Reaction products of C16-18 (even numbered), C18 unsaturated alkylamines with C10-13 alkylbenzenesulfonic acid

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:

January 2020 - July 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

No deviations

Qualifier:

according to guideline

Guideline:

OECD Guideline 208 (Terrestrial Plants, Growth Test)

Version / remarks:

July 2006

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Details on sampling:

Not applicable

Vehicle:

yes

Details on preparation and application of test substrate:

- Method of application to filter paper (if used): /
- Method of mixing into soil (if used): Amines tallow alkyl dodecylbenzenesulfonates was dissolved in acetone as follows (see the table in attached justification) The solvent solutions were applied to dry soil and when the acetone had evaporated, the soil was mixed into the appropriate amount of weighed moist soil. This soil was then moistened with reverse osmosis (RO) water and mixed using a handheld mixer to ensure that the soil had a structure suitable for sowing the seeds.
- Method of application to soil surface (if used): /
- Controls: water control and solvent control. For the untreated water and solvent control groups the same amounts of dry soil and moist soil was used as for the treatment groups. For the water control the dry soil was mixed into the moist soil and for the solvent control the appropriate volume of acetone was added and allowed to evaporate off and then mixed into the moist soil.
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Concentration of vehicle in test medium (stock solution and final test solution): 0,6 - 1,8 - 5,5 - 16,6 - 50 mg a.i / mL (for more details, see the table Preparation of Test Item in attached justification)
- Evaporation of vehicle before use: yes

Species:

Allium cepa

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

- Common name: Onion
- Plant family: Liliaceae
- Variety: /

The seeds were not treated with any type of fungicides, insecticides, or any other type of pesticidal compound. They were stored dry in glass bottles and kept in darkness prior to the start of the study. Germination trials indicated that seed viability was = 75% for all species.

Species:

Lolium perenne

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

- Common name: Ryegrass
- Plant family: Poaceae
- Variety: /
The seeds were not treated with any type of fungicides, insecticides, or any other type of pesticidal compound. They were stored dry in glass bottles and kept in darkness prior to the start of the study. Germination trials indicated that seed viability was = 75% for all species.

Species:

Lycopersicon esculentum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: Tomato
- Plant family: Solanaceae
- Variety: /
The seeds were purchased from commercial suppliers. The seeds were not treated with any
type of fungicides, insecticides, or any other type of pesticidal compound. They were stored
dry in glass bottles and kept in darkness prior to the start of the study. Germination trials
indicated that seed viability was = 75% for all species.

Species:

Lactuca sativa

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: Lettuce
- Plant family: Asteraceae
- Variety: /
The seeds were purchased from commercial suppliers. The seeds were not treated with any
type of fungicides, insecticides, or any other type of pesticidal compound. They were stored
dry in glass bottles and kept in darkness prior to the start of the study. Germination trials
indicated that seed viability was = 75% for all species.

Species:

Daucus carota

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: Carrot
- Plant family: Apiaceae
- Variety: /
The seeds were purchased from commercial suppliers. The seeds were not treated with any
type of fungicides, insecticides, or any other type of pesticidal compound. They were stored
dry in glass bottles and kept in darkness prior to the start of the study. Germination trials
indicated that seed viability was = 75% for all species.

Species:

Brassica oleracea var. capitata

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: Cabbage
- Plant family: Cruciferae
- Variety: /
The seeds were purchased from commercial suppliers. The seeds were not treated with any type of fungicides, insecticides, or any other type of pesticidal compound. They were stored dry in glass bottles and kept in darkness prior to the start of the study. Germination trials
indicated that seed viability was = 75% for all species.

Test type:

seedling emergence and seedling growth test

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

21 d

Post exposure observation period:

none

Test temperature:

Temperatures were maintained at levels suitable for the individual species.
T° Min - Max : 23.2 - 26.2°C

pH:

Soil pH before treatment was 7.2.
During the test, the pH does not vary more than pH 7,2 - 7,7 (all treatments and controls related).

Moisture:

Humidity range 57.65 - 79.10 %. The humidity remain all along the test in the TG OECD range (humidity: 70 % ± 25 %)

Details on test conditions:

TEST SYSTEM
- Testing facility: Covance CRS Limited
- Test container (type, material, size): Non-porous plastic plant pots will be used. These will be labeled with at least study number, application date, plant species, test substance, rate of application and replicate number.10.5 cm internal diameter plastic pots were used.
- Amount of soil: Cabbage, carrot and lettuce : 770 g weight of dry soil. Tomato, ryegrass and onion 1100 g weight of dry soil (for more details please see the tables in attached justification).
- Method of seeding: After application the soil will be transferred into pots and the seeds of the given plant species will be planted as soon as possible. For all species 10.5 cm internal diameter plastic pots were used and each was filled to a depth of approximately 75 mm, the seeds were spaced out on the soil surface and approximately 5 mm of soil was added to cover the seeds. Each pot was labelled with the study number, species name, treatment, rate of application, replicate number, application date and the name of the Study Director. Each application rate was also identified by a colour-coded plastic stick label. After application each pot was placed in an individual water tray and placed impartially within rectangular blocks of evenly spaced pots for each plant species and randomly assigned to a position in the controlled environment area.
- No. of seeds per container: A total of 14 pots of three seeds each were sown for cabbage, carrot and lettuce, 20 pots of two seeds each for tomato and 10 pots of four seeds each for ryegrass and onion.
- No. of plants (retained after thinning): 40 for tomato, ryegrass and onion. 42 for cabbage, carrot and lettuce.
- No. of replicates per treatment group: 14 replicate / treatment for cabbage, carrot and lettuce. 20 replicate/treatment for tomato. 10 replicate/treatment for ryegrass and Onion.
- No. of replicates per control: 14 replicate / control for cabbage, carrot and lettuce. 20 replicate / control for tomato. 10 replicate/ control for ryegrass and Onion.
- No. of replicates per vehicle control: 14 replicate / solvent control for cabbage, carrot and lettuce. 20 replicate / solvent control for tomato. 10 replicate/ solvent control for ryegrass and Onion.

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographic location: /
- Pesticide use history at the collection site: /
- Collection procedures: /
- Sampling depth (cm): /
- Soil texture (if natural soil)/
- Soil taxonomic classification: /
- Soil classification system: Sandy loam
- Composition (if artificial substrate):
% sand: 70% w/w (Sand 2.00-0.063 mm)
% silt: 15% w/w (Silt 0.063-0.002 mm)
% clay: 15% w/w (Clay <0.002 mm)
For more details, please refer to the soil analysis in attached justification
- Organic carbon (%): 0.7% w/w
- Maximum water holding capacity (in % dry weigth): /
- CEC: /
- Pretreatment of soil: The carbon content (1.1% organic matter), pH (pH = 7,5) and particle size of the soil were analysed according to UK classification and it was sieved prior to use to remove all particles greater than 2 mm.
- Storage (condition, duration): not precised

NUTRIENT MEDIUM (if used)
- Description: /

GROWTH CONDITIONS
- Photoperiod: Plants were maintained in controlled environment rooms providing a continuous photoperiod of 16 hours light per day.
- Light source: Temperature, humidity and light intensity, (initially at soil level and at the top of the canopy after germination) were recorded daily throughout the exposure period. A summary of environmental data is given in the table in attached justification.
- Light intensity and quality: Light intensity, in lux, (nominally 4,000 – 15,000 lux)
- Day/night temperatures: Temperatures were maintained at levels suitable for the individual species. A summary of environmental data is given in the table in attached justification.
- Relative humidity (%): Range for all species : 57.65-79.10%. Temperature, humidity and light intensity, (initially at soil level and at the top of the canopy after germination) were recorded daily throughout the exposure period. A summary of environmental data is given in the table in attached justification.
- Wind velocity: /
- Air exchange rate: /
- Watering regime and schedules: Plants were watered as required, maintaining seedling health and growth (visually determined on a per-species and per-treatment basis).
- Water source/type: This soil was then moistened with reverse osmosis (RO) water and mixed using a handheld mixer to ensure that the soil had a structure suitable for sowing the seeds.
- Volume applied: 300 mL for cabbage, carrot and lettuce. 430 mL for tomato, ryegrass and onion.
- Interval of applications: /
- Method of application: mixing
- Any pest control method/fertilization (if used): none

ACCLIMATION PERIOD: /

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : germination, height and dry weight. Pots will be checked daily up to the stage of 50% emergence of control seedlings and thereafter at least every 2 to 3 days for additional emergence and signs of phytotoxicity.

- Phytotoxicity rating system (if used): A visual assessment based on the overall condition of surviving seedlings for each replicate was conducted at the end of the study and at regular intervals throughout. This assessment was conducted using a uniform scoring system based on plant size and vigor where 0 indicated no abnormal effects and 10 for all plants within a replicate being dead. Each pot was inspected daily and the number of emerged plants was recorded, together with any herbicidal symptoms. To see the results and the details of scoring, please refer to the table in attached justification.

VEHICLE CONTROL PERFORMED: yes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: x3
- Justification for using less concentrations than requested by guideline: none
- Range finding study: 0.1, 1, 10, 100 and 1000 mg a.i./kg dry soil
- Test concentrations: Based on the results of the range finding test, nominal treatment rates of 12.4, 37.0, 111.1, 333.3 and 1000 mg a.i./kg of dry soil were selected for a definitive dose-response test.
- Results used to determine the conditions for the definitive study:Emergence in the water control was 100% for cabbage, lettuce and tomato, 86.7% for carrot and 93.8% for onion and ryegrass. Emergence in the solvent control was 100% for lettuce and onion, 93.3% for cabbage and carrot and 93.8% for tomato and ryegrass. There was no significant reduction in emergence in any of the treatment rates compared to the solvent control, with 100% survival in all treatment rates and species. No statistically significant reductions in height in comparison to the solvent control were identified at any treatment rate. A significant reduction of 34% in dry weight was calculated for lettuce at 1000 mg a.i./kg dry soil in comparison to the solvent control. No statistically significant reductions in dry weight were identified in any other species at any treatment rate, however, for cabbage, carrot, tomato, onion and ryegrass at 1000 mg a.i./kg dry soil a reduction of 20%, 10%, 29%, 0% and 30% respectively was observed. No visible effects (e.g. phytotoxicity, wilting or leaf deformity) were observed at any treatment rate in any of the species.

Nominal and measured concentrations:

Definitive test : Based on the results of the range finding test, nominal treatment rates of 12.4, 37.0, 111.1, 333.3 and 1000 mg a.i./kg of dry soil were selected for a definitive dose-response test.

Reference substance (positive control):

no

Key result

Species:

Brassica oleracea var. capitata

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

333 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Remarks:

height

Species:

Brassica oleracea var. capitata

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Remarks:

dry weight

Species:

other: all tested species excepted cabbage

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

growth

Remarks:

height and dry weight

Species:

other: all tested species : Cabbage, Carrot, Lettuce, Tomato, Ryegrass, Onion

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

seedling emergence

Species:

other: all tested species: Cabbage, Carrot, Lettuce, Tomato, Ryegrass, Onion

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

phytotoxicity

Species:

other: all tested species : Cabbage, Carrot, Lettuce, Tomato, Ryegrass, Onion

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

germination

Details on results:

SEED GERMINATION
- Percent seed germination: Germination in the water and solvent controls was 100% for all species. Survival was 100% in all species except onion in the water control which was 97.5%. There were no treatment related effects on germination or survival in any plant species at any treatment rate.
- Root length: /
- Root discolouration/malformation: /
- Other effects: none
- Results with reference substance valid? None

SEEDLING EMERGENCE
- Percent seedling emergence: There were no effects on emergence of any of the species tested and no indications of delayed emergence for any species or treatment.
- Percent survival: 100%
- Plant height: for more details please refer to the height results in the table (attached justification).
- Dry weight: for more details please refer to the dry weight results in the table (attached justification).
- Abnormal seed development or appearance: No visible effect
- Lesions: No visible effect
- Swelling: No visible effect
- Loss of turgour: No visible effect
- Discoloration: No visible effect
- Unusual leaf/plant shape or size: No visible effect
- Other effects: none

VEGETATIVE VIGOUR : none

INJURY RATING SYSTEM: please find the "Uniform scoring and results for visual assessment of plant size and health" in attached background justification.

Results with reference substance (positive control):

none

Reported statistics and error estimates:

Statistical analysis of mean replicate height and dry weight was carried out using the SAS 9.1.3 programme (SAS Institute Inc. 2002).
The height and dry weight data was log transformed prior to analysis to stabilize the variance and the means are presented as back transformed geometric means. The test item treatment solutions were prepared in solvent; therefore, the effects of the test item were determined for all species based on statistical comparisons with the solvent control group using Williams’ tests, Dunnett’s test. The solvent and water controls were also compared for all species, using t-tests.

Major Computerized Systems:
The IT systems listed below were used on this study:
Pristima: Test item management
Rees: Environmental monitoring
SAS 9.1.3: Statistics

For more details, please refer to the statistical analysis into the result tables in attached justification.

Effect on plant growth and health:

The test item treatment solutions were prepared in solvent (acetone); therefore, the effects of the test item were determined for all species based on statistical comparisons with the solvent control group using Williams’ tests and Dunnett’s test. The solvent and water controls were also compared for all species, using t-tests. For carrot, there was a statistically significant difference in height and for lettuce in dry weight between the controls. As the test item was administered with the solvent and we are comparing the treated vs the solvent we are assessing the extra effect of the test item over the solvent which means any effects are attributable to the test item exposure only. The control comparison being significant does not interfere with the required interpretation (the control comparisons assess the impact of the solvent over water).

No significant reductions were observed in height in comparison to the solvent control in carrot, lettuce, tomato, onion and ryegrass at any treatment rate. A significant increase in height was observed in carrot at 37 mg a.i./kg and in the tomato at 12.4 and 333.3 mg a.i./kg dry soil. For cabbage, a significant reduction in height of 14% was observed at 1000 mg a.i./kg dry soil in comparison to the solvent control. No significant reductions at any treatment rate were observed in dry weight in comparison to the solvent control for cabbage, carrot, lettuce, tomato, ryegrass and onion. A significant increase in dry weight was observed in lettuce at 37.0 and 333.3 mg a.i./kg.

The No Observed Effect Concentration (NOEC) values for height compared against the solvent control were 1000 mg a.i./kg dry soil for carrot, lettuce, tomato, ryegrass and onion. For cabbage the NOEC value was 333.3 mg a.i./kg dry soil.

The No Observed Effect Concentration (NOEC) values for dry weight compared against the solvent control were 1000 mg a.i./kg dry soil all species.

Validity criteria fulfilled:

yes

Remarks:

Mean survival of emerged control seedlings was at least 90% for the duration of the study. No visible phytotoxic effects were observed in the water or solvent controls. Control seedling emergence was achieved as there was 100% germination in all species.

Conclusions:

The study is considered valid as the validation criteria were met (according TG OECD 208).

The EC50 values for germination and survival were estimated to be >1000 mg a.i./kg dry soil for all plant species. The NOEC values for effects on germination and survival were 1000 mg a.i./kg dry soil for all plant species. The EC50 values for effects on height and dry weight compared against the solvent control were >1000 mg a.i./kg dry soil for all species.

The EC25 values for effects on height and dry weight compared against the solvent control were >1000 mg a.i./kg dry soil for all species except for the height in lettuce which was 622.01 mg a.i./kg dry soil with confidence limits of 224.54 – 2649.48 mg a.i./kg dry soil. (However, the wide confidence limit indicates that there is not a strong dose response and the upper limit is substantially higher than the highest dose level in the study, overall the estimated EC25 is imprecisely estimated and therefore unreliable).

The No Observed Effect Concentration (NOEC) values for height compared against the solvent control were 1000 mg a.i./kg dry soil for carrot, lettuce, tomato, ryegrass and onion. For cabbage the NOEC value was 333.3 mg a.i./kg dry soil. The No Observed Effect Concentration (NOEC) values for dry weight compared against the solvent control were 1000 mg a.i./kg dry soil all species.

Executive summary:

 

This study was conducted to determine the phytotoxic effects of the test item on seedling emergence and growth of six non-target plant species, consisting of four dicotyledons (cabbage, carrot, lettuce and tomato) and two monocotyledons (ryegrass and onion), during early critical stages in their development. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals (2006) No. 208, " Terrestrial Plant Test: Seedling Emergence and Seedling Growth Test".

 

Following the initial range finding at 0.1, 1, 10, 100 and 1000 mg a.i./kg dry soil, the definitive test was conducted as a dose response test at nominal concentrations of 12.4, 37.0, 111.1, 333.3 and 1000 mg a.i./kg of dry soil for tomato, cabbage, carrot, lettuce, ryegrass and onion. Three seeds were sown in each of 14 pots for cabbage, carrot and lettuce, two seeds in each of 20 pots for tomato and four seeds in each of 10 pots for ryegrass and onion for each application rate of Amines tallow alkyl dodecylbenzenesulfonates and the concurrent water and solvent (acetone) controls. The soil used was a sandy loam, sieved to remove all particles greater than 2mm. After application and sowing, the pots were checked daily and the numbers of emerged seedlings were recorded. Emergence, survival and sub-lethal effects were recorded daily until at least 50% emergence of control seedlings and thereafter every one to three days. Plants were harvested 16 to 20 days after 50% emergence in the water and solvent controls, giving a total exposure period of a minimum of 21 days. The height of each individual plant was measured, and the plants were then dried before being weighed individually to obtain a measurement of dry weight.

 

Effect on emergence and plant survival in the water and solvent controls was 100% for all species. Survival was 100% in all species except onion in the water control which was 97.5%. There were no treatment related effects on emergence or survival in any plant species at any treatment rate. The EC50 values for emergence and survival were estimated to be >1000 mg a.i./kg dry soil for all plant species. The NOEC values for effects on emergence and survival were 1000 mg a.i./kg dry soil for all plant species.

 

Effect on plant growth and health

At harvest, measurement of the height and dry weight for each surviving seedling from each replicate pot was conducted. The test item treatment solutions were prepared in solvent (acetone); therefore, the effects of the test item were determined for all species based on statistical comparisons with the solvent control group using Williams’ tests and Dunnett’s test.

 

The solvent and water controls were also compared for all species, using t-tests. For carrot, there was a statistically significant difference in height and for lettuce in dry weight between the controls. As the test item was administered with the solvent and were compared with the treated vs the solvent, we assessed the extra effect of the test item over the solvent which means any effects are attributable to the test item exposure only. The control comparison being significant does not interfere with the required interpretation (the control comparisons assess the impact of the solvent over water). No significant reductions were observed in height in comparison to the solvent control in carrot, lettuce, tomato, onion and ryegrass at any treatment rate. A significant increase in height was observed in carrot at 37 mg a.i./kg and in the tomato at 12.4 and 333.3 mg a.i./kg dry soil. For cabbage, a significant reduction in height of 14% was observed at 1000 mg a.i./kg dry soil in comparison to the solvent control.

 

No significant reductions at any treatment rate were observed in dry weight in comparison to the solvent control for cabbage, carrot, lettuce, tomato, ryegrass and onion. A significant increase in dry weight was observed in lettuce at 37.0 and 333.3 mg a.i./kg.

No visible effects (e.g. phytotoxicity, wilting or leaf deformity) were observed at any treatment rate in any of the species.

 

The EC50 values for effects on height and dry weight compared against the solvent control were >1000 mg a.i./kg dry soil for all species. The EC25 values for effects on height and dry weight compared against the solvent control were >1000 mg a.i./kg dry soil for all species except for the height in lettuce which was 622.01 mg a.i./kg dry soil with confidence limits of 224.54 – 2649.48 mg a.i./kg dry soil (However, the wide confidence limit indicates that there is not a strong dose response and the upper limit is substantially higher than the highest dose level in the study, overall the estimated EC25 is imprecisely estimated and therefore unreliable).

 

The No Observed Effect Concentration (NOEC) values for height compared against the solvent control were 1000 mg a.i./kg dry soil for carrot, lettuce, tomato, ryegrass and onion. For cabbage the NOEC value was 333.3 mg a.i./kg dry soil. The No Observed Effect Concentration (NOEC) values for dry weight compared against the solvent control were 1000 mg a.i./kg dry soil all species.

 

The study was considered valid as the validation criteria were met and no visible phytotoxic effects were observed in the controls.

Description of key information

According the OECD 208 and GLP study, the chronic toxicity of the registered substance (EC 701-164-2) on terrestrial plants is considered as a worst case value at NOEC = 333 mg a.i/kg dry soil (observed on Cabbage specie after 21 days of exposure).

Key value for chemical safety assessment

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

333 mg/kg soil dw

Additional information

Toxicity to terrestrial plants (OECD 208, GLP):

This study was conducted to determine the phytotoxic effects of the test item (EC 701-164-2), on seedling emergence and growth of six non-target plant species, consisting of four dicotyledons (cabbage, carrot, lettuce and tomato) and two monocotyledons (ryegrass and onion), during early critical stages in their development. The study was considered valid as the validation criteria were met according TG OECD 208 and no visible phytotoxic effects were observed in the controls. The soil used was a sandy loam, sieved to remove all particles greater than 2 mm.

 

The EC50 values for emergence and survival were estimated to be >1000 mg a.i./kg dry soil for all plant species. The NOEC values for effects on emergence and survival were 1000 mg a.i./kg dry soil for all plant species.

The No Observed Effect Concentration (NOEC) values for height compared against the solvent control were 1000 mg a.i./kg dry soil for carrot, lettuce, tomato, ryegrass and onion. For cabbage the NOEC value was 333.3 mg a.i./kg dry soil.

The No Observed Effect Concentration (NOEC) values for dry weight compared against the solvent control were 1000 mg a.i./kg dry soil all species.