N-[3-(dimethylamino)propyl]stearamide monoacetate

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

effects on growth of green algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015-02-02 to 2015-07-23

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Samples were taken at 0 ad 72 h. 10 mL solution sample was collected.

Details on test solutions:

Definitive test stock solution (100 mg/L) was prepared by adding 0.0054 g of test substance to 50 mL BG11 algal medium, followed by ultrasonication for 30 min. Appropriate volumes of stock solution were added to the glass flasks and 100 mL growth solution was added to each flask, it was then ultrasonicated for 20 min.

Test organisms (species):

Chlorella vulgaris

Details on test organisms:

TEST ORGANISM
- Common name: Chlorella vulgaris
- Strain: Beij.
- Source (laboratory, culture collection): Institute of Hydrobiology, Chinese Academy of science
- Age of inoculum (at test initiation): 3 days
- Method of cultivation: The pre-culture was incubated in a shaker exposed to 3000 Lux (+/- 15 %) continuous illumination at 22.6-23.9 degrees Celsius. Rotor speed - 100 (+/-10) rpm.

ACCLIMATION
- Acclimation period: 3 days
- Culturing media and conditions (same as test or not):same as test
- Any deformed or abnormal cells observed: No

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

22.8 - 23.°C

pH:

7.2 - 8.38

Salinity:

NA

Conductivity:

NS

Nominal and measured concentrations:

The nominal and measured concentrations can be seen in Table 1 in the Any other information on materials and methods incl. tables section. The concentrations varied beyond acceptable criteria in the 0.05, 0.10 and 0.20 mg/L nominal concentration groups, the measured concentrations in these were considerably smaller than the nominal.

Details on test conditions:

TEST SYSTEM
- Test vessel: glass flasks, total 100 mL growth medium in each, 250 mL total volume.
- Aeration: Flasks were capped with air-permeable stoppers
- Type of flow-through (e.g. peristaltic or proportional diluter): NA
- Renewal rate of test solution (frequency/flow rate): NA
- Initial cells density: 2.22 x 10^5 cells/mL
- Control end cells density: mean 726 x 10^4 cells/mL
- No. of organisms per vessel: 2.22 x 10^7
- No. of vessels per concentration (replicates):5
- No. of vessels per control (replicates):5


GROWTH MEDIUM
- Standard medium used: no
- Detailed composition if non-standard medium was used: See Overall remarks, attachments section for full components

TEST MEDIUM / WATER PARAMETERS
BG11 Culture medium was used (components can be seen in the Overall remarks, attachments section). Mixed with pure water ad sterilised by autoclaving at 120 degrees Celsius for 20 min. Water source not stated, referred to as "pure water", not specified if deionised as required by guidelines.

OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: Continuous
- Light intensity and quality: 3000 Lux (controlled within -/+ 15% of average light intensity


EFFECT PARAMETERS MEASURED
Algal biomass in each flask was determined by microscope at 24, 48 and 72 h.

TEST CONCENTRATIONS

- Test concentrations: 0.05, 0.10, 0.20, 0.40, 0.80
- Results used to determine the conditions for the definitive study: Range finding test (1, 10 and 100 mg/L concentrations of test substance) resulted in inhibition rates of 98.1, 100 and 102 % respectively.

CULTURING APPARATUS
-Details on culturing apparatus used: Rotor speed: 100 +/-10 rpm

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

0.026 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

0.19 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

During the test period the pH values of blank control mediums was between 7.20 and 8.38. The biomass in the control cultures increased exponentially with specific growth rates of 1.28/day, 1.27/day and 1.16/day between days 0 and 1, 0 and 2 and 0 and 3, respectively. Coefficients of variation of average specific growth rates for the same time period were 1.78 %, 0.67% and 0.33%, respectively and the mean coefficient of variation for section-by-section specific growth rates in the blank control cultures for the three time periods were 1.78%, 2.60 % and 0.79%, respectively. Therefore, the study met the validity criteria in this regard.
As the measured concentrations varied from the nominal, the results were calculated using geometric mean measured concentrations.

The 72 hour EC50 on C. vulgaris Beij. of the test substance was 0.19 mg/L and its LOEC was >0.051 mg/L when based on geometric mean measured concentration. the NOEC of the test substance was 0.026 mg/L if based on geometric mean measured concentration.

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- EC50: 0.90 mg/L (cell concentrations, 72 h, based on nominal concentrations)

Table 2.

Nominal concentration ^1(mg/L)	0 h	24 h		48 h		72 h
	pH	^2Cell conc.	Inhibition Rate (%)	^2Cell conc.	Inhibition Rate (%)	pH	^2Cell conc.	Inhibition Rate (%)
Blank control	7.18	77.3	-	279	-	8.45	731	-
	7.16	80.2		285		8.49	734
	7.15	81.7		273		8.47	715
Specific growth rate (day ^-1)		1.28 (0 ~ 1 d)		1.27 (0 ~ 2 d)		1.16 (0 ~ 3 d)
Coefficients of variation		1.78 (0 ~ 1 d)		0.67 (0 ~ 2 d)		0.33 (0 ~ 3 d)
Specific growth rate (day ^-1)		1.28 (0 ~ 1 d)		1.25 (1 ~ 2 d)		0.96 (2 ~ 3 d)
Coefficients of variation		1.78 (0 ~ 1 d)		2.60 (1 ~ 2 d)		0.79 (2 ~ 3 d)
0.05	7.20	73.0	10.7	288	0.43	8.34	697	2.64
	7.21	67.2		273		8.32	646
	7.22	68.6		267		8.36	665
0.10	7.27	64.3	18.0	275	2.49	8.38	637	4.77
	7.26	62.8		260		8.34	596
	7.24	62.8		251		8.32	614
0.20	7.36	36.7	63.9	122	32.3	8.25	437	15.8
	7.31	35.3		119		8.27	404
	7.34	33.8		128		8.21	415
0.40	7.40	26.6	81.9	38.2	78.1	8.10	32.4	92.6
	7.38	28.0		39.6		8.17	23.7
	7.41	29.5		38.2		8.06	30.9
0.80	7.40	22.2	102	32.4	84.6	7.98	25.1	93.9
	7.39	20.8		35.3		7.85	29.5
	7.43	22.2		30.9		7.91	28.0
ErC50 (mg/L) (based on geometric mean measured concentration)		0.11		0.19		0.19
95 % confidence interval (mg/L)		0.10 ~ 0.11		0.18 ~ 0.20		0.18 ~ 0.20

¹ The nominal cell concentrations of Chorella vulgaris Beij. At 0 h were all 2.22 x 10⁵ cells/mL.

² (x10⁴ cells/mL)

 

 

Validity criteria fulfilled:

yes

Conclusions:

The 72 hour EC50 on C. vulgaris Beij. of the test substance was 0.19 mg/L and its LOEC was >0.10 mg/L based on nominal, 0.051 mg/L when based on geometric mean measured concentration. The NOEC of the test substance was 0.05 mg/L based on nominal concentrations, 0.026 mg/L if based on geometric mean measured concentration. The results based on geometric mean measurements are more reliable because the measured and nominal concentrations in the test varied by more than 20%.

Executive summary:

A study testing the effects on growth inhibition of the algae Chorella vulgaris Beij. by the test substance was carried out. It was done according to "The guidelines for testing of chemicals" (HJ/T 153-2004), "The Guidelines for the Testing of Chemicals-Effects on Biotic Systems" (the 2nd edition, Beijing:China Environment Press. 2013) and with reference to Procedure 201 ("Freshwater Alga and Cyanobacteria, Growth Inhibition Test" (2011)) of the "Guidelines for Testing of Chemicals" by OECD.

An initial range-finding test was carried out using concentrations of 1, 10 and 100 mg/L of the test substance. This resulted in inhibition rates of 98.1, 100 and 102 %, respectively. Based on these results the following nominal concentrations were selected for the definitive test: 0.05, 0.10, 0.20, 0.40 and 0.80 mg/L. In this test the algae were exposed to the different concentrations of the test chemical and the numbers of algae in each replicate were counted at 24, 48 and 72 hours. The resulting growth rates were then compared to blank control algae exposed to the same conditions bar the test substance to establish the differences in growth and calculate inhibition rates. The concentrations of the test chemical within exposures were measured at 0 and 72 hours and the results showed a large deviation from the nominal concentrations in the 0.05, 0.10 and 0.20 mg/L concentrations (54.3, 34.1 and 43.3 % of the nominal concentrations, respectively), therefore the effect concentrations were calculated using geometric mean measurements using the measured rather than nominal concentrations. 

The 72 hour EC50 on C. vulgaris Beij. of the test substance was 0.19 mg/L and its LOEC was >0.051 mg/L (0.10 when based on nominal concentrations). the NOEC of the test substance was 0.026 mg/L (0.05 mg/L if based on nominal concentrations).

Description of key information

72 h ErC50 = 0.19 mg/L (geomean), 72 h NOEC = 0.026 mg/L (geomean) (Chlorella vulgaris Beij.); OECD 201; Zhen (2015)

Key value for chemical safety assessment

EC50 for freshwater algae:

0.19 mg/L

EC10 or NOEC for freshwater algae:

0.026 mg/L

Additional information

The key study tested the effects on growth inhibition of the algae Chorella vulgaris Beij. by the test substance. It was done according to "The guidelines for testing of chemicals" (HJ/T 153-2004), "The Guidelines for the Testing of Chemicals-Effects on Biotic Systems" (the 2nd edition, Beijing:China Environment Press. 2013) and with reference to Procedure 201 ("Freshwater Alga and Cyanobacteria, Growth Inhibition Test" (2011)) of the "Guidelines for Testing of Chemicals" by OECD.

An initial range-finding test was carried out using concentrations of 1, 10 and 100 mg/L of the test substance. This resulted in inhibition rates of 98.1, 100 and 102 %, respectively. Based on these results the following nominal concentrations were selected for the definitive test: 0.05, 0.10, 0.20, 0.40 and 0.80 mg/L. In this test the algae were exposed to the different concentrations of the test chemical and the numbers of algae in each replicate were counted at 24, 48 and 72 hours. The resulting growth rates were then compared to blank control algae exposed to the same conditions bar the test substance to establish the differences in growth and calculate inhibition rates. The concentrations of the test chemical within exposures were measured at 0 and 72 hours and the results showed a large deviation from the nominal concentrations in the 0.05, 0.10 and 0.20 mg/L concentrations (54.3, 34.1 and 43.3 % of the nominal concentrations, respectively), therefore the effect concentrations were calculated using geometric mean measurements using the measured rather than nominal concentrations. 

The 72 hour EC50 on C. vulgaris Beij. of the test substance was 0.19 mg/L and its LOEC was 0.051 mg/L (0.10 mg/L when based on nominal concentrations). the NOEC of the test substance was 0.026 mg/L (0.05 mg/L if based on nominal concentrations).