Reaction mass of Cobaltate(3-), bis[2-[[[3-[2-[1-[[(2-chlorophenyl)amino]carbonyl]-2-(oxo-κO)propyl]diazenyl-κN1]-4-(hydroxy-κO)phenyl]sulfonyl]amino]benzoato(3-)]-, sodium (1:3) and tetrasodium bis[2-[[[3-[[1-[(2-chloroanilino)carbonyl]-2-oxopropyl]azo]-4-hydroxyphenyl]sulphonyl]amino]benzoato(3-)]cobaltate(4-)

Toxicity to aquatic plants other than algae

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic plants other than algae

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2014

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 221 (Lemna sp. Growth Inhibition Test)

Deviations:

yes

Remarks:

No major (the pH increased by more than 1.5 units during the test at several time points. The test is still valid, as it was shown that validity criteria are met).

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Name: FAT 92368/Y
Batch number: 1309001
EAS Test item code: 2014-000177
Appearance / colour: solid / orange brown
Date of analysis: 04 Oct 2013
Stability in solution: sufficient for the test purpose (at least 1h)
Expiry date: 30 Sep 2018
Storage conditions: ambient (+5 to +30 °C), dark, dry

Analytical monitoring:

yes

Details on sampling:

Samples were taken at 0 hours (initial value) from fresh test solution, after 3 and 5 days from fresh and aged test solution and after 7 days from aged test solution from control and all test item concentrations. Nominal test item concentrations of 0.954, 3.05, 9.77, 31.3 and 100 mg/L and control were analysed at t=3d from fresh test solutions and at t=3d and 5d from fresh and aged test solutions.

Vehicle:

no

Details on test solutions:

The necessary amount of test item for preparing the stock solution S1 was weighed and transferred to a volumetric flask. Test medium was added up to the bench mark and the solution was homogenised by shaking. Lower concentrations of the test item were prepared by dilution of the stock solution.

Test organisms (species):

Lemna gibba

Details on test organisms:

The test organism was the aquatic plant duckweed, Lemna gibba G3 (Alismatales: Araceae). The plants used in this study were taken as aseptic clones from the cultivation of the testing facility. They were primarily cultured by Dr. Janet Slovin, Horticulture Crops Quality Laboratory, U.S. Department of Agriculture. BARC-West, Bldg. 050 HH-4, Beltsville, MD 20705, U.S.A. The plants were cultured in appropriate glass beakers under continuous illumination with OSRAM light tubes (L30/32-930 and L30/W72-965), at 6500 - 7000 lux. The temperature was 24 ± 2 °C. Before the test was started, plants were cultured under the same illumination and temperature conditions as those to be used for the toxicity test. Young, light-green plants of similar size and comprising 2 - 5 fronds were transferred onto fresh medium and cultured for approx. 3 weeks prior to testing, with two further transfers onto fresh medium before initiating the test.

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

7 d

Hardness:

none

Test temperature:

22.0 – 25.0 °C

pH:

7.47 – 7.63

Dissolved oxygen:

none

Salinity:

none

Conductivity:

none

Nominal and measured concentrations:

0.954, 3.05, 9.77, 31.3 and 100 mg/L

Details on test conditions:

Light intensity (mean): ~ 6800 lux
Light tubes: OSRAM light tubes (L 30 W77 and L 30 W25)
Exposure to light: continuously
Test vessels: 250 mL glass beakers filled with 150 mL
Number of fronds: 12 per test vessel

Duration:

7 d

Dose descriptor:

EC10

Effect conc.:

0.272 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of frond number

Remarks on result:

other: 0.000545-1.32

Duration:

7 d

Dose descriptor:

EC20

Effect conc.:

0.856 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of frond number

Remarks on result:

other: 0.0116-3.00

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

7.69 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of frond number

Remarks on result:

other: 1.83-32.0

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

< 0.534 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of frond number

Duration:

7 d

Dose descriptor:

LOEC

Effect conc.:

0.534 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of frond number

Duration:

7 d

Dose descriptor:

EC10

Effect conc.:

0.837 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of frond number

Remarks on result:

other: 0.0000125-4.21

Duration:

7 d

Dose descriptor:

EC20

Effect conc.:

4.05 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of frond number

Remarks on result:

other: 0.0165-14.9

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

> 96 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of frond number

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

0.534 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of frond number

Duration:

7 d

Dose descriptor:

LOEC

Effect conc.:

2.01 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of frond number

Duration:

7 d

Dose descriptor:

EC10

Effect conc.:

0.428 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of dry weight

Remarks on result:

other: 0.0120-1.52

Duration:

7 d

Dose descriptor:

EC20

Effect conc.:

1.24 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of dry weight

Remarks on result:

other: 0.102-3.40

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

9.38 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of dry weight

Remarks on result:

other: 3.41-28.2

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

0.534 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of dry weight

Duration:

7 d

Dose descriptor:

LOEC

Effect conc.:

2.01 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield of dry weight

Duration:

7 d

Dose descriptor:

EC10

Effect conc.:

1.33 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of dry weight

Remarks on result:

other: 0.0101-4.84

Duration:

7 d

Dose descriptor:

EC20

Effect conc.:

5.45 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of dry weight

Remarks on result:

other: 0.411-15.3

Duration:

7 d

Dose descriptor:

EC50

Effect conc.:

> 96 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of dry weight

Duration:

7 d

Dose descriptor:

NOEC

Effect conc.:

0.534 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of dry weight

Duration:

7 d

Dose descriptor:

LOEC

Effect conc.:

2.01 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: growth rate of dry weight

Reported statistics and error estimates:

The statistical evaluation for day 7 was performed for yield of frond numbers, growth rate of frond numbers, growth rate dry weight and yield of dry weight. A test for normality of the data was performed by calculating the Shapiro-Wilk’s statistic, a test for homogeneity of the data was performed according to Levene. The NOEC and LOEC were determined by using a multiple comparison method (Dunnett`s t-test and Jonckheere-Terpstra). EC10 and EC20 values were determined by probit analysis following normal or logistic procedure. The EC50 value for yield of frond number and yield of dry weight was determined by probit analysis following normal procedure. The calculation of EC50 values for growth rate of frond numbers and growth rate of dry weight was not indicated since inhibition was below 50 %. The evaluation of data was performed by SAS® (2002-2010).

	Test item [mg/L] nominal
Endpoint	EC10	95 % confidence limit	EC20	95 % confidence limit	EC50	95 % confidence limit	NOEC	LOEC
Yield of frond numbers	0.513^1)	0.00208-2.10	1.44^1)	0.0318-4.43	10.4^1)	2.88 – 37.0	<0.954^2)	0.954^2)
Growth rate of frond numbers	1.41^4)	0.0000861-6.00	5.82^4)	0.0471-18.6	>100	-	0.954^2)	3.05^2)
Yield of dry weight	0.771^1)	0.0313-2.41	2.00^1)	0.213-4.96	12.4^1)	5.00 – 33.2	0.954^3)	3.05^3)
Growth rate of dry weight	2.14^4)	0.0283-6.81	7.61^4)	0.766-19.2	>100	-	0.954^3)	3.05^3)

1) Probit analysis following the normal distribution

2) following Dunnetts-t-test

3) following Jonckheere-Terpstra

4) Probit analysis following the logistic distribution

*based on geometric mean of each concentration level

- not applicable

Validity of test :The  test is valid as the doubling time of control frond numbers was calculated to be 47.7 h, corresponding to 1.987 days at day 7. This is less than the maximum allowable doubling time set in OECD guideline 221 (60 h, 2.5 days).

Validity criteria fulfilled:

yes

Conclusions:

The nominal EC50-value for yield of frond numbers was determined to be 10.4 mg/L, the nominal EC50-value for yield of dry weight was determined to be 12.4 mg/L. The nominal EC50-value for growth rate of frond numbers and growth rate of dry weight was determined to be >100 mg/L. The actual EC50-value for yield of frond numbers was determined to be 7.69 mg/L, the actual EC50-value for yield of dry weight was determined to be 9.38 mg/L. The actual EC50-value for growth rate of frond numbers and growth rate of dry weight was determined to be >96.0 mg/L.

Executive summary:

FAT 92368/Y was tested for toxicity to plants other than algae. This test was conducted according to OECD TG 221. The initial mean measured concentration of FAT 92368/Y in the test item solutions was 88 % of nominal. The mean measured content of FAT 92368/Y in aged test item solutions was 62 %. Therefore, toxicological endpoints were evaluated using nominal concentrations of the test item and actual concentrations based on the geometric mean concentration of each test item concentration level. Significant inhibitory effects were determined for yield of frond numbers at all concentration levels. For growth rate of frond numbers, yield of dry weight and growth rate of dry weight significant inhibitory effects were observed at 3.05, 9.77, 31.3 and 100 mg/L (nominal). The LOEC for yield of frond numbers was determined to be 0.954 mg/L (nominal), the LOEC for growth rate of frond numbers, yield of dry weight and growth rate of dry weight was at 3.05 mg/L (nominal). The nominal EC10-value for yield of frond numbers was determined to be 0.513 mg/L, the nominal EC10-value for yield of dry weight was determined to be 0.771 mg/L. The nominal EC10-value for growth rate of frond numbers was determined to be 1.41 mg/L and the nominal EC10-value for growth rate of dry weight was determined to be 2.14 mg/L. The actual EC10-value for yield of frond numbers was determined to be 0.272 mg/L, the actual EC10-value for yield of dry weight was determined to be 0.428 mg/L. The actual EC10-value for growth rate of frond numbers was determined to be 0.837 mg/L and the actual EC10-value for growth rate of dry weight was determined to be 1.33 mg/L. The nominal EC20-value for yield of frond numbers was determined to be 1.44 mg/L, the nominal EC20-value for yield of dry weight was determined to be 2.00 mg/L. The nominal EC20-value for growth rate of frond numbers was determined to be 5.82 mg/L and the nominal EC20-value for growth rate of dry weight was determined to be 7.61 mg/L. The actual EC20-value for yield of frond numbers was determined to be 0.856 mg/L, the actual EC20-value for yield of dry weight was determined to be 1.24 mg/L. The actual EC20-value for growth rate of frond numbers was determined to be 4.05 mg/L and the actual EC20-value for growth rate of dry weight was determined to be 5.45 mg/L. The nominal EC50-value for yield of frond numbers was determined to be 10.4 mg/L, the nominal EC50-value for yield of dry weight was determined to be 12.4 mg/L. The nominal EC50-value for growth rate of frond numbers and growth rate of dry weight was determined to be >100 mg/L. The actual EC50-value for yield of frond numbers was determined to be 7.69 mg/L, the actual EC50-value for yield of dry weight was determined to be 9.38 mg/L. The actual EC50-value for growth rate of frond numbers and growth rate of dry weight was determined to be >96.0 mg/L. The test is valid as the doubling time of control frond numbers was calculated to be 47.7 h, corresponding to 1.987 days at day 7. This is less than the maximum allowable doubling time set in OECD guideline 221 (60 h, 2.5 days).

Description of key information

FAT 92368/Y was tested for toxicity to plants other than algae. This test was conducted according to OECD TG 221. The initial mean measured concentration of FAT 92368/Y in the test item solutions was 88 % of nominal. The mean measured content of FAT 92368/Y in aged test item solutions was 62 %. Therefore, toxicological endpoints were evaluated using nominal concentrations of the test item and actual concentrations based on the geometric mean concentration of each test item concentration level.

 

Toxicological endpoints for FAT 92368/Y:

Significant inhibitory effects were determined for yield of frond numbers at all concentration levels. For growth rate of frond numbers, yield of dry weight and growth rate of dry weight significant inhibitory effects were observed at 3.05, 9.77, 31.3 and 100 mg/L (nominal). The LOEC for yield of frond numbers was determined to be 0.954 mg/L (nominal), the LOEC for growth rate of frond numbers, yield of dry weight and growth rate of dry weight was at 3.05 mg/L (nominal). The nominal EC10-value for yield of frond numbers was determined to be 0.513 mg/L, the nominal EC10-value for yield of dry weight was determined to be 0.771 mg/L. The nominal EC10-value for growth rate of frond numbers was determined to be 1.41 mg/L and the nominal EC10-value for growth rate of dry weight was determined to be 2.14 mg/L.

 

The actual EC10-value for yield of frond numbers was determined to be 0.272 mg/L, the actual EC10-value for yield of dry weight was determined to be 0.428 mg/L. The actual EC10-value for growth rate of frond numbers was determined to be 0.837 mg/L and the actual EC10-value for growth rate of dry weight was determined to be 1.33 mg/L.

 

The nominal EC20-value for yield of frond numbers was determined to be 1.44 mg/L, the nominal EC20-value for yield of dry weight was determined to be 2.00 mg/L. The nominal EC20-value for growth rate of frond numbers was determined to be 5.82 mg/L and the nominal EC20-value for growth rate of dry weight was determined to be 7.61 mg/L.

 

The actual EC20-value for yield of frond numbers was determined to be 0.856 mg/L, the actual EC20-value for yield of dry weight was determined to be 1.24 mg/L. The actual EC20-value for growth rate of frond numbers was determined to be 4.05 mg/L and the actual EC20-value for growth rate of dry weight was determined to be 5.45 mg/L.

 

The nominal EC50-value for yield of frond numbers was determined to be 10.4 mg/L, the nominal EC50-value for yield of dry weight was determined to be 12.4 mg/L. The nominal EC50-value for growth rate of frond numbers and growth rate of dry weight was determined to be >100 mg/L.

 

The actual EC50-value for yield of frond numbers was determined to be 7.69 mg/L, the actual EC50-value for yield of dry weight was determined to be 9.38 mg/L. The actual EC50-value for growth rate of frond numbers and growth rate of dry weight was determined to be >96.0 mg/L.

 

The test is valid as the doubling time of control frond numbers was calculated to be 47.7 h, corresponding to 1.987 days at day 7. This is less than the maximum allowable doubling time set in OECD guideline 221 (60 h, 2.5 days).

Key value for chemical safety assessment

EC50 for freshwater plants:

96 mg/L

EC10 or NOEC for freshwater plants:

0.837 mg/L