Bis[2-(diethylamino)ethyl] adipate

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2018

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Version / remarks:

Deviations from the Study Plan
The following deviation was documented:
The pre-culture was prepared 5 days before the test start. As all validity criteria were met and exponential growth was observed in the blank control, this was stated as uncritical.
The deviation was assessed and signed by the study director on 17. Apr. 2018.

The following deviation was documented:
The lower treatments were produced by dilution of the highest concentration. This was not clearly described in the study plan. As the test concentrations from the study plan have been reached, this deviation is not to be regarded as critical. The deviation was assessed and signed by the study director on 23. Apr. 2018.

Deviations from the Guideline
The following deviations were documented:
The stock solutions to be autoclaved were autoclaved for 20 minutes instead of the required 15 minutes. This can be considered uncritical, as the goal of sterility is achieved.
The deviation was assessed and signed by the study director on 08. Mar. 2018.

The pre-culture was prepared 5 days before the test start. As all validity criteria were met and exponential growth was observed in the blank control this was stated as uncritical.
The deviations was assessed and signed by the study director on 17. Apr. 2018.

Deviations:

yes

Remarks:

The deviations can be considered uncritical

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Bis[2-(diethylamino)ethyl] adipate

EC Number:

240-610-6

EC Name:

Bis[2-(diethylamino)ethyl] adipate

Cas Number:

16545-00-9

Molecular formula:

C18H36N2O4

IUPAC Name:

1,6-bis[2-(diethylamino)ethyl] hexanedioate

impurity 1

Reference substance name:

2-diethylaminoethanol

EC Number:

202-845-2

EC Name:

2-diethylaminoethanol

Cas Number:

100-37-8

Molecular formula:

C6H15NO

IUPAC Name:

2-(diethylamino)ethanol

Test material form:

liquid

Sampling and analysis

Analytical monitoring:

yes

Test solutions

Vehicle:

no

Details on test solutions:

A saturated solution was prepared for the test. This was done by mixing 100.0 mg/L with the corresponding amount of algal medium (demineralised water enriched with minerals but without algae) and shaking for 2 hours. The resulting solution was filtrated through 0.45 µm PTFE filters. Lower test concentrations were prepared by dilution of this stock solution in algal medium.

Test organisms

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

The culture of Desmodesmus subspicatus was obtained in January 2016 by MBM Sciencebridge GmbH (Institut für Pflanzenphysiologie of Universität Göttingen). The algae are kept as stock culture on solid agar at 2 - 8 °C. From the stock culture, a permanent culture was prepared. From an aliquot of the permanent culture, the pre-culture was prepared.

Study design

Test type:

not specified

Water media type:

freshwater

Total exposure duration:

72 h

Test conditions

Test temperature:

20.1 – 23.6 °C

Nominal and measured concentrations:

1 / 3.2 / 10 / 32/ 100 mg/L nominal concentration
The concentrations to be tested are based on non GLP pre- tests. At the lowest concentration, the measured DOC values were in the same range as in the control. At the beginning, no test item was detectable in this treatment. In the other treatments the measured concentrations lay between 87 % and 104 % of the nominal concentrations at the beginning of the test and between 57 % and 71 % of the nominal concentrations at the end of the test. Because the lowest concentration was necessary for evaluation, the real concentration was calculated on the basis of the geometric mean of the measured concentration in the highest concentration and the respective dilution factor. The respective geometric mean of the individual treatments values was almost identical to the calculated values.

Reference substance (positive control):

yes

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 84.8 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 2

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Any other information on results incl. tables

One valid experiment was performed.

The study was performed using 5 concentrations ranging from 1.0 to 100 mg/L. Incubation time (test system Desmodesmus subspicatus) was 72 hours. The cell concentration of each replicate was determined by measuring the cell numbers every 24 hours with an electronic particle counter. Growth rate µ and the yield[1] were determined from the cell number at the respective observation times.

Significant inhibition of algal growth was observed at all tested concentrations.

A saturated solution was prepared for the test. This was done by mixing 100.0 mg/L with the corresponding amount of algal medium (demineralised water enriched with minerals but without algae) and shaking for 2 hours. The resulting solution was filtrated through 0.45 µm PTFE filters. Lower test concentrations were prepared by dilution of this stock solution in algal medium. At the start and at the end of the test, the content of the test item in the test solutions was estimated by determination of the dissolved organic carbon (DOC) content in the test solutions using a carbon analyser. At the lowest concentration, the measured DOC values were in the same range as in the control. At the beginning no test item was detectable in this treatment. In the other treat-ments the measured concentrations lay between 87 % and 104 % of the nominal concentrations at the beginning of the test and between 57 % and 71 % of the nominal concentrations at the end of the test. Because the lowest concentration was necessary for evaluation, the real concentration was calculated on the basis of the geometric mean of the measured concentration in the highest concentration and the respective dilution factor. The respective geometric mean of the individual treatments values was almost identical to the calculated values. The lower measured concentration at the end of the test was caused by the presence of the test organisms (adsorption ingestion of test item onto the algae cells). That means test item was present in the test system but partly not measurable. Therefore additional the results are also provided for nominal concentrations. The 72h-EC50s of potassium dichromate were determined in a separate reference test. For the estimation of the 72h-EC50s of the positive control, the fits showed sufficient statistical correspondence of the data with the dose-response-equation. The values were within the range of the laboratory. The pH of the blank control should not fluctuate by more than 1.5 units. The change was 0.3 units in the blank control.

[1]Yield (according to OECD Guideline 201) is defined as the biomass at the end of the exposure period minus the biomass at the start of the exposure period. Calculation see under 8.2 Growth and growth inhibition are quantified from measurements of the algal biomass as a function of time. Algal biomass is defined as the dry weight per volume, e.g. mg algae/L test solution. However, dry weight is difficult to measure and therefore surrogate parameters are used. Of these surrogates, cell counts are most often used.

1         

1.1         

The cell concentration in the blank control should increase by a factor of at least 16 within 72h.

1.2         

Mean coefficient of variation of daily growth rates in the blank controls should be 35% at the most.

Coefficient of variation of average growth rate during the whole test period should be 7% at the most.

1.3         

The daily growth rates of the blank controls were calculated. Means, standard deviations and coefficients of variation were determined. Values and assessment can be found in the following tables.

Table11.3‑a        Daily Growth Rates of the blank controls

Growth rates	days 0 – 1	days 1 – 2	days 2 – 3	CV of sectional daily growth rates	days 0 – 3
Replicate 1	1.087	1.731	1.210	25%	1.343
Replicate 2	1.344	1.590	1.140	17%	1.358
Replicate 3	1.248	1.527	1.244	12%	1.340
Replicate 4	1.294	1.484	1.411	7%	1.396
Replicate 5	1.197	1.624	1.402	15%	1.408
Replicate 6	1.282	1.617	1.195	16%	1.364
Mean	1.242	1.595	1.267	15%	1.368
Standard deviation	0.090	0.086	0.113		0.028
CV	7%	5%	9%		2%

CV = Coefficient of Variation

 

Table11.3‑b       Assessment

Parameter	Validity criteria	Observed value	Assessment
Increase factor biomass	factor 16 in 72 h	61	Valid
Mean coefficient of variation of daily growth rates	max. 35%	15%	Valid
Coefficient of variation of average growth rate during the whole test period	max. 7%	2%	Valid

 

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The following results for the test item Bis[2-(diethylamino)ethyl]adipate were determined:
Table 3 a Results of the test item (Measured Conc.)
Endpoint NOEC LOEC EC10 EC50
Growth Rate < 0.80 mg/L ≤ 0.80 mg/L 1.35 mg/L > 84.8 mg/L
Yield < 0.80 mg/L ≤ 0.80 mg/L 0.01 mg/L 5.75 mg/L

Table 3 a Results of the test item (Nominal Conc.)
Endpoint NOEC LOEC EC10 EC50
Growth Rate < 1 mg/L ≤ 1 mg/L 1.65 mg/L > 100 mg/L
Yield < 1 mg/L ≤ 1 mg/L 0.01 mg/L 7.18 mg/L

Executive summary:

One valid experiment was performed.

The study was performed using 5 concentrations ranging from 1.0 to 100 mg/L. Incubation time (test system Desmodesmus subspicatus) was 72 hours. The cell concentration of each replicate was determined by measuring the cell numbers every 24 hours with an electronic particle counter. Growth rate µ and the yield[1]were determined from the cell number at the respective observation times.

Significant inhibition of algal growth was observed at all tested concentrations.

A saturated solution was prepared for the test. This was done by mixing 100.0 mg/L with the corresponding amount of algal medium (demineralised water enriched with minerals but without algae) and shaking for 2 hours. The resulting solution was filtrated through 0.45 µm PTFE filters. Lower test concentrations were prepared by dilution of this stock solution in algal medium.

 

At the start and at the end of the test, the content of the test item in the test solutions was estimated by determination of the dissolved organic carbon (DOC) content in the test solutions using a carbon analyser.

At the lowest concentration, the measured DOC values were in the same range as in the control. At the beginning no test item was detectable in this treatment. In the other treatments the measured concentrations lay between 87 % and 104 % of the nominal concentrations at the beginning of the test and between 57 % and 71 % of the nominal concentrations at the end of the test.

Because the lowest concentration was necessary for evaluation, the real concentration was calculated on the basis of the geometric mean of the measured concentration in the highest concentration and the respective dilution factor.

The respective geometric mean of the individual treatments values was almost identical to the calculated values.

The lower measured concentration at the end of the test was caused by the presence of the test organisms (adsorption ingestion of test item onto the algae cells). That means test item was present in the test system but partly not measurable. Therefore additional the results are also provided for nominal concentrations.

The 72h-EC50s of potassium dichromate (K₂Cr₂O₇, CAS No. 7778-50-9) were determined in a separate reference test. The values lay within the range of the laboratory (growth rate 0.73 - 1.10 mg/L, yield 0.21 – 0.66 mg/L).

 

The following results for the test itemBis[2-(diethylamino)ethyl]adipatewere determined:

Table 3‑a              Results of the test item (Measured Conc.)

Endpoint	NOEC	LOEC	EC10	EC50
Growth Rate	< 0.80 mg/L	≤ 0.80 mg/L	1.35 mg/L	> 84.8 mg/L
Yield	< 0.80 mg/L	≤ 0.80 mg/L	0.01 mg/L	5.75 mg/L

 

Table 3‑a              Results of the test item (Nominal Conc.)

Endpoint	NOEC	LOEC	EC10	EC50
Growth Rate	< 1 mg/L	≤ 1 mg/L	1.65 mg/L	> 100 mg/L
Yield	< 1 mg/L	≤ 1 mg/L	0.01 mg/L	7.18 mg/L