Dodecamethylenediamine

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

07. Nov. 2019 - 05. Feb. 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Source and lot/batch number of test material: B1-5-10
- Expiration date of the lot/batch: 31. Jan. 2020
- Purity test date: not stated

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: The test item was stored in the test facility in a closed vessel at room temperature (16.2 – 23.7 °C).
- Stability under storage conditions: assumed stable
- Stability under test conditions: assumed stable
- Solubility and stability of the test substance in the solvent/dispersant/vehicle/test medium: assumed stable, solubility determined during analytical validation report
- Reactivity of the test substance with the solvent/vehicle /test medium (if applicable): assumed stable

TREATMENT OF TEST MATERIAL PRIOR TO TESTING
A saturated solution was prepared. This was done by mixing the real load of 10.4 mg/L with the corresponding amount of algal medium (demineralised water enriched with min-erals but without algae and shaking vigorously for 24hours. The resulting solution was filtrated through 0.45 µm PTFE filters. The lower treatments (0.032 / 0.1 / 0.32 / 0.1 / 3.2 mg/L nominal) were prepared by dilution of this stock solution with algal medium.

Analytical monitoring:

yes

Details on sampling:

At the beginning and at the end of the test, the content of the test item in the test solutions was determined using LC/MS-MS-determination.

The following dilution factors were used to dilute the concentrations 3.2 mg/L and 10 mg/L with algal medium to stay in the calibrated range:

Concentration dilution factor
3.2 mg/l 3.2
10 mg/L 10

Vehicle:

no

Details on test solutions:

A saturated solution was prepared. This was done by mixing the real load of 10.4 mg/L with the corresponding amount of algal medium (demineralised water enriched with min-erals but without algae and shaking vigorously for 24hours. The resulting solution was filtrated through 0.45 µm PTFE filters. The lower treatments (0.032 / 0.1 / 0.32 / 0.1 / 3.2 mg/L nominal) were prepared by dilution of this stock solution with algal medium.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

6.3.1 Specification
Unicellular freshwater green alga.
Genus Desmodesmus
Species subspicatus
SAG Strain Number 86.81
Taxonomic position Chlorophyta - Chlorophyceae
Origin and Culture
The culture of Desmodesmus subspicatus was obtained in January 2016 by MBM Sci-encebridge 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 cul-ture was prepared. From an aliquot of the permanent culture, the pre-culture was pre-pared.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

no

Hardness:

not determined

Test temperature:

22.8 – 24.7 °C

pH:

7.5 - 8.0

Dissolved oxygen:

not determined

Salinity:

not applicalbe

Conductivity:

not determined

Nominal and measured concentrations:

At the beginning and at the end of the test, the content of the test item in the test solutions was determined using LC/MS-MS-determination. The measured concentrations of the test concentrations 0.1 / 0.32 / 1 / 3.2 / 10 mg/L lay between 91 % and 104 % of the nominal concentrations at the beginning of the test and between 98 % and 112 % of the nominal concentrations at the end of the test. The measured concentration of the test concentra-tion 0.032 mg/L lay lower than the LOQ (0.025 mg/L) at the beginning and the end of the test. In this treatment the test item was detectable, but not quantifiable.
Because all measured concentrations within the calibrated range were within the range of ± 20 % of the nominal concentration, therefore the determination of the biological re-sults was based on the nominal concentrations

Details on test conditions:

Date: 10. – 13. Dec. 2019
Treatments tested: 0.032 / 0.1 / 0.32 / 1 / 3.2 / 100 mg/L nominal concentration
The concentrations to be tested are based on non GLP pre-tests
Number of replicates: 6 replicates for the blank control
3 replicates for each treatment
Vessels: glass flasks total volume 65 mL
Duration: 72 hours
Lighting: within the specified range (4440 – 8880 Lux)
Temperature: 23.0 – 25.5 °C (see chapter 13 Deviations)

Replicates
Treatments: 3 vessels, filled with 45 ±1 mL test solution and algae
1 vessel, filled with 45 ±1 mL test solution without algae for analytical determination
Blank Control: 6 vessels, each filled with 45 ±1 mL algal medium and algae
1 vessel filled with 45 ±1 mL algal medium without algae for analytical determination

Reference substance (positive control):

yes

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

<= 0.03 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.65 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

< 0.03 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks:

as yield: Biomass at the end of the exposure period minus the biomass at the start of the exposure period.

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

< 0.03 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks:

as yield: Biomass at the end of the exposure period minus the biomass at the start of the exposure period.

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.03 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks:

as yield: Biomass at the end of the exposure period minus the biomass at the start of the exposure period.

Results with reference substance (positive control):

The results of the last study with the positive control K2Cr2O7 are presented in the follow-ing table. The study was performed under GLP conditions in September 2019.
Biological Results of K2Cr2O7
Parameter Value 95% confidence interval
72h ErC50 0.80 mg/L 0.76 – 0.85 mg/L
72h EyC50 0.39 mg/L 0.38 – 0.40 mg/L

Reported statistics and error estimates:

Calculation of results was performed with the help of validated software. Calculation was performed with Microsoft Excel® and ToxRat® Professional, version 3.3.0.
Note: All calculations are performed with unrounded values. Therefore, re-calculation with rounded values may lead to slightly different results.

Validity criteria fulfilled:

yes

Conclusions:

The following results for the test item 1,12-Diaminododecane were determined:
EC50 (growth rate, 72 h) = 0.65 mg/L

Executive summary:

Two experiments were performed. In the 1^stexperiment one of the validity criteria was not met. Therefore, the experiment was repeated. The results of the 1^stexperiment are not stated in the report, but will be kept together with the other raw data in the GLP archive of the test facility.

The study was performed using 6 concentrations ranging from 0.032 to 10 mg/L (nominal). Incubation time (test systemDesmodesmus 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 in all test concentrations.

At the beginning and at the end of the test, the content of the test item in the test solutions was determined using LC/MS-MS-determination. The measured concentrations of the test concentrations 0.1 / 0.32 / 1 / 3.2 / 10 mg/L lay between 91 % and 104 % of the nominal concentrations at the beginning of the test and between 98 % and 112 % of the nominal concentrations at the end of the test. The measured concentration of the test concentration 0.032 mg/L lay lower than the LOQ (0.025 mg/L) at the beginning and the end of the test. In this treatment, the test item was detectable, but not quantifiable.

Because all measured concentrations within the calibrated range were within the range of ± 20 % of the nominal concentration, thereforethe determination of the biological results was based on the nominal concentrations (see Guidance Doc. No.23 (§176)).

The 72h-EC₅₀values 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 0.65 - 1.10 mg/L, yield 0.21 – 0.66 mg/L).

 

The following results for the test item1,12-Diaminododecanewere determined:

Table 3           Results of the test item

Endpoint	NOEC	LOEC	EC10	EC50
Growth Rate	< 0.03 mg/L	≤ 0.03 mg/L	0.10 mg/L	0.65 mg/L
Yield	< 0.03 mg/L	≤ 0.03 mg/L	< 0.03 mg/L	0.03 mg/L

 

Because of the very high toxicity of the test item, the EC₁₀(yield) is given in a range.

[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.

Description of key information

The following results for the test item 1,12-Diaminododecane were determined:

EC50 (growth rate, 72 h) = 0.65 mg/L

Key value for chemical safety assessment

EC50 for freshwater algae:

0.65 mg/L

EC10 or NOEC for freshwater algae:

0.1 mg/L