Reaction mass of N,N-ethanediylbis- Hexadecanamide, N-[2-[(1-oxohexadecyl)amino]ethyl]-Octadecanamide, Hexadecanoic acid, 12-hydroxystearic acid, reaction products with ethylenediamine, N,N-ethanediylbis-Octadecanamide and Octadecanoic acid, 12-hydroxystearic acid, reaction products with ethylenediamine

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 Nov - 18 Nov 2005

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

GLP compliance:

yes

Remarks:

according to the Japanese GLP Standard

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
PHYSICO-CHEMICAL PROPERTIES
- Melting point: 146.6 °C
- Water solubility: < 4.9 μg/L

OTHER PROPERTIES
- Other: solubility: < 200 μg/L in hexane, 57 mg/L in chloroform, < 200 μg/L in ethyl acetate, < 200 μg/L in methanol, < 4.9 mg/L in octanol

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 0, 100 mg/L
- Sampling method: test solution for analysis was separately prepared for analysis at the initiation of the test. After exposure, same volume of test solution was sampled directly from each test vessel and cetrifuged (3000 rpm, 10 min) to get rid of alga.
- Sample storage conditions before analysis: analysis was conducted immediately after sampling, no storage

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: test item is immiscible with water and therefore a WAF was prepared by adding the test item in dilution water. After 30 minutes sonication, test water was stirred (750 rpm) for 24 hours at 23± 2 °C. Test water was filtered with No. 2 filter paper.
- Controls: dilution water
- Evidence of undissolved material (e.g. precipitate, surface film, etc): test water was colorless after preparation

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Common name: alga
- Strain: ATCC22662
- Source: American Type Culture Collection
- Method of cultivation: on rotary shaker

ACCLIMATION
- Acclimation period: 5 days (11/11/2005 - 15/11/2005)
- Culturing media and conditions (same as test or not): same as test

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test temperature:

22.0 - 22.7 (23 ± 2 °C)

pH:

7.7 - 9.9

Nominal and measured concentrations:

nominal concentration: 100 mg/L
measured concentration : 0.053 (at the initiation), 0.00619 (after exposure) mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 300 mL Erlenmeyer flask with an air-permeable stopper
- Type: open
- Material, fill volume: glass, 100 mL
- Aeration: no
- Initial cells density: 1000 cells/mL
- Control end cells density: 2883100 ± 120600
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: not stated

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: not adjusted
- Photoperiod: continuously
- Light intensity and quality: 4000 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: counting chamber; electronic particle counter;

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: control, 10.0, 50.0, 100 mg/L
- Results used to determine the conditions for the definitive study: growth inhibition rate: 10.0 mg/l: -8%, 50.0 mg/L: -11%, 100 mg/L: -40%

Reference substance (positive control):

yes

Remarks:

potassium dichromate (reagent grade)

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 39.8 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

refers to N N'-ethylene-bis-octadecanamide

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 39.8 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

refers to N N'-ethylene-bis-octadecanamide

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 39.8 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

refers to N N'-ethylene-bis-octadecanamide

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 39.8 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

refers to N N'-ethylene-bis-octadecanamide

Basis for effect:

biomass

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): not observed
- Unusual cell shape: not observed
- Flocculation: not observed
- Adherence to test vessels: not observed
- Aggregation of algal cells: not observed

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EbC50: 0.430 ± 0.0698 mg/L (n=17)
- Other: this test is performed every 6 months.

Reported statistics and error estimates:

The EC50 values and associated 95% confidence limits could not be determined statistically because the test was conducted at one concentration level. The NOEC were determined by Student's t-testm subsequent to F test for homogeneity of variances. Statistical analyses were performed using Yukms Statlight #3 software (Yukms Corp., Tokyo) and all tests of significant were at α=0.05.

Determined effect concentration based on the measured concentration of the test item:

EbC50: > 0.053 mg/L

NOECb: > 0.053 mg/L

ErC50: > 0.053 mg/L

NOECr: > 0.053 mg/L

Determined effect concentration based on the measured concentration of active ingredient (N N'-ethylene-bis-octadecanamide, CAS 110 -30 -5)

EbC50: > 0.0211 mg/L

NOECb: > 0.0211 mg/L

ErC50: > 0.0211 mg/L

NOECr: > 0.0211 mg/L

Table 1. Measured Concentration of the Test Substance in Test Cultures

Loading Rate (*1) (mg/L)	Measured Concentration (mg/L) (Percent of 0hr)
	0 Hour	72 Hour
Control	<0.002	<0.00002
100	0.053	0.00619 (12)

(*1): The ratio of test substance to water (in mg/L) used in the preparation of the WAF.

Table 2. Cell Densities of Pseudokirchneriella subcapitata during the 72 -Hour Exposure

Loading Rate [Measured Conc. at 0 hr] (mg/L)	Cell Densities (cells/mL)
		0 Hour	24 Hour	48 Hour	72 Hour
Control	Average	10000	82800	756900	2883100
	SD	0	11900	123400	120600
100 [0.053]	Average	1000	83000	778900	3413100
	SD	0	16300	138300	70200

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

refer to chapter 13

Reason / purpose for cross-reference:

read-across source

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

WAF

Basis for effect:

growth rate

Validity criteria fulfilled:

yes

Conclusions:

Based on the result of a toxicity to algae test with a read across substance it can be concluded that the registered substance exhibited no acute toxicity to the freshwater algae Pseudokirchenriella subcapitata within the range of water solubility.

Executive summary:

The toxicity of the read Across Source substance Amides, C16-C18 (even), N,N'-ethylenebis to the freshwater algae Pseudokircheneriella subcapitata was investigated in one study from the Japanese Ministry of Environment (MOE, 2002). This study followed the OECD Guideline No. 201: Algae, Growth Inhibition Test (1984). The results from this test show that after 72 hours exposure to Amides, C16-C18 (even), N,N'-ethylenebis, no effects on growth rate of Pseudokircheneriella subcapitata were observed, up to the highest attainable concentration (0.053 mg/L, limit of solubility). Therefore, the 72h-EC50 was greater than 0.053 mg/L and no toxicity within the range of water solubility was reported.

Description of key information

Based on the result of a toxicity to algae test with a read across substance it can be concluded that the registered substance exhibited no acute toxicity to the freshwater algae Pseudokirchenriella subcapitata within the range of water solubility.

The toxicity of the read Across Source substance Amides, C16-C18 (even), N,N'-ethylenebis to the freshwater algae Pseudokircheneriella subcapitata was investigated in one study from the Japanese Ministry of Environment (MOE, 2002). This study followed the OECD Guideline No. 201: Algae, Growth Inhibition Test (1984). The results from this test show that after 72 hours exposure to Amides, C16-C18 (even), N,N'-ethylenebis, no effects on growth rate of Pseudokircheneriella subcapitata were observed, up to the highest attainable concentration (0.053 mg/L, limit of solubility). Therefore, the 72h-EC50 was greater than 0.053 mg/L and no toxicity within the range of water solubility was reported.