Trisodium 8-hydroxypyrene-1,3,6-trisulphonate

Endpoint summary

Description of key information

Hydrolysis

On the basis of the experimental studies of the structurally and functionally similar read across chemical and applying the weight of evidence approach, the hydrolysis half-life value of the test chemical can be expected to be > 5 days at pH 4, 7 and 9 & at a temperature of 50⁰C, respectively or > 1 years at pH 7 & at a temperature of 20⁰C, respectively.

Biodegradation in water

28-days Closed Bottle test following the OECD guideline 301 D to determine the ready biodegradability of the test item (Experimental study report, 2017). The study was performed at a temperature of 20°C. The test system included control, test item and reference item. Domestic waste water, surface soil and soil samples from polluted sites receiving predominantly domestic waste was collected and were mixed to get diluted suspension. The inoculum was kept aerobic until being used for experiment by supplying organic and inorganic sources required by micro flora to sustain at controlled laboratory conditions. This gave the bacterial count as 107 to 108 CFU/ml. At the regular interval microbial plating was also performed on agar to confirm the vitality and CFU count of microorganism. The concentration of test and reference item (Sodium Benzoate) chosen for both the study was 4 mg/L, while that of inoculum was 32 ml/l. OECD mineral medium was used for the study. ThOD (Theoretical oxygen demand) of test and reference item was determined by calculation. % degradation was calculated using the values of BOD and ThOD for test item and reference item. The % degradation of procedure control (reference item) was also calculated using the values of BOD & ThOD and was determined to be 63.85% at 20 ± 1°C. Degradation of Sodium Benzoate exceeds 78.91% on 7 days & 61.44% on 14th day. As the fluctuation of degradation due to D.O depletion in the blank. The activity of the inoculum was thus verified and the test can be considered as valid. The BOD28 value of test chemical was observed to be 0.44 mgO2/mg. ThOD was calculated as 1.14 mgO2/mg. Accordingly, the % degradation of the test item after 28 days of incubation at 20 ± 1°C according to Closed Bottle test was determined to be 38.59%. Based on the results, the test item, under the test conditions, was considered to be primary inherently biodegradable in nature.

Bioaccumulation: aquatic / sediment

Bioaccumulation endpoint can also be considered for waiver as per inaccordance with column 2 of Annex IX of the REACH regulation, testing for this endpoint is scientifically not necessary and does not need to be conducted since the test chemical has a low potential for bioaccumulation based on logKow ≤ 3.

Adsorption / desorption

Adsorption study was conducted for evaluating the adsorption capacity of test chemical (Einat Magal et. al., 2008). Adsorption study was performed according to batch experiments in sediments at a temperature of 25 deg.C.Two types of sediments were used for the sorption experiments:First, a group of pure minerals include clays (bentonite,kaolinite), quartz, calcite and dolomite. Two different grain sizes of quartz, calcite and dolomite were used.Second, a group representing natural sediments. Samples of sediments were collected from three boreholes drilled along the Dead Sea shore. Test chemical conc. used for the study were 0, 50 ,100, 150, 200, 250 mg/l, respectively.A sediment sample and dyed solution were placed in a 50 ml tube. The tubes were continuously rotated 360 deg at 8 rpm by Labquake tube rotator (Thermolyne, USA) in the dark at room temperature (17–25 deg C) for 12–14 h in order to achieve equilibrium concentration. In the first set of experiments, 5 g of sediment was placed together with 5 ml of solution, except for the clays (bentonite and kaolinite) for which, due to difficulty of separating liquids from the sediment, a higher volume of solution was needed (20 ml for bentonite and 10 ml for kaolinite).The solution was removed from the sediments by centrifuging for 20 min at a rate of 3000 rpm and was analyzed by fluorescence spectrophotometry (Cary Eclipse Fluorescence Spectrophotometer, Varian[1], Palo Alto, CA).The concentration of the dyes in the samples was determined separately for each experiment by using calibration curves prepared with solutions of similar salinity and composition to those of the experiment solutions. Blanks were prepared by repeating experimental procedures with solutions without dyes. The fluorescence intensities (FI) of the blanks were analyzed and calculated to determine the ‘‘apparent dye concentration’’ from the FI measurements. It was found that except for Naph, FI of the blank solutions was close to zero. The FI’s in the emission–excitation wavelength similar to that of Naph in the blank solutions were found to be comparable in values to 20–30 ppb. The amount of dye sorption on sediments equals the amount of dye loss after mixing with sediment during the batch experiment. Other dye losses such as dye precipitation are thought to be negligible since in the experiment the dye concentrations were a few orders of magnitude lower than its solubility (a few grams per liter). Dye sorption was calculated as percentage of initial concentration. Percent adsorption of test chemical was 29–53% (0.01 DSW), 38–59% (0.5 DSW) and 64–88% (DSW), except for sample B3 (41%, 83% and 98%, 0.01 DSW, 0.5 DSW and DSW, respectively). On the basis of this, test chemical have low to moderate sorption on sediments and therefore have moderate to slow migration potential to groundwater.

Additional information

Hydrolysis

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the half-life of the test chemical. The studies are as mentioned below:

 

The half-life of the test chemical was determined at different pH range. The study was performed according to OECD Guideline 111 (Hydrolysis as a Function of pH) at a temperature of 50°C. The average percentage recovery of the test chemical after 5 days was determined to be 99.0, 99.5 and 98.7% at pH 4, 7 and 9, respectively. As no hydrolysis of test item was observed for a period of 5 days, the half-lives was determined to be > 5 days at pH 4, 7 and 9 & at a temperature of 50⁰C, respectively. Based on the half-life values, it is concluded that the test substance is not hydrolysable.

 

In an another study, the half-life of the test chemical was determined at pH 7.0. The study was performed at a temperature of 50°C. The only functional group present is the amide group, which hydrolyzes slowly at neutral pH. The half-life value of the test chemical was determined to be > 1 years at pH 7 & at a temperature of 20⁰C, respectively. Based on the half-life value, it is concluded that test chemical is not hydrolysable.

 

On the basis of the experimental studies of the structurally and functionally similar read across chemical and applying the weight of evidence approach, the hydrolysis half-life value of the test chemical can be expected to be > 5 days at pH 4, 7 and 9 & at a temperature of 50⁰C, respectively or > 1 years at pH 7 & at a temperature of 20⁰C, respectively.

Biodegradation in water

28-days Closed Bottle test following the OECD guideline 301 D to determine the ready biodegradability of the test item (Experimental study report, 2017). The study was performed at a temperature of 20°C. The test system included control, test item and reference item. Domestic waste water, surface soil and soil samples from polluted sites receiving predominantly domestic waste was collected and were mixed to get diluted suspension. The inoculum was kept aerobic until being used for experiment by supplying organic and inorganic sources required by micro flora to sustain at controlled laboratory conditions. This gave the bacterial count as 107 to 108 CFU/ml. At the regular interval microbial plating was also performed on agar to confirm the vitality and CFU count of microorganism. The concentration of test and reference item (Sodium Benzoate) chosen for both the study was 4 mg/L, while that of inoculum was 32 ml/l. OECD mineral medium was used for the study. ThOD (Theoretical oxygen demand) of test and reference item was determined by calculation. % degradation was calculated using the values of BOD and ThOD for test item and reference item. The % degradation of procedure control (reference item) was also calculated using the values of BOD & ThOD and was determined to be 63.85% at 20 ± 1°C. Degradation of Sodium Benzoate exceeds 78.91% on 7 days & 61.44% on 14th day. As the fluctuation of degradation due to D.O depletion in the blank. The activity of the inoculum was thus verified and the test can be considered as valid. The BOD28 value of test chemical was observed to be 0.44 mgO2/mg. ThOD was calculated as 1.14 mgO2/mg. Accordingly, the % degradation of the test item after 28 days of incubation at 20 ± 1°C according to Closed Bottle test was determined to be 38.59%. Based on the results, the test item, under the test conditions, was considered to be primary inherently biodegradable in nature.

Bioaccumulation: aquatic / sediment

Bioaccumulation endpoint can also be considered for waiver as per inaccordance with column 2 of Annex IX of the REACH regulation, testing for this endpoint is scientifically not necessary and does not need to be conducted since the test chemical has a low potential for bioaccumulation based on logKow ≤ 3.

Adsorption / desorption

Experimental result of the test chemical and various supporting studies for its structurally similar read across substance were reviewed for the adsorption end point which are summarized as below:

 

In an experimental key study from peer reviewed journal (Einat Magal et. al., 2008), adsorption experiment was conducted for evaluating the adsorption capacity of test chemical. Adsorption study was performed according to batch experiments in sediments at a temperature of 25 deg.C. Two types of sediments were used for the sorption experiments: First, a group of pure minerals include clays (bentonite,kaolinite), quartz, calcite and dolomite. Two different grain sizes of quartz, calcite and dolomite were used. Second, a group representing natural sediments. Samples of sediments were collected from three boreholes drilled along the Dead Sea shore. Test chemical conc. used for the study were 0, 50 ,100, 150, 200, 250 mg/l, respectively. A sediment sample and dyed solution were placed in a 50 ml tube. The tubes were continuously rotated 360 deg at 8 rpm by Labquake tube rotator (Thermolyne, USA) in the dark at room temperature (17–25 deg C) for 12–14 h in order to achieve equilibrium concentration. In the first set of experiments, 5 g of sediment was placed together with 5 ml of solution, except for the clays (bentonite and kaolinite) for which, due to difficulty of separating liquids from the sediment, a higher volume of solution was needed (20 ml for bentonite and 10 ml for kaolinite).The solution was removed from the sediments by centrifuging for 20 min at a rate of 3000 rpm and was analyzed by fluorescence spectrophotometry (Cary Eclipse Fluorescence Spectrophotometer, Varian[1], Palo Alto, CA).The concentration of the dyes in the samples was determined separately for each experiment by using calibration curves prepared with solutions of similar salinity and composition to those of the experiment solutions. Blanks were prepared by repeating experimental procedures with solutions without dyes. The fluorescence intensities (FI) of the blanks were analyzed and calculated to determine the ‘‘apparent dye concentration’’ from the FI measurements. It was found that except for Naph, FI of the blank solutions was close to zero. The FI’s in the emission–excitation wavelength similar to that of Naph in the blank solutions were found to be comparable in values to 20–30 ppb. The amount of dye sorption on sediments equals the amount of dye loss after mixing with sediment during the batch experiment. Other dye losses such as dye precipitation are thought to be negligible since in the experiment the dye concentrations were a few orders of magnitude lower than its solubility (a few grams per liter). Dye sorption was calculated as percentage of initial concentration. Percent adsorption of test chemical was 29–53% (0.01 DSW), 38–59% (0.5 DSW) and 64–88% (DSW), except for sample B3 (41%, 83% and 98%, 0.01 DSW, 0.5 DSW and DSW, respectively). On the basis of this, test chemical have low to moderate sorption on sediments and therefore have moderate to slow migration potential to groundwater.

 

In a supporting study from study report (2016), adsorption coefficient Koc in soil and in sewage sludge of test chemical was determined by the Reverse Phase High Performance Liquid Chromatographic method according to OECD Guideline No. 121 for testing of Chemicals. The solutions of the test substance and reference substances were prepared in appropriate solvents. A test item solution was prepared by accurately weighing 50mg of test item and diluted with mobile phase up to 100ml. Thus, the test solution concentration was 500mg/l. The pH of test substance was 7.18. Each of the reference substance and test substance were analysed by HPLC at 210 nm. After equilibration of the HPLC system, Urea was injected first, the reference substances were injected in duplicate, followed by the test chemical solution in duplicate. Reference substances were injected again after test sample, no change in retention time of reference substances was observed. Retention time tR were measured, averaged and the decimal logarithms of the capacity factors k were calculated. The graph was plotted between log Koc versus log k(Annex - 2).The linear regression parameter of the relationship log Koc vs log k were also calculated from the data obtained with calibration samples and therewith, log Koc of the test substance was determined from its measured capacity factor. The reference substances were chosen according to estimated Koc range of the test substance and generalized calibration graph was prepared. The reference substances were 4-chloroaniline, 4-methylaniline, N methyl aniline, 2-Nitrophenol, Nitrobenzene, 4-Nitrobenzamide, N,N-dimethylbenzamide, N-Methylbenzamide, Benzamide, Phenanthrene having Koc value ranging from 1.239 to 4.09. The Log Koc value of test chemical was determined to be 3.024 ± 0.020 at 25°C. This log Koc value indicates that the test chemical has a moderate sorption to soil and sediment and therefore have slow migration potential to ground water.

 

For the test chemical, the adsorption coefficient Koc in soil and in sewage sludge of test chemical was determined by the Reverse Phase High Performance Liquid Chromatographic method according to OECD Guideline No. 121 for testing of Chemicals (Experimental study report, 2016). The solutions of the test substance and reference substances were prepared in appropriate solvents. A test item solution was prepared by accurately weighing 50mg of test item and diluted with mobile phase up to 100ml. Thus, the test solution concentration was 500mg/l. The pH of test substance was 8.10. Each of the reference substance and test substance were analysed by HPLC at 210 nm. After equilibration of the HPLC system, Urea was injected first, the reference substances were injected in duplicate, followed by the test chemical solution in duplicate. Reference substances were injected again after test sample, no change in retention time of reference substances was observed. Retention time tR were measured, averaged and the decimal logarithms of the capacity factors k were calculated. The graph was plotted between log Koc versus log k(Annex - 2).The linear regression parameter of the relationship log Koc vs log k were also calculated from the data obtained with calibration samples and therewith, log Koc of the test substance was determined from its measured capacity factor. The reference substances were chosen according to structural similarity with the test substance and calibration graph was prepared. The reference substances were Phenol, Toluene, Xylene, Ethylbenzene, Naphthalene, Phenenthrene having Koc value ranging from 1.32 to 4.09. The Log Koc value of test chemical was determined to be 3.313 ± 0.007 at 25°C. This log Koc value indicates that the test chemical has a moderate sorption to soil and sediment and therefore have slow migration potential to ground water.

 

On the basis of above overall results for test chemical, it can be concluded that thetest chemicalhas a low to moderate strong sorption to soil and sediment and therefore have moderate to slow migration potential to ground water.