2-(2,6-diethyl-4-methyl-phenyl)propanediamide

Toxicity to terrestrial plants

Link to relevant study record(s)

Reference

Endpoint:

toxicity to terrestrial plants: short-term

Remarks:

The study on toxicity to terrestrial plants was conducted solely to comply with a non-EU national registration requirement, and has been provided here in accordance with REACH, Article 22(1)e.

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The study was conducted under a National quality assurance scheme (HJ/T 155-2004) similar to OECD GLP

Qualifier:

according to guideline

Guideline:

other: SEPA HJ/T 153-2004 The guidelines for the testing of chemicals [S]. Beijing: SEPA, 2004

Version / remarks:

2004

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: CRC-MEP The Guidelines for the Testing of Chemicals-Effects on biotic systems [M]. 2nd edition Beijing: China Environmental Science Press 2013, 459 - 463

Version / remarks:

2013

Deviations:

no

GLP compliance:

no

Remarks:

The study was conducted under a National quality assurance scheme (HJ/T 155-2004) similar to OECD GLP

Specific details on test material used for the study:

SOURCE OF TEST MATERIAL
- Expiration date of the lot/batch: 31 May 2018
- Purity test date: 13 July 2015

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: < 30 °C

Analytical monitoring:

yes

Details on sampling:

- Concentrations: Control and 100, 500 and 1000 mg/L test solutions
- Sampling method: A sample was taken from an additional treatment at the start of the test, dissolved in acetonitrile and filtered by 0.22 µm membrane.
- Sample storage conditions before analysis: Not reported

Vehicle:

no

Details on preparation and application of test substrate:

- Method of mixing into soil: The appropriate test solution and deionised water were added to 10 g quartz sand and then 90 g quartz sand was thoroughly mixed.
- Controls: Blank control

Species:

Raphanus sativus

Details on test organisms:

- Common name: Radish
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

other: Capsicum annuum

Details on test organisms:

- Common name: Pepper
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

Lycopersicon esculentum

Details on test organisms:

- Common name: Tomato
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

other: Sesamum indicum

Details on test organisms:

- Common name: Sesame
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

Zea mays

Details on test organisms:

- Common name: Corn
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

Cucumis sativus

Details on test organisms:

- Common name: Cucumber
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

other: Brassica oleracea

Details on test organisms:

- Common name: Kohlrabi
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

Oryza sativa

Details on test organisms:

- Common name: Rice
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

Lactuca sativa

Details on test organisms:

- Common name: Lettuce
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Species:

other: Brassica chinensis L.

Details on test organisms:

- Common name: Cabbage
- Prior seed treatment/sterilization: Only untreated seed (not treated with fungicides, repellents, etc.) taken from the same lot and year or season of collection was used in the test
- Seed storage: Seeds were held at 5 °C in moisture-proof containers with a seed moisture content of less than 10 %

Test type:

seed germination/root elongation toxicity test

Study type:

laboratory study

Substrate type:

other: Quartz sand

Limit test:

no

Total exposure duration:

6 d

Post exposure observation period:

None

Test temperature:

25.0 - 25.1 °C

pH:

6.85 - 7.03

Moisture:

80 - 81 % humidity

Details on test conditions:

TEST SYSTEM
- Testing facility: Climatic chamber
- Test container (type, material, size): Glass dishes sealed with preservative film containing holes
- Amount of soil: 100 g
- Method of seeding: The seed was positioned on the substrate allowing adequate room for anticipated growth. The radicle end of the seed was aligned in the direction of growth.
- No. of seeds per container: 15
- No. of replicates per treatment group: 1
- No. of replicates per control: 1

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- % sand: 100 % quartz sand

GROWTH CONDITIONS
- Photoperiod: Complete darkness
- Relative humidity (%): 80 ± 5 %

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): Number of germinated seeds were counted and root lengths were measured at the end of the study. The test period ended when at least 65 % of the control seed had germinated and developed roots as least 20 mm long.

Temperature in the germination facility was reported at the start and end of the test. Test solution pH was recorded at the start of the test.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2 - 5
- Justification for using less concentrations than requested by guideline: Range-finding study conducted only
- Range finding study: Yes
- Test concentrations: 100, 500 and 1000 mg/L
- Results used to determine the conditions for the definitive study: No significant phytotoxicity was observed in the range-finding test, therefore no definitive testing was carried out

Nominal and measured concentrations:

Nominal concentrations: 100, 500 and 1000 mg/L
Measured concentrations: 100, 519 and 1012 mg/L

Reference substance (positive control):

no

Key result

Species:

other: Brassica chinensis L.

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

Lactuca sativa

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

Oryza sativa

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

other: Brassica oleracea

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

Cucumis sativus

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

Zea mays

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

other: Sesamum indicum

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

Lycopersicon esculentum

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

other: Capsicum annuum

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

Raphanus sativus

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

germination

Key result

Species:

other: Brassica chinensis L.

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

Lactuca sativa

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

Oryza sativa

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

other: Brassica oleracea

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

Cucumis sativus

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

Zea mays

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

other: Sesamum indicum

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

Lycopersicon esculentum

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

other: Capsicum annuum

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Key result

Species:

Raphanus sativus

Duration:

6 d

Dose descriptor:

NOEC

Effect conc.:

1 012 other: mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

other: root elongation

Details on results:

SEED GERMINATION
- Percent seed germination: At the end of the test, over 65 % of the control seeds germinated
- Root length: At the end of the test, control roots were longer than 20 mm
- EC50 values were >1012 mg/L (measured, initial) for all species

SEEDLING EMERGENCE
- Percent seedling emergence: The germination rates of the highest concentration group (1000 mg/L) were < 50 % compared to the blank control
- EC50 values were >1012 mg/L (measured, initial) for all species

Reported statistics and error estimates:

The NOEC (no observed effect concentration) and LOEC (lowest observed effect concentration) for root elongation was determined by comparing the mean growth rate for each concenration with the control mean using a multiple comparison method of Dunnett's tests. The NOEC and LOEC for seed germination was determined using Chi-square Test. The EC50 was not determined as no toxicity was observed.

Table 1. Germination rates for ten seeds

Initial measured concentration (mg/L)	Control		100		519		1012
	Germination rate (%)	Inhibition rate (%)	Germination rate (%)	Inhibition rate (%)	Geramination rate (%)	Inhibition rate (%)	Germination rate (%)	Inhibition rate (%)
Brassica chinensis L.	100	-	93.3	6.67	93.3	6.67	100	0.00
Lactuca sativa	100	-	93.3	6.67	93.3	6.67	100	0.00
Oryza sativa L.	100	-	93.3	6.67	93.3	6.67	93.3	6.67
Brassica oleracea	93.3	-	93.3	0.00	93.3	0.00	93.3	0.00
Cucumis sativus	93.3	-	100	-7.14	93.3	0.00	100	-7.14
Zea mays	100	-	93.3	6.67	100	0.00	93.3	6.67
Sesamum indicum	93.3	-	100	-7.14	100	-7.14	93.3	0.00
Lycopersicon esculentum	93.3	-	100	-7.14	86.7	7.14	100	-7.14
Capsicum annuum	100	-	93.3	6.67	100.0	0.00	93.3	6.67
Raphanus sativus	93.3	-	93.3	0.00	100	-7.14	100	-7.14

Table 2. Root lengths for ten seeds

Initial measured concentration (mg/L)	Control			100			519			1012
	Mean (mm)	RSD (%)	Inhibition rate (%)	Mean (mm)	RSD (%)	Inhibition rate (%)	Mean (mm)	RSD (%)	Inhibition rate (%)	Mean (mm)	RSD (%)	Inhibition rate (%)
Lactuca sativa	25.2	10.6	-	23.6	12.0	6.08	22.8	19.2	9.23	22.6	18.0	10.2
Oryza sativa L.	50.4	6.84	-	50.7	7.10	-0.56	50.0	10.27	0.80	49.0	7.02	2.70
Brassica oleracea	34.1	12.7	-	34.1	11.7	-0.13	31.3	17.4	8.16	30.0	11.9	11.8
Cucumis sativus	50.7	8.33	-	52.2	8.19	-3.02	49.8	8.03	1.75	49.5	5.46	2.31
Zea mays	56.2	13.3	-	57.5	12.7	-2.22	58.8	9.97	-4.49	55.9	11.6	0.57
Sesamum indicum	26.5	11.6	-	26.8	18.0	-1.27	25.2	15.9	4.78	24.0	14.2	9.26
Lycopersicon esculentum	29.5	13.2	-	29.5	15.7	0.11	29.3	13.8	0.50	26.2	9.0	11.1
Capsicum annuum	25.2	6.89	-	23.6	12.0	6.65	23.2	15.7	8.11	22.7	16.4	9.96
Raphanus sativus	30.1	15.5	-	28.9	12.8	3.92	28.9	12.6	3.99	28.2	9.90	6.27
Brassica chinensis L.	32.9	14.2	-	31.3	14.2	4.91	30.1	15.2	8.71	29.9	13.2	9.11

 

Table 3. 6-day LOEC and EC50 values for ten plant species

Seed	Germination		Root elongation
	LOEC (mg/L)	EC50 (mg/L)	LOEC (mg/L)	EC50 (mg/L)
Brassica chinensis L.	> 1012	> 1012	> 1012	> 1012
Lactuca sativa	> 1012	> 1012	> 1012	> 1012
Oryza sativa L.	> 1012	> 1012	> 1012	> 1012
Brassica oleracea	> 10120	> 1012	> 1012	> 1012
Cucumis sativus	> 1012	> 1012	> 1012	> 1012
Zea mays	> 1012	> 1012	> 1012	> 1012
Sesamum indicum	> 1012	> 1012	> 1012	> 1012
Lycopersicon esculentum	> 1012	> 1012	> 1012	> 1012
Capsicum annuum	> 1012	> 1012	> 1012	> 1012
Raphanus sativus	> 1012	> 1012	> 1012	> 1012

Initial measured concentrations are reported, based on test material

Validity criteria fulfilled:

yes

Remarks:

> 65 % of the control seed germinated at the end of the test

Conclusions:

Based on initial measured concentration, the NOEC for seed germination and root elongation of the test material to Brassica chinensis L. (cabbage), Lactuca sativa (lettuce), Oryza sativa L. (rice), Brassica oleracea (kohlrabi), Cucumis sativus (cucumber), Zea mays (corn), Sesamum indicum (sesame), Lycopersicon esculentum (tomato), Capsicum annuum (pepper) and Raphanus sativus (radish) was 1012 mg/L (nominal concentration: 1000 mg/L), the highest concentration tested.

Executive summary:

The effects of the test material on seed germination and root elongation of cabbage (Brassica chinensis L.), lettuce (Lactuca sativa), rice (Oryza sativa L), kohlrabi (Brassica oleracea), cucumber (Cucumis sativus), corn (Zea mays), sesame (Sesamum indicum), tomato (Lycopersicon esculentum), pepper (Capsicum annuum) and radish (Raphanus sativus) was determined in a 6-day range-finding test according to SEPA HJ/T 153-2004 and under GLP. Seedlings were exposed to initial measured concentrations of 100, 519 and 1012 mg/L, alongside a blank control. Each replicate contained 15 seeds and only one replicate was tested for each treatment. Based on initial measured concentration, the NOEC for seed germination and root elongation of the test material was 1012 mg/L for all plant species tested (nominal concetration: 1000 mg/L). Based on initial measured concentration, the LOEC and EC50 were >1012 mg/L (nominal concentration: 1000 mg/L).

Description of key information

6-day NOEC = 1012 mg/L, seed germination and root elongation, 10 plant species, SEPA HJ/T 153 -2004, Zhen 2016

Additional information

The effects of the test material on seed germination and root elongation of cabbage (Brassica chinensis L.), lettuce (Lactuca sativa), rice (Oryza sativa L), kohlrabi (Brassica oleracea), cucumber (Cucumis sativus), corn (Zea mays), sesame (Sesamum indicum), tomato (Lycopersicon esculentum), pepper (Capsicum annuum) and radish (Raphanus sativus) was determined in a 6-day range-finding test according to SEPA HJ/T 153-2004 and under GLP. Seedlings were exposed to initial measured concentrations of 100, 519 and 1012 mg/L, alongside a blank control. Each replicate contained 15 seeds and only one replicate was tested for each treatment. Based on initial measured concentration, the NOEC for seed germination and root elongation of the test material was 1012 mg/L for all plant species tested (nominal concentration: 1000 mg/L). Based on initial measured concentration, the LOEC and EC50 were >1012 mg/L (nominal concentration: 1000 mg/L).