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Toxicity to terrestrial plants

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Reference
Endpoint:
toxicity to terrestrial plants: short-term (with study design considered suitable for long-term assessment)
Remarks:
Seedling Emergence and Seedling Growth Test
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016-10-21 to 2016-11-18
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)
Version / remarks:
July 2006
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Analytical monitoring:
no
Details on sampling:
- Concentrations: 1000, 333, 111, 37.0, and 12.3 mg/kg DW (factor 3). For the plant species sugar beet and carrot 4.12 mg/kg DW were tested additionally.

Vehicle:
no
Details on preparation and application of test substrate:
- Method of mixing into soil (if used): The control and test item treatments were applied once at the beginning of the study. Due to large soil amounts per concentration, several batches were prepared per concentration and applied separately. Due to low water solubility, the respective test item amounts were weighed out for each test item concentration on quartz sand (1 % of the entire soil amount per concentration) and blended thoroughly. Afterwards the test item - quartz sand mixture and an appropriate amount of demineralised water (adjusting the soil to a moisture of 50 % of the maximum water holding capacity (WHC)) were added to the soil. Subsequently, the soil was mixed carefully with a mixer (2.5 min) to ensure a homogeneous distribution.
- Controls: Untreated soil
Species:
Glycine max (G. soja)
Plant group:
Dicotyledonae (dicots)
Details on test organisms:
- Common name: soybean
- Plant family: Fabaceae
- Source of seed: Farmsaaat AG, Everswinkel, Germany
- Seed storage: The seeds were stored in the dark at the test facility at room temperature (22 ± 10 °C) and protected from moisture until use.
Species:
Lactuca sativa
Plant group:
Dicotyledonae (dicots)
Details on test organisms:
- Common name: lettuce
- Plant family: Umbelliferae
- Source of seed: Hild Samen GmbH, Marbach, Germany
- Seed storage: The seeds were stored in the dark at the test facility at room temperature (22 ± 10 °C) and protected from moisture until use.
Species:
Brassica napus
Plant group:
Dicotyledonae (dicots)
Details on test organisms:
- Common name: rape
- Plant family: Brassicaceae
- Source of seed: KWS Saat SE, Einbeck, Germany
- Seed storage: The seeds were stored in the dark at the test facility at room temperature (22 ± 10 °C) and protected from moisture until use.
Species:
Beta vulgaris
Plant group:
Dicotyledonae (dicots)
Details on test organisms:
- Common name: sugar beet
- Plant family: Amaranthaceae
- Source of seed: KWS Saat SE, Einbeck, Germany
- Seed storage: The seeds were stored in the dark at the test facility at room temperature (22 ± 10 °C) and protected from moisture until use.
Species:
Allium cepa
Plant group:
Monocotyledonae (monocots)
Details on test organisms:
- Common name: onion
- Plant family: Amaryllidaceae
- Source of seed: Hild Samen GmbH, Marbach, Germany
- Seed storage: The seeds were stored in the dark at the test facility at room temperature (22 ± 10 °C) and protected from moisture until use.
Species:
Avena sativa
Plant group:
Monocotyledonae (monocots)
Details on test organisms:
- Common name: oats
- Plant family: Poaceae
- Source of seed: KWS Lochow GmbH, Bergen, Germany
- Seed storage: The seeds were stored in the dark at the test facility at room temperature (22 ± 10 °C) and protected from moisture until use.
Test type:
seedling emergence and seedling growth test
Study type:
laboratory study
Substrate type:
natural soil
Remarks:
A 2:1 mixture of natural soil LUFA 2.2 and quartz sand was used.
Limit test:
no
Total exposure duration:
21 d
Remarks:
onion and carrot were additionally checked after 28 days
Test temperature:
nominal 22 °C ± 10 °C
actual 11.9 - 23.7 °C
pH:
soil mixture 6.27 ± 0.04
Moisture:
nominal 70 ± 25 %, actual 46.9 % to 96.2 %
Details on test conditions:
TEST SYSTEM
- Testing facility: climatic hall
- Test container (type, material, size): standard flower pots, plastic, 12 cm diameter
- Amount of soil: 0.7 kg (with 50% of MWHC) per replicate
- Method of seeding: Seeding holes were made with a seedling pistil. One seed was given into each hole and covered with soil.
- No. of seeds per container: 5 seeds/ test container (soybean: 3 seeds)
- No. of replicates per treatment group: 8 replicates/ test concentration
- No. of replicates per control: 8

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)

- Geographic location: Soil collected by LUFA, Speyer, Germany
- Pesticide use history at the collection site: No crop protection products applied during the year of sampling and 4 preceding years.
- Sampling depth (cm): approx. 20 cm
- Soil texture (if natural soil)
- % sand: 74.8% dw
- % silt: 16.6 % dw
- % clay: 8.6 % dw
- Soil classification system: loamy sand
- Composition (if artificial substrate): A 2:1 mixture of natural soil LUFA 2.2 and quartz sand was used.
- Organic carbon (%): 1.61 ±0.15 (soil mixture 0.952%)
- Maximum water holding capacity: 35.6 ± 1.4 g/100g DW

GROWTH CONDITIONS
- Photoperiod: 16 h
- Light source: cool white fluorescent tubes, special light sources for plants
- Light intensity and quality: nominal >3500 lux, actual 4473 ± 438 lux
- Watering regime and schedules: Bottom watering, including fertilizer as necessary.
- Volume applied: as needed

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : During the 21 - 28 day observation period the plants were observed on day 7, 14 and 21, onion and carrot additionally on day 28, for number of emerged seedlings, number of dead plants and visual phytotoxic effects. Observations included all variations, either inhibitory or stimulatory, between the treated replicates and the untreated controls. Such variations may be phytotoxic symptoms (e.g. chlorosis, necrosis, wilting), effects of growth and development rates. At the end of the study the shoot heights (in cm) and the shoot fresh weights of the shoots (in g) were measured additionally, directly after cutting the plants at the soil surface. The shoot fresh weight was determined for all plants from the same pot and considered as one replicate.


VEHICLE CONTROL PERFORMED: no

TEST CONCENTRATIONS
- Spacing factor for test concentrations: factor 3
- Range finding study
- Test concentrations: 10, 100, and 1000 mg/kg DW
- Results used to determine the conditions for the definitive study: The preliminary study was performed under the same environmental conditions and test duration as the definitive study. Based on the results of the preliminary study, the definitive study was conducted with the following test concentrations: 1000, 333, 111, 37.0, and 12.3 mg/kg DW (factor 3). For the plant species sugar beet and carrot 4.12 mg/kg DW were tested additionally.
Nominal and measured concentrations:
12.3, 37.0, 111, 333, and 1000 mg/kg DW (factor 3). For the plant species sugar beet and carrot 4.12 mg/kg DW were tested additionally.
Reference substance (positive control):
no
Remarks:
No reference item is recommended for this test according to the test guideline.
Species:
Avena sativa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
shoot height
Species:
Avena sativa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
shoot height
Species:
Avena sativa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
333 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot Fresh Weight
Species:
Avena sativa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
959 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Remarks on result:
other:
Remarks:
Confidence range 793- >1000 mg/kg dw
Species:
Avena sativa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Avena sativa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Allium cepa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
333 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Allium cepa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
shoot height
Species:
Allium cepa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
333 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Species:
Allium cepa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Species:
Allium cepa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Allium cepa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Beta vulgaris
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
333 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Beta vulgaris
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Beta vulgaris
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
333 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Species:
Beta vulgaris
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot Fresh weight
Species:
Beta vulgaris
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Beta vulgaris
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Brassica napus
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
333 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Brassica napus
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Brassica napus
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
111 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Species:
Brassica napus
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
698 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Remarks on result:
other: confidence range 610-777 mg/kg dw
Species:
Brassica napus
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Brassica napus
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Lactuca sativa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Lactuca sativa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Lactuca sativa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
333 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Species:
Lactuca sativa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Species:
Lactuca sativa
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Lactuca sativa
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Glycine max (G. soja)
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Glycine max (G. soja)
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Remarks:
Shoot height
Species:
Glycine max (G. soja)
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
333 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Species:
Glycine max (G. soja)
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Shoot fresh weight
Species:
Glycine max (G. soja)
Duration:
21 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Species:
Glycine max (G. soja)
Duration:
21 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
seedling emergence
Details on results:
For onion and carrot the rate of emergence of the control plants reached > 50 % within 14 days. For the other species the rate of emergence of the control plants reached > 50 % within 7 days. For the plant species rape stunned growth was observed at the highest test item concentration of 1000 mg/kg soil dry weight. For the other plant species no test item related phytotoxic effects were occurred. No test item related mortality occurred during the test period for all tested plant species up to 1000 mg/kg DW.

For shoot height statistically significant differences were observed for the plant species onion, sugar beet and rape. For these species the NOEC is 333 mg/kg soil dry weight. For the other species the NOEC is ≥ 1000 mg/kg soil dry weight. For shoot fresh weight statistically significant differences were occurred for all plant species. For the species oats, onion, sugar beet , carrot and soybean the NOEC is 333 mg/kg soil dry weight. For rape the NOEC is 111 mg/kg soil dry weight.

For the rate of emergence no statistically significant was occurred for all plant species. So, the NOEC is ≥ 1000 mg/kg soil dry weight.


Reported statistics and error estimates:
NOEC and LOEC values were calculated using One Way Analysis of Variance (ANOVA) and DUNNETT’s Method, respectively. EC-values with confidence ranges of shoot height and shoot fresh weight were calculated out of percentage inhibition, if applicable.
Validity criteria fulfilled:
yes
Remarks:
The validity criteria were fulfilled.
Conclusions:
Potential toxic effects of the test item were assessed on day 7, 14 and 21 (onion and carrot additionally day 28) by visual observations (number of emerged seedlings and phytotoxic effects) and at test end by shoot height and shoot fresh weight determination.

Description of key information

A seedling emergence and seedling growth test with the test substance on six plant species has been conducted over a period of 21 days according to guideline OECD208. The plant species raps revealed a NOEC of 111 mg/kg soil dry weight for shoot fresh weight and a corresponding EC50 of 698 mg/kg soil dry weight.

Key value for chemical safety assessment

Short-term EC50 or LC50 for terrestrial plants:
698 mg/kg soil dw
Long-term EC10, LC10 or NOEC for terrestrial plants:
111 mg/kg soil dw

Additional information

A seedling emergence and seedling growth test with the test substance has been conducted over a period of 21 days according to guideline OECD208 and in GLP compliance. The test systems comprised two monocotyledons (oats, Poaceae; onion, Amaryllidaceae), and four dicotyledons (sugar beet, Amaranthaceae; rape, Brassicaceae; carrot, Umbelliferae; soybean, Fabaceae). The test substance has been tested with five different test concentrations between 12.3 mg/kg soil dry weight (DW) and 1000 mg/kg soil dry weight (factor 3), the plant species sugar beet and carrot were tested additionally with 4.12 mg/kg DW. Test substance related visual phytotoxic effects (stunted growth) has been observed for the plant species rape at the highest tested test concentration of 1000 mg/kg DW. For the remaining plants no test item related visual phytotoxic effects has been observed up to 1000 mg/kg DW. No test item related mortality occurred during the test period for all tested plant species up to 1000 mg/kg DW. For shoot height statistically significant differences has been observed for the plant species onion, sugar beet and rape and revealed a NOEC of 333 mg/kg soil dry weight. For the other species the NOEC for shoot height is >= 1000 mg/kg soil dry weight. For shoot fresh weight statistically significant differences has been observed for all plant species and the NOEC has been determined to be 333 mg/kg soil dry weight for the species oats, onion, sugar beet, carrot and soybean. For rape the NOEC for shoot fresh weight has been shown to be 111 mg/kg soil dry weight. For the rate of emergence no statistically significant was occurred for all plant species. Therefore, the NOEC for seedling emergence is >= 1000 mg/kg soil dry weight for all tested plant species.