Amines, N-C12–14 (even numbered)-alkyltrimethylenedi-, reaction products with chloroacetic acid and diethylenetriamine, N-C12–14 (even numbered)-alkyl derivs.

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

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:

2011

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

2008

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

All concentration levels and the control were analytically verified via LC-MS/MS at the start (0 hours) and at the end of the exposure (72 hours) with algae.

Vehicle:

no

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: green alga
- Strain: HINDÁK, SAG 61.81
- Source (laboratory, culture collection): Sammlung von Algenkulturen (SAG), Pflanzenphysiologisches Institut der Universität Göttingen, Nikolausberger Weg 18, D-37073 Göttingen
- Age of inoculum (at test initiation): three to four day old preculture
- Method of cultivation: Fresh stocks are prepared every month on Z-Agar. Light intensity amounts 5125 to 6411 (mean: 5975) lux for 24 hours per day.

ACCLIMATION
- Culturing media and conditions (same as test or not): Nutrient medium Z according to LÜTTGE et al. (1994).
- Any deformed or abnormal cells observed: no

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

22.5 (mean)

pH:

7.82 - 8.12

Nominal and measured concentrations:

0.00266, 0.00841, 0.0266, 0.0841, 0.266 mg/L (nominal dry content concentration )

Details on test conditions:

TEST SYSTEM
- Test vessel: Sterile Erlenmeyer flasks (vol. 250 mL) with cotton wool plugs, fill volume: 100 mL
- Aeration: Test containers were placed on a rotary shaker and oscillated at approximately 70 rpm
- Initial cells density: 6839 cells/mL
- Control end cells density: 1626745 (mean)
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Detailed composition if non-standard medium was used: natural river water, see Any other information on materials and methods

TEST MEDIUM / WATER PARAMETERS
- Supplemental Dilution Water: OECD TG 201 medium
natural river water, see Any other information on materials and methods

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 24 hours/day light
- Light intensity and quality: Approximately 4440 to 8880 lux, corresponding to 60 to 120 µE*m-2*s-1

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Chlorophyll measurement: yes
- Other: Microscopic evaluation (unusual cell shapes, colour differences, differences in chloroplast morphology, flocculation, adherence of algae to test containers and agglutination of algae cells)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: sqrt(10)
Range finding study
- Test concentrations: 0.0100 - 0.100 - 1.00 mg/L
- Results used to determine the conditions for the definitive study:
Nominal Test Item Concentration
[mg/L] Growth Rate Inhibition [%] Yield Inhibition [%]
1.00 100 100
0.100 6 29
0.0100 0 2

Reference substance (positive control):

yes

Remarks:

Potassium dichromat

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.008 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

0.027 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Remarks on result:

other: 95% c.i.

Remarks:

0.0264 – 0.0288 mg/L

Duration:

72 h

Dose descriptor:

EC20

Effect conc.:

0.031 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Remarks on result:

other: 95% c.i.

Remarks:

0.0296 – 0.0375 mg/L

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.039 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth rate

Remarks on result:

other: 95% c.i.

Remarks:

0.0346 – 0.0472

Validity Criteria

 

Validity Criteria	Required	This study
Increase of the cell growth in the control cultures	Exponentially, > 16-fold corresponding to a specific growth rate of 0.92 day^-1	238-fold (specific growth rate 1.82 day^-1)
Mean coefficients of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control cultures	≤35%	28.1%
Coefficient of variation of average specific growth rates during the whole test period in replicate control cultures	≤7%	0.59%

 

 

Nominal dry content concentration of the test item	Replicate	Growth rate		Inhibition of growth rate	Yield		Inhibition of yield
[mg/L]	No.	[d^-1]		[%]	[cells/mL]		[%]
0.266	1		n.a.	100		n.a.	100
	2		n.a.	100		n.a.	100
	3		n.a.	100		n.a.	100
	Mean	(+)	n.a.	100	(+)	n.a.	100
0.0841	1		-0.143	100		-2382	100
	2		-0.044	100		-846	100
	3		0.036	98		784	100
	Mean	(+)	-0.050	99	(+)	-815	100
0.0266	1		1.66	9		983949	39
	2		1.68	8		1043230	36
	3		1.66	9		974352	40
	Mean	(+)	1.66	9	(+)	1000510	38
0.00841	1		1.77	3		1382588	15
	2		1.77	3		1382939	15
	3		1.82	0		1577693	3
	Mean	(-)	1.79	2	(-)	1447740	11
0.00266	1		1.83	0		1634224	-1
	2		1.84	-1		1709714	-6
	3		1.83	0		1658509	-2
	Mean	(-)	1.83	-1	(-)	1667482	-3
Control	1		1.81			1529122
	2		1.84			1673666
	3		1.83			1625095
	4		1.83			1642416
	5		1.83			1654238
	6		1.82			1594898
	Mean		1.82			1619906

n.a. = data not determinable

Validity criteria fulfilled:

yes

Conclusions:

The NOEC, EC10 and EC50 values based on growth rate were 8.41 µg/L, 27.3 and 38.7 µg/L, respectively.

Executive summary:

In a 72 hour acute toxicity study, the cultures of Pseudokirchneriella subcapitata were exposed to Reaction product of lauryl-PDA/lauryl-DETA with chloroacetic acid (27% a.i.) at nominal concentrations of 0 (control), 0.00266, 0.00841, 0.0266, 0.0841 and 0.266 mg/L (nominal dry content concentration) under static conditions in accordance with OECD Guideline 201 (2011) and EU method C3 (2008). The study was conducted with an initial cell density of 6839 cells/mL with natural river water.

 

The exposure concentrations of the test item were analytically verified via LC‑MS/MS at the start of the exposure (0 hours) and at the end of the exposure (72 hours) in all concentration levels and the control. Details of the analytical method are presented in section13. The measured concentrations of the test item in fresh media at the start of the exposure (0 hours) were in the range of < LOQ to 83% of the nominal values and 72 to 190% at the end of the exposure (72 hours).

Since the test is a UVCB substance all effect values given are based on the nominal dry content concentrations of the test item.

 

The NOEC, EC10 and EC50 values based on growth rate were 8.41 µg/L, 27.3 and 38.7 µg/L, respectively. The NOEC, EC10 and EC50 values based on yield were 8.41 µg/L, 14.2 and 30.7 µg/L, respectively. Microscopic examination of the cells at the start and end of the incubation period revealed no abnormal cells.

 

Results Synopsis

Test Organism: Pseudokirchneriella subcapitata

Test Type: Static

 

72 hr EC50:  38.7 µg a.i./L (95% c.i. 34.6 – 47.2 µg a.i./L)

72 hr EC10: 27.3 µg a.i./L (95% c.i. 26.4 – 28.8 µg a.i./L)

72 hr NOEC:  8.41 µg a.i./L

Endpoint(s) Effected: growth rate

Description of key information

72 hr EC50: 38.7 µg a.i./L (95% c.i. 34.6 – 47.2 µg a.i./L)

72 hr EC10: 27.3 µg a.i./L (95% c.i. 26.4 – 28.8 µg a.i./L)

72 hr NOEC: 8.41 µg a.i./L

OECD TG 201 / EU method C.3; Pseudokirchneriella subcapitata

Key value for chemical safety assessment

EC50 for freshwater algae:

38.7 µg/L

EC10 or NOEC for freshwater algae:

27.3 µg/L

Additional information

In a 72 hour acute toxicity study, the cultures of Pseudokirchneriella subcapitata were exposed to Reaction product of lauryl-PDA/lauryl-DETA with chloroacetic acid (27% a.i.) at nominal concentrations of 0 (control), 0.00266, 0.00841, 0.0266, 0.0841 and 0.266 mg/L (nominal dry content concentration) under static conditions in accordance with OECD Guideline 201 (2011) and EU method C.3 (2008). The study was conducted with an initial cell density of 6839 cells/mL with natural river water.

 

The exposure concentrations of the test item were analytically verified via LC‑MS/MS at the start of the exposure (0 hours) and at the end of the exposure (72 hours) in all concentration levels and the control. The measured concentrations of the test item in fresh media at the start of the exposure (0 hours) were in the range of < LOQ to 83% of the nominal values and 72 to 190% at the end of the exposure (72 hours). Since the test is a UVCB substance all effect values given are based on the nominal dry content concentrations of the test item.

 

The NOEC, EC10 and EC50 values based on growth rate were 8.41 µg/L, 27.3 and 38.7 µg/L, respectively. The NOEC, EC10 and EC50 values based on yield were 8.41 µg/L, 14.2 and 30.7 µg/L, respectively. Microscopic examination of the cells at the start and end of the incubation period revealed no abnormal cells.

The 72 h NOErC of the source substance DOPA-Glycinate in Pseudokirchneriella subcapitata was 9.55 µg a.i./L; the 72 h ErC50 was 23.7 µg a.i./L (23.2–24.5 µg/L).