:

Study was conducted to assess the effect of test chemical on the mortality of fish Danio rerio. Test conducted according to OECD Guideline 203 (Fish, Acute Toxicity Test).The test substance was soluble in water. Therefore, the test solution was prepared by dissolving 1 gm of the test substance in 1 liters of potable water (passed through reverse osmosis system) with 1 hour continuous stirring for achieving test concentrations of 6.25 mg/L,12.5mg/L,25mg/L,50mg/L and 100mg/L, respectively.

Bowl aquaria containing 2 liters of potable water (passed through reverse osmosis system) were loaded with 8 fishes. A static procedure was used for the study and it was conducted in compliance with the OECD guideline 203. After 96 hours of exposure to test item to various nominal test concentrations, LC50 was determine to be> 50 mg/L but < 100 mg/L.  . Based on the LC50, it can be consider that the chemical was toxic and can be consider to be classified aq aquatic chronic 3 as per the CLP classification criteria.

Short-term toxicity to aquatic invertebrates:

The effect of test material was evaluated based on the predicted data for test chemical and further the prediction was supported by data for structurally similar read across substance.

Based on the prediction done by EPI suite, ECOSAR version 1.1, on the basis of similarity of structure to chemicals for which the aquatic toxicity has been previously measured by structure-activity relationships (SARs) program, the LC 50 value for short term toxicity to aquatic invertebrates was predicted.

On the basis of EPI suite, ECOSAR version 1.1, the LC 50 value for short term toxicity to aquatic invertebrates was predicted to be 80.912 mg/l for test material in 48 hrs.Based on this value it can be concluded that the substance is considered to be  toxic to aquatic environment and can be classified in aquatic chronic 3 category as per the criteria mentioned in CLP regulation.

The above prediction was supported by data for structurally similar read across substance , experimental study for inhibition of the mobility of daphnids was carried out with the benzimidazole according to OECD Guideline 202. The animals used for the test shall be less than 24 h old and should not be of first brood progeny. Test solutions of required concentration were prepared by mixing the stock solution of the test sample with reconstituted test water. The test substance was tested at the concentrations 0, 7.5, 15, 30, 60 and 120 mg/L. The test was performed under static conditions in a static fresh water system at 20±1°C temperature. Effects on immobilisation were observed for 48 hours. EC50 was calculated using non linear regression by the software Prism 4.0. After the experiment, the median effective concentration (EC50) for the test substance, benzimidazole, in Daphnia magna was determined to be 55.5 mg/L on the basis of immobilisation effects. Based on the EC50 value chemical consider to be toxic and classified as aquatic chronic 3 as per the CLP classification criteria.

Further for another structurally similar read across substance ,determination of the inhibition of the mobility of daphnids was carried out with the substance according to OECD Guideline 202. The stock solution (200 g/L) was prepared by dissolving white powder in DMSO. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with reconstituted test water. The test substance was tested at the concentrations 0, 5, 10, 20, 30, 40, 80 mg/L and the immobilisation effects were observed for 48 hours. Effects on immobilisation were observed for 48 hours. 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. The median effective concentration (EC50) for the test substance, in Daphnia magna was determined to be 26 mg/L on the basis of mobiity inhibition effects in a 48 hour study. This value indicates that the substance is likely to be hazardous to aquatic invertebrates can can be classified as Aquatic chronic category 3 as per the CLP criteria.

The above data was supported by data for another structurally similar read across substance , determination of the inhibition of the mobility of daphnids was carried out with the substance,according to OECD Guideline 202.The stock solution (100 g/L) was prepared by dissolving white powder in acetone. The test solutions of required concentrationas were prepared by mixing the stock solution of the test sample with reconstituted test water. The test solution were kept 10 min in ultrasonic bath.The test substance was tested at the concentrations 0, 6, 10, 17, 29, 50 and 100 mg/L. Effects on immobilisation were observed for 48 hours. The median effective concentration (EC50) for the test substance, in Daphnia magna was determined to be 29.8 mg/L for immobilisation effects.

These value indicates that the substance is likely to be hazardous to aquatic invertebrates and can be classified as Aquatic Chronic category 3 as per the CLP criteria.

Toxicity to aquatic algae and cyanobacteria:

The study was designed to assess the toxic effects of the test compound on the green algaPseudokirchneriella subcapitata. Test was conducted in compliance with the OECD guideline 201 (Alga, Growth Inhibition Test).

Test was carried out in 100mL conical flasks which were carefully autoclaved and sterilized. The test solution in each of these test vessels was kept constant which is 60 ml so that a sufficient amount of head space was left.The stock solution of test material was prepared by adding 300 mg of test item directly in 300 ml of OECD Media.The dose concentrations were selected on the basis solubility of test item in OECD media. To prepare 200 mg/L of stock for test, 125.554 ml from above stock and volume was makeup to 300 ml with OECD medium. The remaining test solutions were prepared by dilution from the above stock solution 200 mg/l. The recommended cell density of the culture as per OECD guideline was 5 X 103 to 1 X 104cells/ml.

For the assessment of algal growth, the test was conducted in replicates. The control flask was maintained in triplicates as recommended in the OECD guideline and the test concentration were selected in geometric series which were maintained in duplicates. To obtain a quantitative concentration-response relationship by regression analysis, a linearizing transformation of the response data into probit was performed. Using the same, effective concentration (EC) were determined.

Algal growth was calculated daily by counting the cells microscopically with the help of haemocytometer. For microscopic observations the cultures were observed daily with the help of a microscope to verify a normal and healthy appearance of the algal culture and also to observe any abnormal appearance of the algae (as may be caused by the exposure of the test item). Apart from this, the cell count of each test vessel was also noted with the help of a microscope and haemocytometer. By spectrophotometer the absorbance values of each test vessel and control vessel was noted at 680nm.The BBM was taken as blank for both control and test vessels. The absorbance value of each vessel was in line with the average specific growth rate.

As per OECD 201, the biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72 hr test period. This corresponds to a specific growth rate of 0.92 per day. Thus, the observed specific growth rate in the control cultures during the experiment was 0.358 per day. Secondly the mean coefficient of variation for section by section specific growth rates (days 0-1, 1-2 & 2-3, for 72 hr tests) in the control cultures must not exceed 35%. Thus, the observed mean coefficient of variation in the control cultures during the experiment was 33.42%. Thirdly the coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 10%. Thus, the observed coefficient of variation of average specific growth rates during the experiment in control cultures was 8.26%. Hence, the test is considered valid as per OECD guideline, 201

After 72 hours of exposure to test item to6.25mg/l,12.5mg/l,25mg/l,50mg/l,100mg/l,200mg/lnominal test concentrations, EC10 was determine to be 15.740 mg/l graphically and through probit analysis. At concentration 200 mg/l 47.44 % inhibition was observed. Based on the percentage inhibition of growth, it can be concluded that the chemical was not toxic and can be consider to be not classified as per the CLP classification criteria.

Toxicity to microorganisms:

The test exposes luminescent organisms in Microtox Acute Reagent to aqueous samples, and measures the increase or decrease in light output by the test organisms.Reagent contains living luminescent bacteria that have been grown under optimal conditions, harvested,and then lyophilized (freeze-dried). The lyophilized bacteria are rehydrated with Reconstitution Solution to provide a ready-to-use suspension of organisms. The test system measures the light output of the luminescent bacteria after they have been challenged by a sample and compares it to the light output of a control (reagent blank) that contains no sample. A difference in light output (between the sample and the control) is attributed to the effect of the sample on the organisms.The median effective concentration (EC50) for the test substance, in microorganism (Photobacterium phosphoreum) in a 5-30 minute study on the basis of viability and sensitivity effect was observed to be 86.4 mg/L.