Fatty acid chlorides, C18 unsatd., reaction products with sodium N-methyltaurinate

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:

2012-12-10 to 2012-12-13

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

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 1.00 - 3.16 - 10.0 - 31.6 - 100 - 316 mg/L
- Sampling method: The concentrations of the test item were analytically determined from all test concentrations and the control at the start of the exposure (0 h) and at the end of the exposure (72 h). Samples at the start of the exposure were taken from an additional replicate without algae. At the end of the exposure samples were taken directly from the pooled test replicates.
- Sample storage conditions before analysis:
All original samples were stored at room temperature until sample preparation. Prepared samples were stored at room temperature until analysis.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution of 316 mg/L was freshly prepared with dilution water.
- Eluate: DIlution water
- Differential loading: 1.00 - 3.16 - 10.0 - 31.6 - 100 - 316 mg/L
- Controls: Six replicates without test item were tested under the same test conditions as the test replicates.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: Desmodesmus subspicatus Chodat
- Strain: SAG 86.81
- Source (laboratory, culture collection): SAG Pflanzenphysiologisches Institut der Universitaet Goettingen, Nikolausberger Weg 18, D-37073 Goettingen, Germany
- Age of inoculum (at test initiation): 3 days
- Method of cultivation: Fresh stocks were prepared every month on Z-Agar. Light intensity amounted to 35 - 70 µE x m-2 x s-1 for 24 h per day.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

No

Hardness:

0.24 mmol Ca+Mg/l

Test temperature:

min.: 22.0 max.: 23.0, mean value: 22.5 °C

pH:

Nominal test item concentration pH-value
Test item [mg/L] Active ingredient [mg/L a.i.] Start; 0 hours End; 72 hours
316 193 7.73 7.90
100 61.0 7.73 8.01
31.6 19.3 7.76 8.14
10.0 6.10 7.79 8.29
3.16 1.93 7.83 8.38
1.00 0.610 7.89 8.54
Control 7.97 8.71

Dissolved oxygen:

Not measured

Salinity:

Not measured, freshwater conditions

Nominal and measured concentrations:

See window "Any other information on materials and methods incl. tables"

Details on test conditions:

TEST SYSTEM
- Test vessel: 250 mL Erlenmeyer flasks
- Type (delete if not applicable): open, cotton wool plugs
- Material, size, headspace, fill volume: sterile 250 mL Erlenmeyer flasks, test volume 100 mL
- Aeration: Test containers were placed on a rotary shaker and oscillated at appr. 70 rpm
- Type of flow-through (e.g. peristaltic or proportional diluter): None
- Renewal rate of test solution (frequency/flow rate): One application at test start
- Initial cells density: Nominal: approximately 2 - 5 x 103 cells/mL, Actual: mean 4332 cells/mL
- Control end cells density: Mean 717179 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6



GROWTH MEDIUM
- Standard medium used: yes



TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water:
Composition of Dilution Water
according to the guideline
Component Concentration [mg/L]
NH4Cl 15
MgCl2  6 H2O 12
CaCl2  2 H2O 18
MgSO4  7 H2O 15
KH2PO4 1.6
FeCl3  6 H2O 0.064
Na2EDTA  2 H2O 0.1
H3BO3 0.185
MnCl2  4 H2O 0.415
ZnCl2 3 x 10-3
Na2MoO4  2 H2O 7 x 10-3
CoCl2  6 H2O 1.5 x 10-3
CuCl2  2 H2O 1 x 10-5
NaHCO3 50
pH 8.1  0.2




OTHER TEST CONDITIONS
- Sterile test conditions: no
- Adjustment of pH: no
- Photoperiod: 24 h
- Light intensity and quality: 60 - 120 µE x m-2 x s-1, mean value: 67.9



EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: [counting chamber; electronic particle counter; fluorimeter; spectrophotometer; colorimeter] fluorimeter, Cell density and self fluorescence was measured daily via Chlorophyll-a- fluorescence, excitation at 436 nm, emission at 685 nm. Dilution water was used as background signal.



TEST CONCENTRATIONS

Results of the Range Finding Test (0-72 hours)
Nominal Test Item Concentration
[mg/L] Growth Rate Inhibition [%] Yield Inhibition [%]
100 56 95
10.0 13 48
1.00 2 9
0.1 3 13
0.01 1 8

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

117 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

is UVCB means 100% act. ingr.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI 99.7-138 mg/L

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

11.1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

is UVCB means 100% act.ingr.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI 6.75-16.5 mg/L

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test):
- Unusual cell shape: No
- Colour differences: No
- Flocculation: No
- Adherence to test vessels: No
- Aggregation of algal cells: No
- Other: No
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: No

Results with reference substance (positive control):

- Results with reference substance valid? Yes

Current Study Valid Range (average ± 3 x SD)
Growth Rate Inhibition
ErC50 0.631 0.682 ± 0.452
95 %
confidence
interval 0.583 - 0.687
Yield Inhibition
EyC50 0.256 0.314 ± 0.144
95 %
confidence
interval 0.247 - 0.265
SD = Standard deviation

Reported statistics and error estimates:

NOEC/LOEC were determined by calculation of statistical significance of growth rate and yield. As standard One Way Analysis of Variance (ANOVA) and DUNNETT’s test were used for NOEC/LOEC calculations. When running a One Way Analysis of Variance a Normality test and an Equal Variance test were done first. The SHAPIRO-WILK-Test was used to test for normally distributed populations. The Levene median test was used for equal variance. P-values for both Normality and Equal Variance tests are 0.05. The -value (acceptable probability of incorrectly concluding that there is a difference) is =0.05.
Detection of statistically significant differences were considered negligible when inhibition values were ≤ 10 %. These findings were marked “biologically not significant”. Following the recommendations of the UBA (HEGER ET AL. 1998), inhibitions > 10 % are significant.

Cell Densities

	Nominal concentration			Replicate		Cell density [cells/mL]
Test item [mg/L]		Active ingredient [mg/L a.i.]	No.		0 hours		24 hours	48 hours	72 hours
316		193	1		4332		9329	7797	7492
			2		4332		7747	8332	10219
			3		4332		8511	7036	7851
			Mean		4332		8529	7722	8521
100		61.0	1		4332		10254	19431	78554
			2		4332		10757	22157	84787
			3		4332		10552	20069	76038
			Mean		4332		10521	20552	79793
31.6		19.3	1		4332		10493	30973	159493
			2		4332		9395	33876	186408
			3		4332		9448	31350	197636
			Mean		4332		9779	32066	181179
10.0		6.10	1		4332		13531	81365	641408
			2		4332		14748	90193	544070
			3		4332		16622	85042	546272
			Mean		4332		14967	85533	577250
3.16		1.93	1		4332		16088	124647	600491
			2		4332		16553	102739	558286
			3		4332		17739	95706	533817
			Mean		4332		16793	107697	564198
1.00		0.610	1		4332		21777	107248	622003
			2		4332		22598	138151	731638
			3		4332		25721	137742	794129
			Mean		4332		23365	127714	715923
Control			1		4332		17506	119822	636187
			2		4332		19148	137245	789380
			3		4332		21604	108771	774253
			4		4332		18852	113425	648673
			5		4332		16056	123602	699543
			6		4332		22925	129389	755037
			Mean		4332		19349	122042	717179

Evaluation after 72 hours

                   Statistically significant differences of growth rates and yield compared to

                   control values are marked (+), not significant differences are marked (-).

Nominal concentration		Replicate	Growth rate		Growth rate inhibition	Yield		Inhibition of yield
Test item [mg/L]	Active ingredient [mg/L a.i.]	No.	[d^-1]		[%]	[cells/mL]		[%]
316	193	1		0.183	89.3		3160	99.6
		2		0.286	83.2		5887	99.2
		3		0.198	88.4		3519	99.5
		Mean	(+)	0.222	86.9	(+)	4189	99.4
100	61.0	1		0.966	43.3		74222	89.6
		2		0.991	41.8		80455	88.7
		3		0.955	43.9		71706	89.9
		Mean	(+)	0.971	43.0	(+)	75461	89.4
31.6	19.3	1		1.20	29.4		155161	78.2
		2		1.25	26.3		182076	74.5
		3		1.27	25.2		193304	72.9
		Mean	(+)	1.24	27.0	(+)	176847	75.2
10.0	6.10	1		1.67	2.12		637076	10.6
		2		1.61	5.34		539738	24.3
		3		1.61	5.26		541940	24.0
		Mean	(+)^1	1.63	4.24	(+)	572918	19.6
3.16	1.93	1		1.64	3.41		596159	16.4
		2		1.62	4.84		553954	22.3
		3		1.61	5.71		529485	25.7
		Mean	(+)^1	1.62	4.65	(+)	559866	21.5
1.00	0.610	1		1.66	2.72		617671	13.4
		2		1.71	-0.46		727306	-2.03
		3		1.74	-2.07		789797	-10.8
		Mean	(-)	1.70	0.06	(-)	711591	0.18
Control		1		1.66			631855
		2		1.74			785048
		3		1.73			769921
		4		1.67			644341
		5		1.70			695211
		6		1.72			750705
		Mean		1.70			712847

Section-by-Section and Average Specific Growth Rates of the Control Group

                   (0 – 72 hours)

	Replicate No.	Specific growth rate [d^-1]			Mean (0 - 72 h)	SD ±	CV [%]	Mean CV [%]
		section-by-section
		0 - 24 hours	24 - 48 hours	48 - 72 hours
Control	1	1.40	1.92	1.67	1.66	0.264	15.8	12.7
	2	1.49	1.97	1.75	1.74	0.242	13.9
	3	1.61	1.62	1.96	1.73	0.203	11.7
	4	1.47	1.80	1.74	1.67	0.174	10.4
	5	1.31	2.04	1.73	1.70	0.367	21.7
	6	1.67	1.73	1.76	1.72	0.0497	2.89
				Mean	1.70
				SD ±	0.03
				CV [%]	1.81

Validity criteria fulfilled:

yes

Conclusions:

In this study Sodium methyl oleyl taurate was found to inhibit the growth of the freshwater green alga Desmodesmus subspicatus after 72 hours. The NOEC-values of the test item for inhibition of growth rate and inhibition of yield after 72 hours were 10.0 mg/L and 1.00 mg/L, respectively. The LOEC-values of the test item for inhibition of growth rate and inhibition of yield after 72 hours were 31.6 mg/L and 3.16 mg/L, respectively. The EC50-values of the test item for inhibition of growth rate (ErC50) and yield (EyC50) with 95 % Confidence Intervals after 72 hours were 117 (99.7 – 138) and 19.5 (14.6 – 24.9) mg/L, respectively.
The NOEC-values of the active ingredient for inhibition of growth rate and inhibition of yield after 72 hours were 6.10 mg/L a.i. and 0.610 mg/L a.i., respectively. The LOEC-values of the active ingredient for inhibition of growth rate and inhibition of yield after 72 hours were 19.3 mg/L a.i. and 1.93 mg mg/L a.i., respectively. The EC50-values of the active ingredient for inhibition of specific growth rate (ErC50) and yield (EyC50) with 95 % Confidence Intervals after 72 hours were 71.6 (60.8 – 84.3) and 11.9 (8.90 – 15.2) mg/L a.i., respectively.
All effect levels are given based on nominal concentrations of Sodium methyl oleyl taurate.

Executive summary:

The toxicity of Sodium methyl oleyl taurate(batch number: DEB4003485)to the unicellular freshwater green alga Desmodesmus subspicatus was determined according to the principles of OECD 201 (2006) at Dr.U.Noack-Laboratorienin 31157 Sarstedt, Germany from 2012-12-10 to
2012-12-13. The aim of the study was to assess the effects on growth rate and yield over a period of 72 hours.

The study was conducted under static conditions with an initial cell density of 4332 cells/mL. Based on a preliminary test 6 concentration levels were tested in a geometrical series with a nominal dilution factor of : 1.00 - 3.16 - 10.0 - 31.6 - 100 - 316 mg/L corresponding to active ingredient (a.i.) concentrations of 0.610 - 1.93 - 6.10 - 19.3 - 61.0 - 193 mg a.i./L. Three replicates were tested for each test item concentration and six replicates for the control. Environmental conditions were determined to be within the acceptable limits.

The concentrations of the test item Sodium methyl oleyl taurate (C18:1-part) were analytically verified by LC-MS/MS at the start and the end of the exposure of all test concentrations and the control.
The measured concentrations of the test item Sodium methyl oleyl taurateat the start of the exposure were in the range of 93 to 103 % and after 72 hours in the range of 82 to 98 % of the nominal values. Therefore, all effect values given are based on the nominal test item and active ingredient (a.i.) concentrations of Sodium methyl oleyl taurate.

IMPORTANT NOTE TO AVOID CONFUSION:

As the registration substance is a UVCB this means that the test item is 100% active ingredient.

Table 1:    NOEC, LOEC, EC - values and 95 % Confidence Intervals of Sodium methyl oleyl taurate
(0-72 hours)

                   based on nominal test item and active ingredient (a.i.) concentrations [mg/L]

                               

	Growth Rate Inhibition
	Nominal [mg/L]
NOEC	10.0
LOEC	31.6
ErC10	11.1 (6.75 – 16.5)
ErC20	29.2 (22.6 – 36.3)
ErC50	117 (99.7 – 138)
	Inhibition of Yield
NOEC	1.00
LOEC	3.16
EyC10	2.55 ( – 9.81)
EyC20	9.28 (5.65 – 14.4)
EyC50	19.5 (14.6 – 24.9)

Description of key information

In this study Sodium methyl oleyl taurate was found to inhibit the growth of the freshwater green alga Desmodesmus subspicatus after 72 hours. The NOEC-values of the test item for inhibition of growth rate and inhibition of yield after 72 hours were 10.0 mg/L and 1.00 mg/L, respectively. The LOEC-values of the test item for inhibition of growth rate and inhibition of yield after 72 hours were 31.6 mg/L and 3.16 mg/L, respectively. The EC50-values of the test item for inhibition of growth rate (ErC50) and yield (EyC50) with 95 % Confidence Intervals after 72 hours were 117 (99.7 – 138) and 19.5 (14.6 – 24.9) mg/L, respectively.

The NOEC-values of the active ingredient for inhibition of growth rate and inhibition of yield after 72 hours were 6.10 mg/L a.i. and 0.610 mg/L a.i., respectively. The LOEC-values of the active ingredient for inhibition of growth rate and inhibition of yield after 72 hours were 19.3 mg/L a.i. and 1.93 mg mg/L a.i., respectively. The EC50-values of the active ingredient for inhibition of specific growth rate (ErC50) and yield (EyC50) with 95 % Confidence Intervals after 72 hours were 71.6 (60.8 – 84.3) and 11.9 (8.90 – 15.2) mg/L a.i., respectively.

All effect levels are given based on nominal concentrations of Sodium methyl oleyl taurate.

Key value for chemical safety assessment

EC50 for freshwater algae:

117 mg/L

EC10 or NOEC for freshwater algae:

11.1 mg/L