4,4'-bis(dimethylamino)-4''-(methylamino)trityl alcohol

Endpoint summary

Description of key information

Short term toxicity to fish

An acute toxicity test was conducted for 96 hrs for assessing the effect of test chemical on Zebra fish (Danio rerio). The test was performed in accordance to OECD guideline No. 203 “Fish Acute Toxicity Test”. Zebra fish (Danio rerio) of average length of 2.3 cm was used as a test organism for the study. Test fishes were kept in a static tank in tap water passed through reverse osmosis system, under natural conditions along with proper feed and aeration. During the housing period, test fishes were fed once daily with standard brand fed. Aeration in test vessels was provided till 1 day before the start of the experiment. The test chemical was prepared by dissolving 5000 mg of test chemical in 5000 ml of RO water. After stirring the stock solution was filtered and analytically detected and the concentration found to be 1000 mg/L.  The remaining test solutions were prepared by dilution from the above stock solution. Test chemical concentrations were analytically determined by UV-VIS spectrophotometer. Test chemical conc. used for the study were 0, 0.13, 0.2, 0.3, 0.45, 0.68 mg/l (factor of 1.5, respectively). Total 7 fishes were exposed to test chemical in a 7 lit of glass aquarium containing 4000 ml of potable water. The test vessels were placed in a room at a temperature of 21.4°C, pH of 8.0 and DO of 7.2 mg/l and under a photoperiod of 16:8 hr light: dark conditions, respectively. At regular intervals of 24 hours vessels were observed for any behavioural changes and mortality along with that the DO, pH and temperature was measured. Test chemical was sampled from test vessel for analytical determinations at 0 hour and 96 hours. All the test concentrations were analytical determined and at 96 hours of the exposure durations which were maintained in the range of 80 -120%. Therefore, the analysis of the results was based on nominal concentration. After the 96 hours of exposure, there was no mortality in the control group, and the dissolved oxygen concentration was 85.06% (i.e. above 60%) of the air saturation value throughout the exposure period. Thus, the study fulfilling the validity criterion as mentioned in the OECD TG 203. On the basis of effect of test chemical on mortality of the test organism, the 96 hr median lethal concentration [LC50 (96 h)] for test chemical on Danio rerio (Zebra Fish) was determined to be 0.257 mg/L (nomnal conc.). Thus, based on LC50 value, test chemical was considered as toxic to aquatic fishes and hence, considered to be classified in 'aquatic acute/ chronic 1' category as per the CLP classification criteria.

Short term toxicity to aquatic invertebrates

This study was designed (as per OECD 202, adopted in 2004) to assess the acute toxicity of test chemical following exposure of daphnids up to 48h by static method. The test daphnids were acclimatized 48 hours prior to the test chemical exposure. After exposure on day 0, daphnids were observed for immobilization at 24 and 48 h. M7 medium was used as control, and the same was used for test chemical formulation . 25 mL glass beakers having a solution volume of 20 mL were used in the test. . Main study (using a spacing factor of 2) was conducted using 0 (control), 0.625, 1.25, 2.5, 5 and 10 mg/L mg/L concentrations. 4 replicates/concentration having 5 daphnids/replicate was used for the main study. UV-Visible spectrophotometer method was used for active ingredient analysis along with stability of the test item in the test medium. Test item was found to be stable in the test medium. The active ingredient content results were considered acceptable (80-120%) as during main study 0 h and after 48 hours of exposure, which were found in acceptable range. Hence the results were based on nominal concentration since the deviation in the initial measured concentration didn’t exceed 20%. Environmental parameters such as pH (7.2 -7.6), temperature (20-21°C), dissolve oxygen (7.1 - 7.4 mg/L), hardness (>140 mg CaCO3/L), photoperiod (16 h light- 8 h dark) was maintained in acceptable range throughout the test. Thus, fulfilling the validity criteria. Feed was not provided during the test. Normal behavioural response and no immobilization (0% mortality) were observed up to 48 h followed by control groups but 35%, 95%, 100%, 100% and 100% immobilisation were observed in the test concentrations of 0.625, 1.25, 2.5, 5 and 10 mg/L, respectively. The 48-h EC50 of test chemical to daphnid, Daphnia magna are 0.671 mg/l. The 48-h EC50 of reference item (Potassium dichromate) to daphnid, Daphnia magna(found to be in acceptable range) is 0.65 mg/L. Hence, the results of the test with reference item establish the acceptability of the test system response, test procedures followed, and results obtained with test item. Thus, test chemical was considered as toxic to aquatic invertebrates at environmental relevant concentrations and hence, considered to be classified in 'aquatic acute/chronic category 1' as per the CLP classification criteria.

Toxicity to aquatic algae and cyanobacteria

A freshwater algal growth inhibition test was conducted for 72 hrs for assessing the effect of test chemical on green algae Pseudokirchneriella subcapitata. The test was performed in accordance to OECD guideline No. 201 – Alga growth inhibition test under static condition. Initial cell density of the culture was kept at 10000 cells/ml. OECD medium composed of macronutrients, micronutrients, alkaline EDTA solution and iron solution was used as a growth medium. The test solution was prepared by dissolving 500 mg of test chemical in 500 ml of OECD media, and allowed for stirring for 96 hours, which was then filtered and the final stock solution obtained was 1000 mg/L, verified analytically by UV-Vis Spectrophotometer. To have a better growth and visibility of cells, the initial cell count of the culture was kept 10,000 cells/mL. Further, exposure concentrations of 0, 0.14, 0.22, 0.35, 0.56 and 0.896 mg/l (factor of 1.6), respectively was from the stock test concentrations. Test vessel were placed in orbital shaking incubator for 72 hrs at a temperature of 21 to 24 ± 2°C and with a continuous uniform illumination of 7000 -10000 lux light intensity, respectively. The speed of the orbital shaking incubator was set at a 120 revolutions per minute throughout the study period. The cultures were counted and observed daily with the help of a microscope. Potassium dichromate (K2Cr2O7) was used as a reference substance. The 72 hr EC50 value of the reference substance was determined to be 0.868 mg/l. The biomass in the control vessel have increased exponentially by a factor of 16, the coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 7% (i.e., reported to be 6.03%) and the mean coefficient of variation of specific growth rate was not exceeded 35% (i.e., reported to be 12.43%), respectively thus fulfilling the validity criterion. As the concentration of the test chemical being tested has not been satisfactorily maintained within ± 20 % of the nominal concentration throughout the test. Therefore, the analysis of the results was based on measured concentration. On the basis of growth rate of the test organism, the 72 hr median effect concentration (ErC50) value was determined to be 0.37 mg/l (measuered conc.) (calculated from equation through probit analysis) and 0.57 mg/l (nominal conc.), respectively.Thus, test chemical was considered as toxic to aquatic algae at environmental relevant concentrations and hence, considered to be classified in 'aquatic acute/chronic category 1' as per the CLP classification criteria.

Toxicity to microorganisms

On the basis of the experimental studies of the structurally similar read across chemical and applying the weight of evidence approach, the EC50 value of the test chemical on test organism can be expected to be in the range of 0.53 to > 39.9 mg/l.

Additional information

Short term toxicity to fish

An acute toxicity test was conducted for 96 hrs for assessing the effect of test chemical on Zebra fish (Danio rerio). The test was performed in accordance to OECD guideline No. 203 “Fish Acute Toxicity Test”. Zebra fish (Danio rerio) of average length of 2.3 cm was used as a test organism for the study. Test fishes were kept in a static tank in tap water passed through reverse osmosis system, under natural conditions along with proper feed and aeration. During the housing period, test fishes were fed once daily with standard brand fed. Aeration in test vessels was provided till 1 day before the start of the experiment. The test chemical was prepared by dissolving 5000 mg of test chemical in 5000 ml of RO water. After stirring the stock solution was filtered and analytically detected and the concentration found to be 1000 mg/L.  The remaining test solutions were prepared by dilution from the above stock solution. Test chemical concentrations were analytically determined by UV-VIS spectrophotometer. Test chemical conc. used for the study were 0, 0.13, 0.2, 0.3, 0.45, 0.68 mg/l (factor of 1.5, respectively). Total 7 fishes were exposed to test chemical in a 7 lit of glass aquarium containing 4000 ml of potable water. The test vessels were placed in a room at a temperature of 21.4°C, pH of 8.0 and DO of 7.2 mg/l and under a photoperiod of 16:8 hr light: dark conditions, respectively. At regular intervals of 24 hours vessels were observed for any behavioural changes and mortality along with that the DO, pH and temperature was measured. Test chemical was sampled from test vessel for analytical determinations at 0 hour and 96 hours. All the test concentrations were analytical determined and at 96 hours of the exposure durations which were maintained in the range of 80 -120%. Therefore, the analysis of the results was based on nominal concentration. After the 96 hours of exposure, there was no mortality in the control group, and the dissolved oxygen concentration was 85.06% (i.e. above 60%) of the air saturation value throughout the exposure period. Thus, the study fulfilling the validity criterion as mentioned in the OECD TG 203. On the basis of effect of test chemical on mortality of the test organism, the 96 hr median lethal concentration [LC50 (96 h)] for test chemical on Danio rerio (Zebra Fish) was determined to be 0.257 mg/L (nomnal conc.). Thus, based on LC50 value, test chemical was considered as toxic to aquatic fishes and hence, considered to be classified in 'aquatic acute/ chronic 1' category as per the CLP classification criteria.

Short term toxicity to aquatic invertebrates

This study was designed (as per OECD 202, adopted in 2004) to assess the acute toxicity of test chemical following exposure of daphnids up to 48h by static method. The test daphnids were acclimatized 48 hours prior to the test chemical exposure. After exposure on day 0, daphnids were observed for immobilization at 24 and 48 h. M7 medium was used as control, and the same was used for test chemical formulation . 25 mL glass beakers having a solution volume of 20 mL were used in the test. . Main study (using a spacing factor of 2) was conducted using 0 (control), 0.625, 1.25, 2.5, 5 and 10 mg/L mg/L concentrations. 4 replicates/concentration having 5 daphnids/replicate was used for the main study. UV-Visible spectrophotometer method was used for active ingredient analysis along with stability of the test item in the test medium. Test item was found to be stable in the test medium. The active ingredient content results were considered acceptable (80-120%) as during main study 0 h and after 48 hours of exposure, which were found in acceptable range. Hence the results were based on nominal concentration since the deviation in the initial measured concentration didn’t exceed 20%. Environmental parameters such as pH (7.2 -7.6), temperature (20-21°C), dissolve oxygen (7.1 - 7.4 mg/L), hardness (>140 mg CaCO3/L), photoperiod (16 h light- 8 h dark) was maintained in acceptable range throughout the test. Thus, fulfilling the validity criteria. Feed was not provided during the test. Normal behavioural response and no immobilization (0% mortality) were observed up to 48 h followed by control groups but 35%, 95%, 100%, 100% and 100% immobilisation were observed in the test concentrations of 0.625, 1.25, 2.5, 5 and 10 mg/L, respectively. The 48-h EC50 of test chemical to daphnid, Daphnia magna are 0.671 mg/l. The 48-h EC50 of reference item (Potassium dichromate) to daphnid, Daphnia magna(found to be in acceptable range) is 0.65 mg/L. Hence, the results of the test with reference item establish the acceptability of the test system response, test procedures followed, and results obtained with test item. Thus, test chemical was considered as toxic to aquatic invertebrates at environmental relevant concentrations and hence, considered to be classified in 'aquatic acute/chronic category 1' as per the CLP classification criteria.

Toxicity to aquatic algae and cyanobacteria

A freshwater algal growth inhibition test was conducted for 72 hrs for assessing the effect of test chemical on green algae Pseudokirchneriella subcapitata. The test was performed in accordance to OECD guideline No. 201 – Alga growth inhibition test under static condition. Initial cell density of the culture was kept at 10000 cells/ml. OECD medium composed of macronutrients, micronutrients, alkaline EDTA solution and iron solution was used as a growth medium. The test solution was prepared by dissolving 500 mg of test chemical in 500 ml of OECD media, and allowed for stirring for 96 hours, which was then filtered and the final stock solution obtained was 1000 mg/L, verified analytically by UV-Vis Spectrophotometer. To have a better growth and visibility of cells, the initial cell count of the culture was kept 10,000 cells/mL. Further, exposure concentrations of 0, 0.14, 0.22, 0.35, 0.56 and 0.896 mg/l (factor of 1.6), respectively was from the stock test concentrations. Test vessel were placed in orbital shaking incubator for 72 hrs at a temperature of 21 to 24 ± 2°C and with a continuous uniform illumination of 7000 -10000 lux light intensity, respectively. The speed of the orbital shaking incubator was set at a 120 revolutions per minute throughout the study period. The cultures were counted and observed daily with the help of a microscope. Potassium dichromate (K2Cr2O7) was used as a reference substance. The 72 hr EC50 value of the reference substance was determined to be 0.868 mg/l. The biomass in the control vessel have increased exponentially by a factor of 16, the coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 7% (i.e., reported to be 6.03%) and the mean coefficient of variation of specific growth rate was not exceeded 35% (i.e., reported to be 12.43%), respectively thus fulfilling the validity criterion. As the concentration of the test chemical being tested has not been satisfactorily maintained within ± 20 % of the nominal concentration throughout the test. Therefore, the analysis of the results was based on measured concentration. On the basis of growth rate of the test organism, the 72 hr median effect concentration (ErC50) value was determined to be 0.37 mg/l (measuered conc.) (calculated from equation through probit analysis) and 0.57 mg/l (nominal conc.), respectively.Thus, test chemical was considered as toxic to aquatic algae at environmental relevant concentrations and hence, considered to be classified in 'aquatic acute/chronic category 1' as per the CLP classification criteria.

Toxicity to microorganisms

Data available for its structurally similar read across chemicals has been reviewed to determine the toxic effect of the test chemical on microorganisms. The studies are as mentioned below:

 

Toxicity to microorganisms study was carried out for 30 mins. The study was performed in accordance with the ISO 11348-2 guideline. Bacteria V. fischeri (NRRL B-1117) was supplied as freeze-dried reagent, BioFix Lumi, from Macherey-Nagel, Du¨ ren, Germany. Test chemcal was dissolved in distilled water; osmolality adjusted to 2 % NaCl. Test chemical concentration used for the study was <=39.9 mg/L. Light production from luminescent bacteria was measured with a luminometer equipped with a constant temperature water bath (15°C). On the basis of the effect of test chemical on light production of the test organism V. fischeri, the 30 mins EC50 value was determined to be > 39.9 mg/l.

 

Another toxicity to micro-organism study was carried out using Bacillus subtilis as test organism. On the basis on effect of test chemical on growth rate of the test organism, the EC50 value was determined to be 0.53 mg/l.

 

On the basis of the experimental studies of the structurally similar read across chemical and applying the weight of evidence approach, the EC50 value of the test chemical on test organism can be expected to be in the range of 0.53 to > 39.9 mg/l.

On the basis of the available experimental information of aquatic toxicity studies, it can be concluded that the test chemical was considered as toxic to aquatic organisms at environmental relevant concentrations and hence, considered to be classified in 'aquatic acute/chronic category 1' as per the CLP classification criteria.