Trisodium dihydrogen -N,N-[bis[2-[bis(carboxylatomethyl)amino]ethyl]]glycinate

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

activated sludge respiration inhibition testing

Type of information:

migrated information: read-across based on grouping of substances (category approach)

Adequacy of study:

key study

Study period:

2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP, guideline study

Justification for type of information:

The target substance (DTPA 3Na) is the trisodium salt of the source substance Diethylenetriaminepentaacetic acid (DTPA). The purity of the source substance used for the ready biodegradation test was 100%. Since the target substance is > 99.9% pure and contains no detectable impurities, the extrapolation of toxicity to microorgaisms from the source to the target substance is considered valid. The source material test was conducted according to OECD test guideline 209 and is considered reliable without restriction (Category1).

Reason / purpose for cross-reference:

reference to same study

Reason / purpose for cross-reference:

reference to other study

Qualifier:

according to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Not applicable

Analytical monitoring:

yes

Details on sampling:

The contents of the first control vessel were transferred to a BOD bottle and stirred using a magnetic stirrer. Dissolved oxygen concentrations were measured every 10 seconds over a 10 minute period or until the D.O. dropped below 1.0 mg/L. The respiration rate in subsequent vessels was determined in an identical manner at 15 minute intervals. The contact time for each concentration of reference or test substance with the activated sludge was three hours.

Vehicle:

no

Details on test solutions:

The test substance was dosed by direct weight addition. This method was deemed appropriate based on the limited solubility of the test substance in water.

Test organisms (species):

activated sludge

Details on inoculum:

Activated sludge collected from the Denton Wastewater Treatment Facility, Denton, Maryland on 22 June 2010 was utilized as the inoculum for the test. The Denton facility receives wastes from predominately domestic sources. The sludge was sieved using a 2 mm screen and allowed to settle for
approximately 30 minutes. The supernatant above the settled solids was removed and the total suspended solids (TSS) concentration of the settled sludge was determined. Total suspended solids in the settled sludge were adjusted to a nominal concentration of approximately 4000 mg/L by dilution with municipal water. Fifty mL of synthetic sewage (Appendix I of Protocol) was added to each liter of adjusted sludge. The sludge was maintained at a temperature of 20 ± 2°C and continuously aerated overnight. The pH and total suspended solids concentration of the activated sludge were determined on the day of use in the study.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

3 h

Post exposure observation period:

Not applicable

Hardness:

No data

Test temperature:

19.6 – 20.8°C

pH:

8.0

Dissolved oxygen:

No data

Salinity:

No data

Nominal and measured concentrations:

Nominal-Control, 10, 30, 100, 300, and 1000 mg/L.

Details on test conditions:

Test System
The biological test system was a consortium of microorganisms common to the activated sludge treatment process. The organisms responsible for the decomposition of organic materials are principally aerobic and facultative aerobic bacteria. The test system was chosen because it is representative of a treatment process that may receive the test substance.

Test Conditions and Apparatus
Control, reference, and treatment test mixtures were incubated at 20 ± 2ºC and aerated for three hours at a rate sufficient to provide aerobic conditions and magnetically stirred to maintain solids in suspension. The mixtures were prepared and aerated in 1000 mL Erlenmeyer flasks and then transferred to fill 300 mL biochemical oxygen demand (BOD) bottles to conduct dissolved oxygen (DO) measurements. Test mixtures were identified by project number, test substance identification, test concentration, and bottle number.

Procedure
Test mixtures were prepared at 15 minute intervals starting with the first control. The control contained 16.0 mL of synthetic sewage, 200 mL of inoculum, and enough municipal water to bring the total volume up to 500 mL. The mixture was promptly aerated and stirred using a magnetic stirrer at a rate sufficient to provide aerobic conditions and keep the solids in suspension. Subsequent mixtures contained 16.0 mL of synthetic sewage, 200 mL of inoculum, the appropriate amount of test substance or reference substance stock solution, and enough municipal water to bring the total volume up to 500 mL. Finally, a second control was prepared. All mixtures were aerated for three hours.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

Temperatures measured throughout the period of sludge maintenance and during the test ranged from 19.6 – 20.8°C. The initial total suspended solids (TSS) concentration of the activated sludge after removal of the supernatant layer measured 10,527 mg/L. The measured TSS concentration of the sludge on the day of testing was 4080 mg/L. The measured pH of the sludge on the day of testing was 8.0.

The respiration rates observed in the two controls were 45.9 and 52.9 mg O2/L/hr, or a difference of approximately 14.3%. The respiration
rate for the first control was determined using the DO concentrations 6.2 and 2.5 mg O2/L at 20 and 310 seconds respectively. The respiration rate of the second control was determined using the DO concentrations 5.1 and 2.6 mg O2/L at 30 and 200 seconds respectively. The DO concentration of 5.1 mg O2/L was used based on the linear portion of the respiration curve. The respiration rate of the 30 mg/L test mixture was determined using the DO concentrations 4.7 and 2.5 mg O2/L at 20 and 140 seconds respectively. The DO concentration of 4.7 mg O2/L was used based on the linear portion of the respiration curve. Confidence limits (95%) for the EC50 and the EC50 value itself, were calculated using binomial probability with nonlinear interpolation. The observed percent inhibitions for DTPA (Diethylenetriamine Pentaacetic Acid) ranged from –33.6 to 27.1%.

Results with reference substance (positive control):

The EC50 value for the reference substance was 16.2 mg/L and was within the 5 to 30 mg/L range considered acceptable for the test.

Reported statistics and error estimates:

No data

None

Validity criteria fulfilled:

yes

Conclusions:

Inhibitory effects upon respiration by the test substance at the concentrations evaluated in this study were observed; however, the test substance did not exhibit a concentration dependent dose response pattern. The EC50 value for the test substance is therefore greater than 1000 mg/L, the highest
concentration tested.

Executive summary:

The effect of DTPA (Diethylenetriamine Pentaacetic Acid) on activated sludge microorganisms maintained in an aerobic environment was assessed by the Activated Sludge Respiration Inhibition Test Method (OECD Guideline 209). The test contained control, reference and treatment groups. The control replicates were used to determine the background respiration rate of the sludge and were not dosed with the test or reference substance. The reference group was dosed with 3,5-dichlorophenol, a known inhibitor of respiration, at concentrations of 3, 15 and 50 mg/L. The treatment group was dosed with DTPA (Diethylenetriamine Pentaacetic Acid) at concentrations of 10, 30, 100, 300, and 1000 mg/L. After an exposure period of three hours, the respiration rates of the test solutions were measured using a YSI Model 50B-115 Dissolved Oxygen Meter. The respiration rates in the two controls were 45.9 and 52.9 mg O2/L/hr. The difference between the two control respiration rates was approximately 14.3%, and was within the 15% difference limit established for the test. The validity of the test was further supported

by the results from the 3,5-dichlorophenol reference group, which resulted in an EC50 value of 16.2 mg/L, with 95 percent confidence limits of 3 mg/L and 50 mg/L. The EC50 for the reference substance was within the 5 to 30 mg/L range considered acceptable for the test. An inhibitory dose response effect was not observed for the treatment group. Therefore, the EC50 value for DTPA (Diethylenetriamine Pentaacetic Acid) is greater than 1000 mg/L, the highest concentration tested. The

following is a summary of the results:

Test Substance Nominal Concentration (mg/L) Respiration Rate (mg O2/L/hr-1) Percent Respiration Inhibition*

Control 1 0 45.9 NA

Control 2 0 52.9 NA

3,5-dichlorophenol 3 38.1 22.9%

3,5-dichlorophenol 15 25.6 48.2%

3,5-dichlorophenol 50 11.6 76.5%

Diethylenetriamine Pentaacetic Acid 10 48.5 1.8%

Diethylenetriamine Pentaacetic Acid 30 66.0 -33.6%

Diethylenetriamine Pentaacetic Acid 100 45.3 8.3%

Diethylenetriamine Pentaacetic Acid 300 36.0 27.1%

Diethylenetriamine Pentaacetic Acid 1000 38.1 22.9%

*Calculations performed using MS Excel 2000 in full precision mode. Manual calculations may differ slightly.

NA - Not Applicable

Reference 2

Endpoint:

activated sludge respiration inhibition testing

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

The target substance (DTPA 3Na) is the trisodiun salt of the source substance Diethylenetriaminepentaacetic acid (DTPA), and is therefore structurally very similar. The two substances have high water solubility and would be fully dissociated during the respiratory inhibition test. The source material is neutral (pH 7.5) in solution which is comparable to the test conditions used for the test with the acid (pH 8). The extrapolation of microbial inhibition potential from the source to the target material is therefore considered valid.

Reason / purpose for cross-reference:

read-across source

Key result

Duration:

3 h

Dose descriptor:

EC50

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Conclusions:

The EC50 respiratory inhibition value for DTPA trisodium salt is predicted to be greater than 1000 mg/L.

Executive summary:

Using a read-across approach from an OECD 209 study conducted on the structural analogue DTPA acid, the respiratory inhibition EC50 value of DTPA trisodium salt is predicted to be greater than 1000 mg/L. The two substances are structurally similar, and virtually identical at the pH (8.0) at which the DTPA acid study was undertaken.

Description of key information

Using a read-across approach from an OECD 209 study conducted on the structural analogue DTPA acid, the respiratory inhibition EC50 value of DTPA trisodium salt is predicted to be greater than 1000 mg/L. The two substances are structurally similar, and essentially identical at the pH (8.0) at which the DTPA acid study was undertaken.

 

Key value for chemical safety assessment

EC50 for microorganisms:

1 000 mg/L