Ethanol, 2,2'-iminobis-, N-tallow alkyl derivs., N-oxides

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:

2019-03-05 to 2019-03-29

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

23 March 2006

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

A water accommodated fraction (WAF) equilibration trail was performed prior to exposure. The purpose of the equilibration trail is to determine the optimum mixing duration to be used for WAF preparation. One individual WAF was prepared at each of three loading levels 1.0, 50.0 and 100 mg/L. At 2, 4, 6 and 24 hours after initiation of mixing, mixing was stopped and the solutions were allowed to settle for one hour. A sample of WAF was removed from each loading level and mixing was resumed. The concentration of organic carbon that had solubilized into the WAF from the test substance was measured using the TOC analyzer (TOC Analyzer/TNML; Make: Shimadzu; Model: LCPH E200 ROHS).

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Strain: ATCC 22662
- Source: American Type Culture Collection
- Method of cultivation: In OECD alga medium
- Age of inoculum (at test initiation): In order to obtain exponentially growing algal cells, the pre-culture (in two flasks containing 100 mL of culture medium) was initiated 3 days prior to dose ranging study and limit test, respectively to use in the test culture as inoculum. The algal cell density of pre-cultures (No. 1 and 2) during range finding study and limit test was 41025 and 40575 cells/mL at start.
- Method of cultivation: Stock culture was periodically subcultured (at least once in a week) and maintained with the illumination of 4440 to 8880 lux at a temperature of 21 to 24 °C. From healthy axenic culture of alga, pre-culture was prepared of desired cell density to provide the inoculum for test cultures.

ACCLIMATION
- Acclimation period: 2 - 4 days
- Culturing media and conditions: Same as test
- Any deformed or abnormal cells observed: no

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test temperature:

22.1 to 22.8 (during limit test)

pH:

8.01 to 8.14 at the beginning (0 h) and 8.02 to 8.12 at the end of the test (72 h; during limit test)

Nominal and measured concentrations:

Nominal loading rates: 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L
No measured concentrations

Details on test conditions:

TEST SYSTEM
- Test vessel: Conical flasks (Erlenmeyer flasks)
- Type (delete if not applicable): Closed with air-permeable stoppers
- Material, size, fill volume: Glass; nominal volume: 250 mL; fill volume: 100 mL
- Aeration: Culture flasks were maintained in suspension by using an orbital shaker at 100 ± 2 revolutions per minute during the test period
- Initial cells density: 5462 cells/mL
- No. of vessels per concentration: 3
- No. of vessels per control: 6

GROWTH MEDIUM
- Standard medium used: Yes (OECD Medium)

OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: No
- Photoperiod: 24 hours light
- Light intensity and quality: 4440 to 8880 lux

EFFECT PARAMETERS MEASURED:
- Determination of cell concentrations: By a haemocytometer and by manual counting using microscope

RANGE FINDING STUDY
- Test concentrations: 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L along with negative control
- Results used to determine the conditions for the definitive study: Yes. In the range finding study the growth rate inhibition of 0.36, 0.08, 0.48, 0.02, -0.23 and -0.45%, was observed at the tested nominal loading rates of 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L of the test item, respectively, as compared with the control. Percent inhibition in yield of 1.8, 0.4, 2.4, 0.1, 2.8 and -2.2% was observed at the tested nominal loading rates of 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L of the test item, respectively, as compared with the control.
Based on the results of dose range finding study, a limit test was conducted at the nominal loading rate of 100 mg/L along with control having 6 replicates.

Reference substance (positive control):

yes

Remarks:

3, 5 - Dichlorophenol

Key result

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: growth rate and yield

Key result

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: growth rate and yield

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- ErL50: 1.11 mg/L
- EyL50: 0.29 mg/L

Table 1: Percent Inhibition in Growth Rate

Dose Range Finding Study		Limit test
Nominal Loading Rate (mg/L)	% Inhibition in Growth Rate	Nominal Loading Rate (mg/L)	% Inhibition in Growth Rate
0.0	-	-	-
0.01	0.36	100	0.01
0.1	0.08
1.0	0.48
10.0	0.02
50.0	-0.23
100.0	-0.45

 

Table 2: Percent inhibition in Yield

Dose Range Finding Study		Limit test
Nominal Loading Rate (mg/L)	% Inhibition in Yield	Nominal Loading Rate (mg/L)	% Inhibition in Yield
0.0	-	-	-
0.01	1.8	100	0.1
0.1	0.4
1.0	2.4
10.0	0.1
50.0	2.8
100.0	-2.2

 

Table 3: TOC analysis during exposure (Dose Range Finding Study)

Total Organic Carbon During Exposure (mg)
Group	Nominal Loading rate (mg/L)	Day 0 (Fresh)	Day 3 (Aged)
G1	0.0	-	-
G2	0.01	0.0234	0.0221
G3	0.1	0.2145	0.2113
G4	1.0	0.7117	0.7081
G5	10.0	0.0300	2.728
G6	50.0	18.470	17.640
G7	100.0	45.810	45.430

-: Not applicable.

 

 

Table 4: TOC analysis during exposure (Limit Test)

Total Organic Carbon During Exposure (mg)
Group	Nominal Loading rate (mg/L)	Day 0 (Fresh)	Day 3 (Aged)
G1	0.0	-	-
G2	100.0	57.9500	56.5200

-: Not applicable.

 

Validity criteria of the test:

- The biomass in the control cultures increased exponentially by a factor of 155.27 and 159.28 within 72 h test period during range finding test and limit test, respectively. This corresponds to an average specific growth rate of 1.68 and 1.69 day-1 in the range finding and limit test, respectively (validity criterion: biomass should increase at least by factor of 16 and specific growth rate of 0.92 day-1).
- The mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2, and 2-3, for 72-hours test) in the control cultures was 20.38 and 30.07 during the range finding and limit test respectively (validity criterion: must not exceed 35%).

Validity criteria fulfilled:

yes

Conclusions:

The 72 hours ErL50 (growth rate) and EyL50 (yield) of Humectol LYS MOD on Pseudokirchneriella subcapitata is >100 mg/L. The 72 hours No Observed Effect Loading Rate (NOELR) and the Lowest Observed Effect Loading Rate (LOELR) is >100 mg/L of Humectol LYS MOD.

Executive summary:

The test item Humectol LYS MOD was assessed for Growth Inhibition on Pseudokirchneriella subcapitata according to OECD Guideline 201.

A dose range finding study was conducted with six nominal loading rates of 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L along with control. Each loading rate contains 2 replicates except control possessing 3 replicates. The algal growth (cell density) was assessed at 24, 48 and 72 hours of post-exposure during the test. During the dose range finding study, exponentially growing cells (5709 cells/mL) were exposed to range of selected nominal loading rates. A growth rate inhibition of 0.36, 0.08, 0.48, 0.02, -0.23 and -0.45%, was observed at the tested nominal loading rates of 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L of Humectol LYS MOD respectively as compared with the control. Percent inhibition in yield of 1.8, 0.4, 2.4, 0.1, 2.8 and -2.2% was observed at the tested nominal loading rates of 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L of Humectol LYS MOD respectively as compared with the control.
Based on the results of dose range finding study, a limit test was conducted at the nominal loading rate of 100 mg/L along with control having 6 replicates. The algal growth (cell density) was assessed at 24, 48 and 72 hours post-exposure. Exponentially growing alga cells (5462 cells/mL) were exposed to the limit nominal loading rate. A growth rate inhibition of 0.01 % was observed at the nominal loading rate of 100 mg/L as compared with the control. Similarly, inhibition in yield of 0.1% was observed at the nominal loading rate of 100 mg/L as compared with the control.
From the experiment conducted under standard experimental conditions, the 72 hours ErL50 (growth rate) and EyL50 (yield) of Humectol LYS MOD on Pseudokirchneriella subcapitata is >100 mg/L. The 72 hours No Observed Effect Loading Rate (NOELR) and the Lowest Observed Effect Loading Rate (LOELR) is >100 mg/L of Humectol LYS MOD.

Description of key information

In a 72 hours Growth Inhibition Test on Pseudokirchneriella subcapitata according to OECD guideline 201 an ErL50 (growth rate) and EyL50 (yield) of Humectol LYS MOD of >100 mg/L was determined, respectively. The 72 hours No Observed Effect Loading Rate (NOELR) and the Lowest Observed Effect Loading Rate (LOELR) is >100 mg/L of Humectol LYS MOD, respectively.

Additional information

The test item Humectol LYS MOD was assessed for Growth Inhibition on Pseudokirchneriella subcapitata according to OECD Guideline 201.

A dose range finding study was conducted with six nominal loading rates of 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L along with control. Each loading rate contains 2 replicates except control possessing 3 replicates. The algal growth (cell density) was assessed at 24, 48 and 72 hours of post-exposure during the test. During the dose range finding study, exponentially growing cells (5709 cells/mL) were exposed to range of selected nominal loading rates. A growth rate inhibition of 0.36, 0.08, 0.48, 0.02, -0.23 and -0.45%, was observed at the tested nominal loading rates of 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L of Humectol LYS MOD respectively as compared with the control. Percent inhibition in yield of 1.8, 0.4, 2.4, 0.1, 2.8 and -2.2% was observed at the tested nominal loading rates of 0.01, 0.1, 1.0, 10.0, 50.0 and 100.0 mg/L of Humectol LYS MOD respectively as compared with the control.
Based on the results of dose range finding study, a limit test was conducted at the nominal loading rate of 100 mg/L along with control having 6 replicates. The algal growth (cell density) was assessed at 24, 48 and 72 hours post-exposure. Exponentially growing alga cells (5462 cells/mL) were exposed to the limit nominal loading rate. A growth rate inhibition of 0.01 % was observed at the nominal loading rate of 100 mg/L as compared with the control. Similarly, inhibition in yield of 0.1% was observed at the nominal loading rate of 100 mg/L as compared with the control.
From the experiment conducted under standard experimental conditions, the 72 hours ErL50 (growth rate) and EyL50 (yield) of Humectol LYS MOD on Pseudokirchneriella subcapitata is >100 mg/L. The 72 hours No Observed Effect Loading Rate (NOELR) and the Lowest Observed Effect Loading Rate (LOELR) is >100 mg/L of Humectol LYS MOD.