1,4-Benzenedicarboxylic acid, 1,4-diisononyl ester

Toxicity to terrestrial plants

Administrative data

Endpoint:

toxicity to terrestrial plants: short-term

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2010-02-10 to 2011-03-17

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Report date:

2011

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

(2006)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

1,4-Benzenedicarboxylic acid, 1,4-diisononyl ester

EC Number:

700-453-0

Cas Number:

59802-05-0

Molecular formula:

C26H42O4

IUPAC Name:

1,4-Benzenedicarboxylic acid, 1,4-diisononyl ester

Test material form:

liquid

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Based on the information from the range finding test, a single test concentration of four replicates, of 1000 mg/kg was tested for each species in the definitive test. The test item concentration in the test samples was determined by high performance liquid chromatography (HPLC).
- Sampling method: Samples were taken from the acetone solvent and the 25 g/250 mL solvent stock solution on Day 0 for quantitative analysis.
- Sample storage conditions before analysis: Duplicate samples were taken and stored at approximately -20°C for further analysis if necessary.

Test substrate

Vehicle:

yes

Remarks:

acetone

Details on preparation and application of test substrate:

- Method of mixing into soil (if used): An amount of test item (25 g) was dissolved in acetone and the volume adjusted to 250 mL to give solvent stock solution of 25 g / 250 mL. The solvent stock solution was inverted several times to ensure adequate mixing and homogeneity. An aliquot (51.0 mL)
of acetone and the 25 g/250 mL solvent stock solution was separately added to approximately 200 g of sand, in duplicate, and the solvent allowed to evaporate off prior to incorporation into 4.7 kg sand and 5.1 kg (wet weight) soil using a mixer to give the solvent control and the test concentration of 1000 mg/kg.
- Controls: control without the addition of solvent or test item and vehicle control
- Chemical name of vehicle: acetone
- Concentration of vehicle in test medium: An amount of test item (25 g) was dissolved in acetone and the volume adjusted to 250 mL to give solvent stock solution of 25 g / 250 mL.
- Evaporation of vehicle before use: The solvent was allowed to evaporate off prior to incorporation into sand and soil using a mixer.

Test organisms

1

Species:

Glycine max (G. soja)

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: soybean (Glycine max)
- Variety: amsoy
- Source of seed: Herbiseed, Twyford, UK
- Prior seed treatment/sterilization: supplied untreated
- Seed storage: Upon arrival the seeds were stored under cool, dry conditions.

2

Species:

Lycopersicon esculentum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: tomato (Lycopersicon esulentum)
- Variety: Orange Pixie
- Source of seed: Herbiseed, Twyford, UK
- Prior seed treatment/sterilization: supplied untreated
- Seed storage: Upon arrival the seeds were stored under cool, dry conditions.

3

Species:

Avena sativa

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

- Common name: oat (Avena sativa)
- Variety: S W Argyle
- Source of seed: Senova Ltd., Cambridge, UK
- Prior seed treatment/sterilization: supplied untreated
- Seed storage: Upon arrival the seeds were stored under cool, dry conditions.

Study design

Test type:

seedling emergence and seedling growth test

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

yes

Total exposure duration:

21 d

Test conditions

Test temperature:

The temperature was maintained at 9 °C to 26 °C with a photoperiod of approximately 16 hours light and 8 hours darkness.

Moisture:

relative humidity at 39% to 77%

Details on test conditions:

TEST SYSTEM
- Test facility: greenhouse equipped with artificial lighting and heating
- Test container (type, material, size): Oolypropylene pots with a diameter of 13 cm
- Amount of soil: 850 g (dry weight) substrate
- No. of seeds per container: At the start of the test five seeds were placed in each test and control pot at random.
- No. of replicates per treatment group: 4 for each species
- No. of replicates per control: 4 for each species
- No. of replicates per vehicle control: 4 for each species

SOURCE AND PROPERTIES OF SUBSTRATE
- Source: Sporting Surface Supplies Ltd, Reigate, Surrey, UK
- Soil composition (if artificial substrate): The soil was a sandy loam with 3.8% organic matter content and a pH of 8.0. The organic matter content of the soil was in excess of 3%. Therefore the soil was mixed 50:50 w/w with sand prior to use to give a nominal organic carbon content of 0.87% (equivalent to 1.5% organic matter).
- Organic carbon: 0.87%
- Pretreatment of soil: The organic matter content of the soil was in excess of 3%. Therefore the soil was mixed 50:50 w/w with sand prior to use to give a nominal organic carbon content of 0.87% (equivalent to 1.5% organic matter).

GROWTH CONDITIONS
- Photoperiod: 16 hours light, 8 hours darkness
- Day/night temperatures: 9 °C to 26 °C
- Relative humidity (%): 39% to 77%
- Watering regime and schedules: Following sowing, the pots were surface irrigated to encourage germination and then placed in a randomised fashion in a greenhouse. the initial watering the pots were watered as necessary using sub-irrigation.
- Method of application: sub-irrigation

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
All the pots were monitored daily after sowing for emergence and the date when 50% emergence occurred in the solvent control pots for each species was recorded. For 21 days following 50% emergence in the solvent control pots, the number of seedlings emerged and any mortalities and/or morphological abnormalities were determined daily in the test and control pots for each species. On Day 21 the above ground portion of each seedling was harvested and the dry weight determined by placing in pre-weighed containers and drying in an oven at approximately 60°C.

VEHICLE CONTROL PERFORMED: yes

TEST CONCENTRATIONS
- Spacing factor for test concentrations: limit test 1000 mg/kg test item for each species
- Justification for using less concentrations than requested by guideline: Based on the results of the range-finding test a "Limit test" was conducted at a concentration of 1000 mg/kg to confirm that at the maximum concentration given in the OECD Test Guidelines no effect on seedling emergence or
morphological abnormalities were observed.
- Range finding study
- Test concentrations: 0, 10, 100 and 1000 mg/kg soil dry weight
- Results used to determine the conditions for the definitive study: There were no significant differences (P≥0.05) between the solvent control and all the test groups in terms of emergence for each species. There were no significant decreases in shoot dry weight (P≥0.05) between the solvent
control and all the test groups at test termination for each species.

Nominal and measured concentrations:

1000 mg/kg nominal concentration

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

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

Effect concentrations 2

Key result

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

Effect concentrations 3

Key result

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

Effect concentrations 4

Key result

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

Effect concentrations 5

Key result

Species:

Lycopersicon esculentum

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

Effect concentrations 6

Key result

Species:

Lycopersicon esculentum

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

Effect concentrations 7

Key result

Species:

Lycopersicon esculentum

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

Effect concentrations 8

Key result

Species:

Lycopersicon esculentum

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

Effect concentrations 9

Key result

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

Effect concentrations 10

Key result

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

Effect concentrations 11

Key result

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

Effect concentrations 12

Key result

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

Details on results:

Emergence
Soybean (Glycine max), tomato (Lycoperisicon esculentum) and oat (Avena sativa) reached 50% emergence in the solvent control on Days 14, 20 and 5 respectively after sowing. Statistical analysis using Bartlett's test for homogeneity of variance (Sokal and Rohlf 1981) and a Students t-test was carried out on the emergence data for each species.
The No Observed Effect Concentration for soybean, tomato and oat was 1000 mg/kg. The No Observed Effect Concentration is based upon no significant differences (P0.05) between the test concentrations and the solvent control in terms of seedling emergence.

Morphological Effects
No morphological abnormalities were observed for each species throughout the test.

Shoot Dry Weight Data
Statistical analysis using Bartlett's test for homogeneity of variance (Sokal and Rohlf 1981) and a Students t-test was carried out on the shoot dry weight data for each species. There were no significant differences (P≥0.05) between the solvent control and 1000 mg/kg test group in terms of shoot dry weight at test termination for all three species.
The shoot weights for the control for tomato were found to be significantly greater (P≥0.05) compared to the solvent control and 1000 mg/kg test group, however as there were no significant differences in terms of dry weight between the solvent control and 1000 mg/kg test group, and the direct comparison was between the solvent control and 1000 mg/kg test concentration, it was considered that these differences did not affect the outcome or validity of the test. Review of the data suggests that these differences were likely due to a small number of particularly large plants in the control group.

The No Observed Effect Concentration for soybean, tomato and oat was 1000 mg/kg. The No Observed Effect Concentration is based upon no significant differences (P≥0.05) between the test concentrations and the solvent control in terms of final shoot dry weight.

Analysis of the 25 g/250 ml solvent stock solution used to prepare the test concentrations on Day 0 showed a measured test concentration of 100% of nominal value and so the results are based on nominal test concentrations only

Reported statistics and error estimates:

Statistical analysis using Bartlett's test for homogeneity of variance (Sokal and Rohlf 1981) and a Students t-test was carried out on the emergence data and on shoot dry weight data for each species.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The No Observed Effect Concentration for soybean, tomato and oat was 1000 mg/kg.
The No Observed Effect Concentration is based upon no significant differences (P >=0.05) between the test concentrations and the solvent control in terms of seedling emergence.

Executive summary:

Introduction. A study was performed to assess the effects of the test item on the emergence and growth of three plant species. The method followed was designed to be compatible with the OECD Guidelines for Testing of Chemicals: Guideline 208: Terrestrial (Non-target) Plant Test: Seedling Emergence and Seedling Growth Test (July 2006).

Methods. Following a preliminary range-finding test, three plant species; two dicotylendonous species, soybean (Glycine max) and tomato (Lycopersicon esulentum) and one monocotylendonous species, oat (Avena sativa) were exposed to a single concentration of 1000 mg/kg. The number of seedlings emerged and any mortalities and/or morphological abnormalities were determined daily for 21 days after 50% emergence in the solvent control for each species.

Results. The EC50 (emergence) and EC50 (growth based on final dry weight) for the test item based on nominal test concentrations for the three species tested were as follows:

Species	EC50 (emergence) (mg/kg)	No Observed Effect Concentration (mg/kg)	EC50 (growth) (mg/kg)	No Observed Effect Concentration (mg/kg)*
Soybean	>1000	1000	>1000	1000
Tomato	>1000	1000	>1000	1000
Oat	>1000	1000	>1000	1000

Analysis of the solvent stock solution used to prepare the test concentrations on Day 0 showed a measured test concentration of 100% of nominal value and so the results are based on nominal test concentrations only.

* No Observed Effect Concentration (growth) based on the concentration where no significant decrease was observed for dry weight compared to the solvent control and no morphological abnormalities were observed.