Bis(dibutyldithiocarbamato-S,S')copper

Toxicity to terrestrial plants

Link to relevant study record(s)

Reference

Endpoint:

toxicity to terrestrial plants: long-term

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 208 (Terrestrial Plants Test: Seedling Emergence and Seedling Growth Test)

Deviations:

yes

Remarks:

Due to deficiencies occurred in the control groups during the definitive test for 3 species (validity criteria not met), an additional definitive test was conducted with 3 different species. This study report contains results from both tests #1 and #2.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

not required

Details on sampling:

No sampling.

Vehicle:

no

Details on preparation and application of test substrate:

APPLICATION OF TEST ITEM

The experimental soils preparation was determined from the following parameters:
- the total amount of soil per treatment
- the amount of water to be added to the soil to reach the chosen WHC
- the 5M soil moisture determined at the test facility
- the treatment rate

As the test item is a solid powder chemical which is insoluble in water, it was directly mixed with the test soils.


TEST #1
For each treatment group 9 kg of 5M soil (corresponding to 8.19 kg dry weight) were necessary. To achieve the WHC80 and considering the soil moisture (9% ), a total of 868 mL tap water was added then thoroughly manually mixed. The test item was then mixed with the experimental soils. Directly after soil preparation, the pots were filled with soil and the seeds were sown.
Detailed information about the preparation of the test soil can be found in Table below.


TEST #2
For each treatment group 7.5 kg of 5M soil (corresponding to 7.035 kg dry weight) were necessary. To achieve the WHC40 and considering the soil moisture (6.2%), a total of 555 mL tap water was added then thoroughly manually mixed. The test item was then mixed with the experimental soils. Directly after soil preparation, the pots were filled with soil and the seeds were sown.
Detailed information about the preparation of the test soil can be found in Table below.

Species:

Sorghum vulgare

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

#2

Monocotyledonae
Name Sorghum
Family Poaceae
Species Sorghum bicolore
Seed supplier AGPM
Date of sowing 20 March 2018
Date of 50% emergence
in the control group 27 March 2018
Date of observation period end 10 April 2018
Ø Pot size Around 12 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Species:

Cucumis sativus

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

#2

Dicotyledonae
Name Cucumber
Family Cucurbitaceae
Species Cucumis sativus
Seed supplier Le Paysan
Date of sowing 20 March 2018
Date of 50% emergence
in the control group 30 March 2018
Date of observation period end 13 April 2018
Ø Pot size Around 12 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Species:

Pisum sativum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

#2

Dicotyledonae
Name Pea
Family Fabaceae
Species Pisum sativum
Seed supplier Le Paysan
Date of sowing 20 March 2018
Date of 50% emergence
in the control group 26 March 2018
Date of observation period end 10 April 2018
Ø Pot size Around 12 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Species:

Allium cepa

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

#1

Monocotyledonae
Name Onion
Family Liliaceae
Species Allium cepa
Seed supplier Vilmorin
Date of sowing 26 January 2017
Date of 50% emergence
in the control group NC
Date of observation period end 21 February 2017
Ø Pot size Around 9 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Species:

Triticum aestivum

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

#1

Monocotyledonae
Name Wheat
Family Poaceae
Species Triticum monococcum
Seed supplier Primeal
Date of sowing 26 January 2017
Date of 50% emergence
in the control group 31 January 2017
Date of observation period end 15 February 2017
Ø Pot size Around 12 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Species:

other: Solanum Lycopersicum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

#1

Dicotyledonae
Name Tomato
Family Solanaceae
Species Solanum Lycopersicum
Seed supplier Vilmorin
Date of sowing 26 January 2017
Date of 50% emergence
in the control group 2 February 2017
Date of observation period end 22 February 2017
Ø Pot size Around 12 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Species:

Daucus carota

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

#1

Dicotyledonae
Name Carrot
Family Apiaceae
Species Daucus carota
Seed supplier Vilmorin
Date of sowing 26 January 2017
Date of 50% emergence
in the control group 03 February 2017
Date of observation period end 22 February 2017
Ø Pot size Around 9 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Species:

Helianthus annuus

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

#1

Dicotyledonae
Name Sunflower
Family Asteraceae
Species Helianthus annuus
Seed supplier AGPM
Date of sowing 26 January 2017
Date of 50% emergence
in the control group 31 January 2017
Date of observation period end 16 February 2017
Ø Pot size Around 12 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Species:

Vicia sativa

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

#1

Dicotyledonae
Name Vetch
Family Fabaceae
Species Vicia sativa
Seed supplier Magellan
Date of sowing 26 January 2017
Date of 50% emergence
in the control group NC
Date of observation period end 21 February 2017
Ø Pot size Around 9 cm
Number of treatment groups 7
Number of seeds/treatment 30
Number of seeds/pot 6
Number of pots/treatment 5
Prior seed treatment/sterilization: no fungicides, insecticides or repellents treatments

Test type:

seedling emergence and seedling growth test

Study type:

laboratory study

Substrate type:

natural soil

Limit test:

no

Total exposure duration:

28 d

Remarks:

From 3 to 4 weeks exposure

Test temperature:

#1: 19.7°C - 27.7 °C (min -max)
#2: 21.4°C - 22.3 °C (min -max)

pH:

7.3

Moisture:

#1: 9%
#2: 6.2%

Details on test conditions:

SOILS TYPE AND GROWING CONDITIONS

The soil substrates used for this study was two loamy sand soils provided by Lufa Speyer (Soil type 5M, batch F5M4816 & F5M0818). The specifications of each soils are presented in Table below. Both soils were not pasteurized nor heat treated; no fertiliser or plant protection products were used.

The test plants were grown in an air-conditioned chamber that allows test pots to be incubated under accurately controlled conditions as follows:
- temperature : range of 20 to 24°C, controlled at ± 2°C;
- relative humidity: 70% ± 25%
- photoperiod: 16 hours light/8 hours darkness
- light intensity: 350 ± 50 µE/m²/s

Plant plastic pots were used for the study (see details in Test Organisms paragraph), their position was rotated randomly every seven days after sowing to ensure a uniform light regime for all plants.

Bottom watering was done regularly with tap water to the plant saucer of each pot, providing the plant root with a sufficient water supply: during the first week and at least three times, some potting containers were randomly weighed in order to check the loss of water through evaporation, then the quantity lost was reintroduced in all pots.


ENVIRONMENTAL CONDITIONS

Temperature and humidity were recorded as follows:

Test #1: temperature and humidity ranged from 19.7 to 27.7 °C and from 62.3 to 99.8%, respectively. The light intensity was measured in a range from 110 to 180 µE/m²/s.
Test #2: temperature and humidity ranged from 21.4 to 22.3 °C and from 64.2 to 89.5%, respectively. The light intensity was measured in a range from 113 to 557 µE/m²/s.

These records imply a deviation from the test conditions recommended in the study plan. However, the variances were of a short term and not significant for all retained species as the test validity criteria for the retained species were met


OBSERVATIONS

The plants were observed at least 3 days per week for emergence, mortality and visual phytotoxicity. Visible detrimental effects on different parts of the plant were assessed (abnormalities in appearance of the emerged seedlings, stunted growth, chlorosis, discoloration, mortality, and effects on plant development).

At the end of the test, percentage of emergence and post-emergence survival were calculated. The final biomass was measured using final average dry shoot weight of surviving plants, by harvesting the shoots at the soil surface and drying them at around 60°C during around 24h.

According to the OECD Test Guideline 208, the observation period is typically 14 to 21 days. The calendar assessment was adjusted in order to ensure sufficient time within a single work day to properly assess each of the biological parameters for each species.

Nominal and measured concentrations:

Nominal concentrations (the spacing factor slightly exceed 3 ):
0 ; 1.6 ; 5.0 ; 16.0 ; 51.2 ; 163.8 ; 524.0 mg/kg soil dry weight

Reference substance (positive control):

no

Species:

Sorghum bicolor

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Species:

Sorghum bicolor

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

seedling emergence

Species:

Daucus carota

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Species:

Daucus carota

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

seedling emergence

Species:

Cucumis sativus

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Species:

Cucumis sativus

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

seedling emergence

Species:

Pisum sativum

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Species:

Pisum sativum

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

seedling emergence

Species:

other: Solanum lycopersicum

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Species:

other: Solanum lycopersicum

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

seedling emergence

Species:

other: Triticum monococcum

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

51.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

other: inhibition of shoot dry weight

Species:

other: Triticum monococcum

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Species:

other: Triticum monococcum

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 524 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

seedling emergence

Details on results:

Note 1: due to deficiencies occurred in the control groups during the definitive test (i.e. low emergence rates for onions (Allium cepa) and vetch (Vicia sativa)and low survival rates for sunflowers (Helianthus annuus)), the test validity criteria for these species were not met and thus an additional definitive test was conducted with three different species. This study report contains results from both tests #1 and #2.

Note 2: An unexpected result was observed for the species Sorghum at the 163.8 mg/Kg/sdw test concentration: no ermergence was observed in one pot (replicate d). A technical mistake was suspected, this replicate was not considered for the final calculation.

SEEDLING EMERGENCE AND SURVIVAL

Seedling emergence and survival results are summarized in Tables below.

The deficiencies occurred in the control groups during the test #1 (i.e. low emergence rates for onions (Allium cepa) and vetch (Vicia sativa) and low survival rates for sunflowers (Helianthus annuus) are highlighted. As the validity criteria were not fulfilled these species are not further considered in this Study Report. Nevertheless all obtained data are archived with the study raw data.

Seedling emergence was not statistically significantly reduced compared to the control (i.e. NOEC is superior or equal to the highest tested concentration) for all valid species tested. Inhibitions of seedling emergence did not follow any dose-response relationship, as indicated in the appended statistical analysis. As a consequence EC10/50 for seedling emergence could not be determined in the course of this study.

In addition, the statistical analysis did not detect any significant difference in terms of emergence or post-emergence survival between the control groups and any treatment in any tested species (i.e. NOEC is superior or equal to the highest tested concentration).

No hazard was identified up to the highest tested concentration for seedling emergence or survival for any plant species tested during the course of this study.


PHYTOTOXICITY
Only slight visual symptoms of chlorosis or wilting could be observed in the course of this study for Pea and Cucumber. No other phytotoxicity was reported. .


SHOOT WEIGHT
Dry weights details for each species are presented in Tables below.

Shoot dry weight was not statistically significantly reduced compared to the control (i.e. NOEC is superior or equal to the highest tested concentration) for the following tested species: Tomato (Solanum Lycopersicum), Sorghum (Sorghum bicolore), Carrot (Daucus carota) and Pea (Pisum sativum).

Inhibitions of shoot dry weight for Wheat (Triticum monococcum) was determined to be statistically significantly reduced compared to the control from the 163.8 mg/Kg/sdw tested concentration: as a consequence LOEC = 163.8 mg/Kg/sdw and NOEC = 51.2 mg/Kg/sdw.

Inhibitions of shoot dry weight for Cucumber (Cucumis sativus) did not follow any dose-response relationship, as indicated in the appended statistical analysis. As a consequence NOEC & EC10/50 for dry weight inhibition could not be determined for this species.

For any species no dose-response relationship was detectable, EC10/50 for shoot dry weight inhibition could thus not be determined during the course of this study

Reported statistics and error estimates:

All statistical calculations were performed with the ToxRatPro software (Version 3.2.1), using the default template for the OECD 208 guideline

Seedling emergence and survival– Summary (Definitive Test #1)

Treatment (mg/Kg dw)	Onion			Wheat			Tomato			Carrot			Sunflower			Vetch
	Nb E	% E	% S	Nb E	% E	% S	Nb E	% E	% S	Nb E	% E	% S	Nb E	% E	% S	Nb E	% E	% S
Control	2	6.7	100.0	25	83.3	100.0	23	76.7	100.0	24	80.0	100.0	29	96.7	86.2	8	26.7	62.5
1.6	7	23.3	100.0	30	100.0	100.0	26	86.7	100.0	24	80.0	100.0	27	90.0	92.6	14	46.7	100.0
5.0	8	26.7	100.0	23	76.7	100.0	27	90.0	100.0	26	86.7	100.0	30	100.0	90.0	15	50.0	86.7
16.0	6	20.0	83.3	27	90.0	100.0	30	100.0	96.7	17	56.7	100.0	29	96.7	96.6	13	43.3	84.6
51.2	13	43.3	46.2	29	96.7	100.0	29	96.7	93.1	20	66.7	100.0	29	96.7	89.7	14	46.7	100.0
163.8	9	30.0	55.6	19	63.3	100.0	27	90.0	100.0	21	70.0	90.5	29	96.7	82.8	17	56.7	100.0
524.0	11	36.7	100.0	26	86.7	100.0	25	83.3	100.0	20	66.7	100.0	29	96.7	89.7	20	66.7	95.0

Colored cells correspond to invalid criteria

Seedling emergence and survival– Summary (Definitive Test #2)

Treatment (mg/Kg dw)	Sorghum			Cucumber			Pea
	Nb E	% E	% S	Nb E	% E	% S	Nb E	% E	% S
Control	21	70.0	100.0	25	83.3	100.0	25	83.3	96.0
1.6	20	66.7	100.0	29	96.7	100.0	25	83.3	96.0
5.0	21	70.0	100.0	28	93.3	96.4	26	86.7	100.0
16.0	26	86.7	100.0	26	86.7	100.0	27	90.0	100.0
51.2	22	73.3	100.0	27	90.0	100.0	23	76.7	100.0
163.8	19*	63.3	100.0	28	93.3	100.0	26	86.7	96.2
524.0	22	73.3	100.0	28	93.3	100.0	28	93.3	100.0

Nb E: number of emerged seeds

% E: % of emerged seeds

% S: % of survival

*The replicate d was not considered for the final calculation

Raw data – Fresh and dry weight (Definitive test #1 / Wheat)

Treatment (mg/Kg dw)	Replicate	Fresh weight				Dry weight
		Per pot	Mean	SD	% inhib	Per pot	Mean	SD	% inhib
Control	a	1,3022	1,2827	0,1094	n.a.	0,1619	0,1576	0,0272	n.a.
	b	1,1905				0,1589
	c	1,4511				0,1989
	d	1,2901				0,1263
	e	1,1797				0,1420
1.6	a	1,1744	1,4389	0,2380	-12,2%	0,1429	0,1782	0,0298	-13,0%
	b	1,7119				0,2091
	c	1,3973				0,1747
	d	1,6553				0,2077
	e	1,2557				0,1564
5.0	a	0,6311	1,1915	0,3574	7,1%	0,0734	0,1499	0,0489	4,9%
	b	1,0353				0,1294
	c	1,4107				0,1917
	d	1,4223				0,1741
	e	1,4581				0,1808
16.0	a	1,3424	1,2835	0,1684	-0,1%	0,1707	0,1553	0,0242	1,4%
	b	1,3076				0,1551
	c	1,0250				0,1216
	d	1,4879				0,1846
	e	1,2544				0,1447
51.2	a	1,5213	1,3930	0,1915	-8,6%	0,1991	0,1763	0,0215	-11,9%
	b	1,2900				0,1675
	c	1,1616				0,1499
	d	1,3462				0,1670
	e	1,6459				0,1981
163.8	a	0,7071	0,8192	0,1397	36,1%	0,0888	0,1019	0,0181	35,3%
	b	0,7987				0,0994
	c	0,9826				0,1289
	d	0,6660				0,0833
	e	0,9415				0,1091
524.0	a	0,5044	0,9273	0,3318	27,7%	0,0596	0,1202	0,0453	23,7%
	b	0,7011				0,0940
	c	1,0754				0,1390
	d	1,3552				0,1784
	e	1,0005				0,1300

n.a.: not applicable

Raw data – Fresh and dry weight (Definitive test #1 / Tomato)

Treatment (mg/Kg dw)	Replicate	Fresh weight				Dry weight
		Per pot	Mean	SD	% inhib	Per pot	Mean	SD	% inhib
Control	a	1,7810	1,2709	0,3168	n.a.	0,1833	0,1362	0,0378	n.a.
	b	1,2974				0,1493
	c	1,2538				0,1511
	d	1,0631				0,1109
	e	0,9593				0,0866
1.6	a	2,1021	1,6784	0,4528	-32,1%	0,2333	0,1929	0,0501	-41,6%
	b	1,4995				0,1784
	c	2,0410				0,2479
	d	0,9922				0,1215
	e	1,7573				0,1834
5.0	a	2,1921	1,6737	0,3424	-31,7%	0,2369	0,2009	0,0314	-47,5%
	b	1,8185				0,2245
	c	1,5527				0,2046
	d	1,3087				0,1627
	e	1,4963				0,1759
16.0	a	1,6902	1,3295	0,2135	-4,6%	0,1699	0,1518	0,0182	-11,5%
	b	1,2726				0,1569
	c	1,1754				0,1516
	d	1,3385				0,1592
	e	1,1708				0,1216
51.2	a	1,0537	1,5506	0,3646	-22,0%	0,1147	0,1652	0,0408	-21,2%
	b	1,6439				0,1709
	c	1,3425				0,1638
	d	2,0079				0,2270
	e	1,7049				0,1495
163.8	a	1,1476	1,4485	0,2263	-14,0%	0,1149	0,1530	0,0290	-12,3%
	b	1,4096				0,1635
	c	1,7727				0,1896
	d	1,3949				0,1629
	e	1,5175				0,1340
524.0	a	0,9259	1,0282	0,2640	19,1%	0,0943	0,1089	0,0298	20,1%
	b	1,4814				0,1610
	c	0,9519				0,1057
	d	0,7926				0,0885
	e	0,9890				0,0950

n.a.: not applicable

Raw data – Fresh and dry weight (Definitive test #1 / Carrot)

Treatment (mg/Kg dw)	Replicate	Fresh weight				Dry weight
		Per pot	Mean	SD	% inhib	Per pot	Mean	SD	% inhib
Control	a	0,1255	0,2436	0,0974	n.a.	0,0164	0,0343	0,0142	n.a.
	b	0,1514				0,0216
	c	0,2923				0,0416
	d	0,3205				0,0453
	e	0,3285				0,0465
1.6	a	0,1851	0,2255	0,0584	7,4%	0,0262	0,0339	0,0104	1,2%
	b	0,2802				0,0449
	c	0,1435				0,0200
	d	0,2579				0,0400
	e	0,2609				0,0383
5.0	a	0,2180	0,2604	0,0388	-6,9%	0,0345	0,0394	0,0048	-14,8%
	b	0,2394				0,0378
	c	0,3000				0,0454
	d	0,2412				0,0358
	e	0,3035				0,0433
16.0	a	0,2370	0,1723	0,0671	29,3%	0,0317	0,0237	0,0083	30,7%
	b	0,0911				0,0141
	c	0,2468				0,0332
	d	0,1451				0,0207
	e	0,1416				0,0190
51.2	a	0,2058	0,1808	0,0576	25,8%	0,0279	0,0250	0,0071	27,0%
	b	0,0996				0,0144
	c	0,2551				0,0335
	d	0,1856				0,0267
	e	0,1581				0,0227
163.8	a	0,2353	0,1834	0,0512	24,7%	0,0297	0,0230	0,0060	32,9%
	b	0,1859				0,0250
	c	0,1444				0,0181
	d	0,1202				0,0155
	e	0,2310				0,0267
524.0	a	0,1069	0,1347	0,0524	44,7%	0,0162	0,0179	0,0058	47,8%
	b	0,1476				0,0191
	c	0,2121				0,0270
	d	0,1361				0,0157
	e	0,0709				0,0114

n.a.: not applicable

Raw data – Fresh and dry weight (Definitive test #1 / Sunflower)

Treatment (mg/Kg dw)	Replicate	Fresh weight				Dry weight
		Per pot	Mean	SD	% inhib	Per pot	Mean	SD	% inhib
Control	a	9,7304	8,7924	1,1674	n.a.	0,8283	0,7872	0,1109	n.a.
	b	8,8853				0,8681
	c	10,1147				0,8990
	d	7,7664				0,6942
	e	7,4651				0,6464
1.6	a	8,8628	9,2260	1,8457	-4,9%	0,8042	0,8717	0,2100	-10,7%
	b	10,5137				1,0544
	c	11,1745				1,0866
	d	9,1884				0,8442
	e	6,3907				0,5693
5.0	a	5,7971	9,1124	2,0820	-3,6%	0,4940	0,8628	0,2262	-9,6%
	b	11,0685				1,0138
	c	10,6862				1,0768
	d	9,1088				0,8601
	e	8,9015				0,8693
16.0	a	10,2207	9,2100	0,6016	-4,7%	0,8957	0,8468	0,0731	-7,6%
	b	9,0356				0,9030
	c	9,2571				0,8767
	d	8,7266				0,8335
	e	8,8100				0,7253
51.2	a	10,3067	9,3262	1,6885	-6,1%	0,9069	0,8910	0,2294	-13,2%
	b	10,1294				1,0436
	c	10,6130				1,0696
	d	9,1003				0,9352
	e	6,4818				0,4998
163.8	a	9,2346	8,8538	2,8649	-0,7%	0,8712	0,8697	0,3029	-10,5%
	b	11,1105				1,1332
	c	9,9226				1,0308
	d	10,1299				0,9574
	e	3,8716				0,3559
524.0	a	8,1988	8,8292	1,3460	-0,4%	0,7953	0,8553	0,1512	-8,6%
	b	9,9456				1,0165
	c	10,4845				0,9941
	d	8,2927				0,8181
	e	7,2243				0,6524

n.a.: not applicable

Raw data – Fresh and dry weight (Definitive test #1 / Vetch)

Treatment (mg/Kg dw)	Replicate	Fresh weight				Dry weight
		Per pot	Mean	SD	% inhib	Per pot	Mean	SD	% inhib
Control	a		0,5960	0,3315	n.a.		0,0848	0,0482	n.a.
	b	0,9785				0,1404
	c	0,4163				0,0590
	d
	e	0,3931				0,0549
1.6	a	0,5463	0,8559	0,5023	-43,6%	0,0818	0,1259	0,0778	-48,5%
	b	1,4320				0,2172
	c	0,8307				0,1081
	d	0,2100				0,0300
	e	1,2606				0,1922
5.0	a	0,8001	1,1488	0,2278	-92,8%	0,1123	0,1617	0,0350	-90,8%
	b	1,0351				0,1404
	c	1,2713				0,1730
	d	1,3285				0,1839
	e	1,3090				0,1989
16.0	a	1,3237	0,9591	0,4735	-60,9%	0,1853	0,1361	0,0715	-60,5%
	b	1,2503				0,1730
	c	0,5472				0,0763
	d	0,3474				0,0430
	e	1,3269				0,2027
51.2	a	0,9890	1,3450	0,2633	-125,7%	0,1346	0,1789	0,0352	-111,0%
	b	1,3996				0,1854
	c	1,6985				0,2125
	d	1,2160				0,1508
	e	1,4220				0,2111
163.8	a	1,6604	1,4799	0,5972	-148,3%	0,2449	0,2094	0,0954	-147,0%
	b	0,9852				0,1292
	c	2,2388				0,3345
	d	0,7705				0,0991
	e	1,7448				0,2393
524.0	a	1,6470	1,2510	0,4487	-109,9%	0,2293	0,1700	0,0679	-100,6%
	b	0,4858				0,0545
	c	1,2790				0,1718
	d	1,3777				0,1902
	e	1,4657				0,2043

n.a.: not applicable

Empty cells: no emergence

 Raw data – Fresh and dry weight (Definitive test #2 / Sorghum)

Treatment (mg/Kg dw)	Replicate	Fresh weight				Dry weight
		Per pot	Mean	SD	% inhib	Per pot	Mean	SD	% inhib
Control	a	0,6972	0,6937	0,1029	n.a.	0,1057	0,1075	0,0177	n.a.
	b	0,5617				0,0886
	c	0,8440				0,1367
	d	0,6503				0,1045
	e	0,7153				0,1018
1.6	a	0,5442	0,7217	0,1435	-4,0%	0,0734	0,1006	0,0206	6,4%
	b	0,6652				0,0923
	c	0,8977				0,1265
	d	0,8375				0,1148
	e	0,6640				0,0958
5.0	a	0,3940	0,6825	0,2607	1,6%	0,0592	0,0964	0,0398	10,3%
	b	0,5684				0,0741
	c	0,5378				0,0711
	d	0,9065				0,1277
	e	1,0056				0,1497
16.0	a	0,8753	0,7411	0,2020	-6,8%	0,1256	0,1118	0,0291	-4,0%
	b	0,8424				0,1205
	c	0,9371				0,1486
	d	0,5672				0,0830
	e	0,4836				0,0812
51.2	a	0,8908	0,7676	0,1995	-10,7%	0,1415	0,1131	0,0325	-5,2%
	b	0,5329				0,0818
	c	1,0169				0,1492
	d	0,6057				0,0794
	e	0,7919				0,1135
163.8	a	0,6761	0,9348	0,3349	-34,8%	0,0911	0,1152	0,0513	-7,2%
	b	1,4270				0,1919
	c	0,8139				0,0833
	d	*				*
	e	0,8222				0,0945
524.0	a	0,6443	0,7140	0,1995	-2,9%	0,0912	0,0996	0,0267	7,3%
	b	0,5464				0,0727
	c	0,5584				0,0793
	d	0,7987				0,1208
	e	1,0221				0,1342

n.a.: not applicable

* no ermergence was observed in replicate d, a technical mistake was suspected: this replicate was not considered for the final calculation

Raw data – Fresh and dry weight (Definitive test #2 / Cucumber)

Treatment (mg/Kg dw)	Replicate	Fresh weight				Dry weight
		Per pot	Mean	SD	% inhib	Per pot	Mean	SD	% inhib
Control	a	3,0717	2,6109	0,5714	n.a.	0,3943	0,3716	0,0629	n.a.
	b	2,8898				0,4112
	c	2,2958				0,3594
	d	3,0413				0,4250
	e	1,7558				0,2679
1.6	a	3,7014	3,4022	0,4304	-30,3%	0,3507	0,3471	0,0550	6,6%
	b	3,9173				0,3895
	c	3,1772				0,3880
	d	2,8231				0,2545
	e	3,3921				0,3527
5.0	a	3,0615	2,9269	0,4217	-12,1%	0,2995	0,3036	0,0588	18,3%
	b	3,2292				0,3580
	c	3,0922				0,3070
	d	3,0691				0,3454
	e	2,1825				0,2083
16.0	a	2,6310	2,6629	0,2510	-2,0%	0,3397	0,3596	0,0213	3,2%
	b	2,8795				0,3717
	c	2,2723				0,3581
	d	2,8889				0,3889
	e	2,6430				0,3394
51.2	a	2,9484	2,9778	0,1282	-14,1%	0,4094	0,4530	0,0348	-21,9%
	b	3,0519				0,4593
	c	3,0026				0,4956
	d	3,1109				0,4737
	e	2,7753				0,4271
163.8	a	2,3067	3,2022	0,6183	-22,6%	0,1764	0,2620	0,0555	29,5%
	b	2,8081				0,2434
	c	3,7147				0,3204
	d	3,6513				0,2949
	e	3,5304				0,2748
524.0	a	3,1056	2,8345	0,3068	-8,6%	0,4755	0,4712	0,0385	-26,8%
	b	2,7787				0,5067
	c	2,5325				0,4060
	d	3,1980				0,4897
	e	2,5577				0,4783

n.a.: not applicable

Raw data – Fresh and dry weight (Definitive test #2 / Pea)

Treatment (mg/Kg dw)	Replicate	Fresh weight				Dry weight
		Per pot	Mean	SD	% inhib	Per pot	Mean	SD	% inhib
Control	a	3,9583	3,4882	0,3546	n.a.	0,5913	0,5343	0,0580	n.a.
	b	3,2399				0,4973
	c	3,0680				0,4655
	d	3,4774				0,5206
	e	3,6972				0,5969
1.6	a	3,5560	3,3869	1,1208	2,9%	0,4752	0,4564	0,1472	14,6%
	b	3,6764				0,4864
	c	4,1863				0,5684
	d	4,0775				0,5490
	e	1,4384				0,2028
5.0	a	2,3896	3,3682	0,6295	3,4%	0,3285	0,4660	0,0934	12,8%
	b	3,1111				0,4137
	c	3,6848				0,5129
	d	3,9673				0,5578
	e	3,6880				0,5169
16.0	a	3,7884	3,9094	0,3088	-12,1%	0,5748	0,5961	0,0482	-11,6%
	b	3,6089				0,5378
	c	3,7550				0,5874
	d	4,4057				0,6674
	e	3,9891				0,6132
51.2	a	3,5070	3,6366	0,8451	-4,3%	0,5196	0,5551	0,1393	-3,9%
	b	4,0388				0,6205
	c	4,6820				0,7371
	d	3,5765				0,5399
	e	2,3788				0,3584
163.8	a	4,2390	3,4610	0,7195	0,8%	0,5344	0,4405	0,0995	17,6%
	b	4,1201				0,5368
	c	3,1220				0,4201
	d	2,5163				0,2980
	e	3,3077				0,4131
524.0	a	3,4016	4,1041	0,4160	-17,7%	0,5408	0,5748	0,1181	-7,6%
	b	4,0613				0,3866
	c	4,4071				0,6893
	d	4,3864				0,6405
	e	4,2639				0,6170

n.a.: not applicable

 

 

 

Validity criteria fulfilled:

yes

Conclusions:

VALIDATION CRITERIA

For all these species (Triticum monococcum, Sorghum bicolor, Solanum lycopersicum, Daucus carota, Cucumis sativus, Pisum sativum), the test met the validity criteria of the test guideline detailed as follows:
- the seedling emergence was at least 70%;
- the seedlings did not exhibit visible phytotoxic effects (e.g. chlorosis, necrosis, wilting, leaf and stem deformations) and the plants did exhibit only normal variation in growth and morphology for that particular species;
- the mean survival of emerged control seedlings was at least 90% for the duration of the study;
- environmental conditions for a particular species were identical and growing media contain the same amount of soil matrix, support media, or substrate from the same source.


DISCUSSION/CONCLUSION
For all these species (Triticum monococcum, Sorghum bicolor, Solanum lycopersicum, Daucus carota, Cucumis sativus, Pisum sativum), seedling emergence and post-emergence survival were statistically not significantly reduced compared to the controls: the test item could be considered not affecting terrestrial plants seedling emergence and post-emergence survival up to the highest tested concentration ( i.e. 524 mg/kg soil dw).
Furthermore, only slight visual symptoms of chlorosis were observed in the course of this study. These effects were not considered adverse.
An application of the test item resulted in a statistically significant inhibition of shoot dry weight for only one species (Wheat, Triticum monococcum): LOEC = 163.8 mg/Kg/sdw and NOEC = 51.2 mg/Kg/sdw.

Executive summary:

A first dose response test (#1) was conducted according to OECD guideline 208 on the following species:

 

Monocotyledonae:

- Onion (Allium cepa)

- Wheat (Triticum monococcum).

 

Dicotyledonae:

- Tomato (Solanum lycopersicum)

- Carrot (Daucus carota)

- Sunflower (Helianthus annuus)

- Vetch (Vicia sativa)

 

Due to deficiencies occurred in the control groups during the definitive test (i.e.low emergence rates for onions (Allium cepa) and vetch (Vicia sativa)and low survival rates for sunflowers (Helianthus annuus)), the test validity criteria for these species were not met and thus an additional definitive test (#2) was conducted with the three following species:

 

Monocotyledonae:

- Sorghum (Sorghum bicolor)

 

Dicotyledonae:

- Cucumber (Cucumis sativus)

- Pea (Pisum sativum)

 

As the test item is a solid powder chemical which is insoluble in water, it was directly mixed with the test soils: after its incorporation into as many batches of soil as number of treatments and thorough mixing, the test soil was evenly distributed among the pots. Each treatment group consisted of a total of 30 seeds evenly distributed among 5 pots. The plant seeds were sown manually.

The plants were evaluated for adverse effects at least 3 days per week after 50% emergence of the seedlings in the control group. During this period, plants were assessed for emergence, mortality and phytotoxicity symptoms. The effects on plant shoot dry weight were determined at test termination.

For all these species (Triticum monococcum, Sorghum bicolor, Solanum lycopersicum, Daucus carota, Cucumis sativus, Pisum sativum), seedling emergence and post-emergence survival were statistically not significantly reduced compared to the controls:the test item could beconsidered not affecting terrestrial plants seedling emergence andpost-emergence survival up to the highest tested concentration ( i.e. 524 mg/kg soil dw).

Furthermore, only slight visual symptoms of chlorosis were observed in the course of this study. These effects were not considered adverse.

An application of the test item resulted in a statistically significant inhibition of shoot dry weight for only one species (Wheat,Triticum monococcum): LOEC = 163.8 mg/Kg/sdw and NOEC = 51.2 mg/Kg/sdw.

For all these species (Triticum monococcum, Sorghum bicolor, Solanum lycopersicum, Daucus carota, Cucumis sativus, Pisum sativum)

the test met the validity criteria of the test guideline detailed as follows:

- the seedling emergence was at least 70%;

- the seedlings did not exhibit visible phytotoxic effects (e.g.chlorosis, necrosis, wilting, leaf and stem deformations) and the plants did exhibit only normal variation in growth and morphology for that particular species;

- the mean survival of emerged control seedlings was at least 90% for the duration of the study;

-  environmental conditions for a particular species were identical and growing media contain the same amount of soil matrix, support media, or substrate from the same source.

Description of key information

Two definitive tests were conducted according to the OECD 208 Guideline due to deficiencies.

For all these species (Triticum monococcum, Sorghum bicolor, Solanum lycopersicum, Daucus carota, Cucumis sativus, Pisum sativum) seedling emergence and post-emergence survival were statistically not significantly reduced compared to the controls: the test item could be considered not affecting terrestrial plants seedling emergence and post-emergence survival up to the highest tested concentration ( i.e. 524 mg/kg soil dw).

Furthermore, only slight visual symptoms of chlorosis were observed in the course of this study. These effects were not considered adverse.

An application of the test item resulted in a statistically significant inhibition of shoot dry weight for only one species (Wheat,Triticum monococcum): LOEC = 163.8 mg/Kg/sdw and NOEC = 51.2 mg/Kg/sdw.

Key value for chemical safety assessment

Long-term EC10, LC10 or NOEC for terrestrial plants:

51.2 mg/kg soil dw