N-cyclohexylbenzothiazole-2-sulfenamide

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:

24 Aug 2020 - 03 Feb. 2021

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:

adopted 19 July 2006

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Details on preparation and application of test substrate:

Defined amounts of test item and quartz sand for the respective treatment (corresponding to the nominal concentrations) were finely ground with a pestle and mortar for three minutes. Thereafter the sand-test item mixture was mixed homogeneously with the test soil for 5 min (A. cepa: 3 min) using an electric stirrer equipped with a three helix stirrer rod.
The concentration of sand added to the soil was 1% (i.e. 10 g/kg soil). The control soil received sand which had not been supplemented with the test item.

Species:

other: Solanum lycopersicum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: tomato
- Plant family: Solanaceae
- Variety: Hellfrucht
- Source of seed: Enza Zaden Deutschland GmbH & Co. KG
- Lot No.: Z513050

Species:

Pisum sativum

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: pea
- Plant family: Fabaceae
- Variety: Ambrosia
- Source of seed: Demeter Mühlenbachhof
- Lot No.: 14520/19

Species:

Cucumis sativus

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: cucumber
- Plant family: Cucurbitaceae
- Variety: Marketmore
- Source of seed: Enza aden Deutschland GmbH & Co. KG
- Lot No.: 4382332

Species:

Brassica rapa

Plant group:

Dicotyledonae (dicots)

Details on test organisms:

- Common name: turnip
- Plant family: Brassicaceae
- Variety: Namenia
- Source of seed: Bioland Mühlenbachhof
- Lot No.: 252-0525-100116Z

Species:

Avena sativa

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

- Common name: oat
- Plant family: Poaceae
- Variety: Bio Sommerhafer
- Source of seed: Sativa Rheinau AG
- Lot No.: 21257

Species:

Allium cepa

Plant group:

Monocotyledonae (monocots)

Details on test organisms:

- Common name: Onion
- Plant family: Amaryllidaceae
- Variety: Stuttgarter Riesen
- Source of seed: Enza Zaden Deutschland GmbH & Co. KG
- Lot No.: 4367035

Test type:

seedling emergence and seedling growth test

Study type:

laboratory study

Substrate type:

natural soil

Limit test:

no

Total exposure duration:

15 d

Remarks:

Seeds were planted in a natural sandy loam soil immediately after test item application and left to grow under controlled conditions for either 14 days (P. sativum, S. lycopersicum), or 15 days (A. cepa, A. sativa, B. rapa, C. sativus) following 50% emerg

Test temperature:

The test plants were cultivated in an air conditioned test room.

Air temperature [°C]:

mean: 25.0 ( 23.4)
min: 19.2 (17.5)
max: 31.6 (26.7)

() = values in brackets refer to the test with A. cepa.

pH:

Properties of LUFA Soil Type 2.3: pH (0.01 M CaCl2): 6.1 +- 0.4

Moisture:

Rel. air humidity [%]:
mean: 43.5 ( 40.4)
min: 25.4 (23.2)
max: 75.9 (64.6)

() = values in brackets refer to the test with A. cepa.

Details on test conditions:

TEST SYSTEM
- Test container (type, material, size): Common plant pots made of polypropylene (diameter 11 cm, height 8.5 cm)
- Amount of soil: Each pot was filled with 470 ± 5 g (A. cepa 475 ± 5 g) soil fresh weight
- Method of seeding: the seeds were sown by pressing them into the soil with a pair of tweezers.
- No. of seeds per container: 6 (a. cepa, a. sativa) or 3 (b. rapa, c. sativus, p. sativum, s. lycopersicum)
- No. of replicates per treatment group: 4 (a. cepa, a. sativa) or 7 (b. rapa, c. sativus, p. sativum, s. lycopersicum)
- No. of replicates per control: 4 (a. cepa, a. sativa) or 7 (b. rapa, c. sativus, p. sativum, s. lycopersicum)

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographic location: LUFA soil Type 2.3 was collected from an uncultivated field at Offenbach, Rhineland-Palatinate, Germany
- Pesticide use history at the collection site: The field had neither received any organic fertilisers nor plant protection substances during the last four years before sampling
- Soil texture (if natural soil): sandy loam
- Organic carbon (%): 0.65 +- 0.08
- Maximum water holding capacity (in % dry weigth): 35.2 +- 1.8
- CEC: 6.8 +- 1.4
- Pretreatment of soil: sieved (<= 2 mm)
- Storage: ambient temperature

GROWTH CONDITIONS
- Photoperiod: 16 h
- Light source: SON-T-Agro high pressure metal halide lamps, Philips GmbH
- Light intensity [µE m-2s-1]: mean 224 min 207 max 237
- Watering regime and schedules: Soils were supplied with water and a nutrient solution as needed by bottom watering.
- Water source/type: deionised water
- Volume applied: as needed
- Interval of applications: after 0 and 7 days
- Method of application: bottom watering
- Any pest control method/fertilization (if used): “Substral®”, 7 ml/L H2O

Nominal and measured concentrations:

nominal test concentrations [mg/kg soil dw]: 0, 1, 3.2, 10, 31.6, 100, 316, 1,000
nominal test concentrations [mg/kg soil dw] for a. cepa: 0, 1, 3.2, 10, 31.6, 100

Reference substance (positive control):

no

Species:

Pisum sativum

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

>= 316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

Cucumis sativus

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/kg soil ww

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Key result

Species:

other: Solanum lycopersicum

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

31.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Key result

Species:

other: Solanum lycopersicum

Duration:

14 d

Dose descriptor:

LOEC

Effect conc.:

100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

other: Solanum lycopersicum

Duration:

14 d

Dose descriptor:

EC50

Effect conc.:

176.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

other: Solanum lycopersicum

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

>= 316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

other: Solanum lycopersicum

Duration:

14 d

Dose descriptor:

LOEC

Effect conc.:

> 316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

other: Solanum lycopersicum

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

>= 316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Species:

other: Solanum lycopersicum

Duration:

14 d

Dose descriptor:

LOEC

Effect conc.:

> 316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Key result

Species:

Pisum sativum

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

31.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Key result

Species:

Pisum sativum

Duration:

14 d

Dose descriptor:

LOEC

Effect conc.:

100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Pisum sativum

Duration:

14 d

Dose descriptor:

EC50

Effect conc.:

755.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Remarks on result:

other: disregarded/not reliable extrapolated value exceeding the highest tested test item concentration

Species:

Pisum sativum

Duration:

14 d

Dose descriptor:

EC10

Effect conc.:

12.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Remarks on result:

other: disregarded/not reliable confidence limits span more than two test concentrations

Species:

Pisum sativum

Duration:

14 d

Dose descriptor:

LOEC

Effect conc.:

> 316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

Pisum sativum

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

>= 316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Species:

Pisum sativum

Duration:

14 d

Dose descriptor:

LOEC

Effect conc.:

> 316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Species:

Cucumis sativus

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Cucumis sativus

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

316 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Cucumis sativus

Duration:

15 d

Dose descriptor:

EC50

Effect conc.:

664.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Cucumis sativus

Duration:

15 d

Dose descriptor:

EC10

Effect conc.:

183.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Cucumis sativus

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

Cucumis sativus

Duration:

15 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:

Cucumis sativus

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Key result

Species:

Brassica rapa

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

31.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Key result

Species:

Brassica rapa

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Brassica rapa

Duration:

15 d

Dose descriptor:

EC10

Effect conc.:

18.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Remarks on result:

other: disregarded/not reliable confidence limits span more than two test concentrations

Species:

Brassica rapa

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

Brassica rapa

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

Brassica rapa

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Species:

Brassica rapa

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Species:

Avena sativa

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Avena sativa

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Avena sativa

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

Avena sativa

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

Avena sativa

Duration:

15 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:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Key result

Species:

Allium cepa

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

31.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Key result

Species:

Allium cepa

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Key result

Species:

Allium cepa

Duration:

15 d

Dose descriptor:

EC50

Effect conc.:

95.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Key result

Species:

Allium cepa

Duration:

15 d

Dose descriptor:

EC10

Effect conc.:

37.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: shoot fresh weight

Species:

Allium cepa

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: survival

Species:

Allium cepa

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: Survival

Species:

Allium cepa

Duration:

15 d

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Species:

Allium cepa

Duration:

15 d

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

seedling emergence

Details on results:

Emergence, survival and shoot fresh weight of the seedlings at the end of the test were considered for statistical evaluation. Effective concentrations could not be determined for either seedling emergence or seedling survival due to the lacking of effects on these end points.
The lowest no-observed-effect concentration (NOEC) was 31.6 mg/kg soil dry weight and was observed with shoot fresh weight in A. cepa, B. rapa, P. sativum and S. lycopersicum.
The lowest 10% effect concentration (EC10) was 37.6 mg/kg soil dw and was observed with shoot fresh weight of A. cepa.
The lowest EC50 was 95.2 mg/kg soil dw and was observed in A. cepa.

Reported statistics and error estimates:

Number of emerged and number of survived seedlings as well as shoot fresh weight at the end of the test was considered for statistical data analysis. Statistical methods were applied to evaluate whether differences between test item treatments and control were significant. Arithmetic means for each pot were calculated from the single plants within a pot. Since each pot constituted a replicate, the mean values were used for statistical analysis.
Emergence and survival data were checked by the Chi2 2x2 table test with Bonferroni correction or in case of a non-monotone dose response with the Fisher's exact binominal test with Bonferroni correction to determine the NOEC and LOEC. Significance level was alpha = 0.05, one-sided greater1. The 3-Parameter normal cumulative distribution function was used to determine the effective concentrations EC10 and EC50 for the end point shoot fresh weight.
According to the criteria given in Annex 6 of the OECD guidance document No. 210, (2013) and EFSA (2019) the ECx and its 95% confidence limits were declared to be not reliable if at least one of the following criteria was met:
• The confidence limits span more than two test concentrations
• The confidence limits contain zero
• The ECx was more than 25% below the lowest test concentration or above the highest test concentration
Normal distribution and homogeneity of variances were checked by the Shapiro-wilk’s test and Levene’s test, respectively. NOEC and LOEC of shoot fresh weight were determined by applying either the Williams multiple sequential t-test (monotonous dose-response relationship). In case of inhomogeneous variances Welch t-Test was applied. Significance level was alpha = 0.05, one-sided smaller. In case the normal distribution was found to be poor, the non-parametric step-down Jonckheere-Terpstra test procedure was applied.
Statistical evaluations were performed using the ToxRat Pro software Version 3.3. (ToxRat Solutions GmbH, 2010).

Seedling Emergence and Survival:

Number of seeds introduced, number of emerged seeds, and number of survived seedlings in the untreated control pots.

Plant species	Seeds [No.]^1	Day 0 [d]^2	Emerged [No.]^3	Survived [No.]^4	Emerged [%]	Survived [%]
A. cepa	24	8	19	19	79.2	100
A. sativa	24	4	24	24	100	100
B. rapa	21	4	19	19	90.5	100
C. sativus	21	4	20	20	95.2	100
P. sativum	21	5	17	17	81.0	100
S. lycopersicum	21	5	18	18	85.7	100
(1) Total number of seeds introduced; (2) day 0 of the test given as days after sowing; (3) emerged until test end; (4) survived until test end.

 

Seedling emergence and seedling survival at the end of the test period.

Code	Test item [mg/kg soil dw]	A. cepa	A. sativa	B. rapa	C. sativus	P. sativum	S. lycopersicum
Number of seeds planted
C	0	24	24	21	21	21	21
T1	1.0	24	---	21	---	---	---
T2	3.2	24	---	21	---	21	21
T3	10.0	24	24	21	21	21	21
T4	31.6	24	24	21	21	21	21
T5	100	24	24	21	21	21	21
T6	316	---	24	---	21	21	21
T7	1,000	---	24	---	21	---	---
Number of seeds emerged (in % of planted seeds)
C	0.0	19 (79.2)	24 (100)	19 (90.5)	20 (95.2)	17 (81.0)	18 (85.7)
T1	1.0	18 (75.0)	---	18 (85.7)	---	---	---
T2	3.2	13 (54.2)	---	20 (95.2)	---	18 (95.7)	19 (90.5)
T3	10.0	14 (58.3)	23(95.8)	20 (95.2)	20 (95.2)	19 (90.5)	18 (85.7)
T4	31.6	17 (70.8)	24 (100)	20 (95.2)	17 (81.0)	19 (90.5)	19 (90.5)
T5	100	18 (75.0)	24 (100)	17 (81.0)	19 (90.5)	18 (85.7)	20 (95.2)
T6	316	--	24 (100)	---	18 (85.7)	19 (90.5)	21 (100)
T7	1,000	---	22 (91.7)	---	20 (95.2)	---	---
Number of seeds survived (in % of emerged seeds)
C	0.0	19 (100)	24 (100)	19 (100)	20 (100)	17 (100)	18 (100)
T1	1.0	17 (94.4)	---	18 (100)	---	---	---
T2	3.2	13 (100)	---	20 (100)	---	18 (100)	19 (100)
T3	10.0	14 (100)	23 (100)	20 (100)	20 (100)	17 (89.5)	18 (100)
T4	31.6	17 (100)	24 (100)	20 (100)	17 (100)	19 (100)	19 (100)
T5	100	18 (100)	24 (100)	17 (100)	19 (100)	16 (89.0)	20 (100)
T6	316	---	24 (100)	---	18 (100)	19 (100)	20 (95.2)
T7	1,000	---	21 (95.5)	---	18 (90.0)	---	---
C = control; T1 to T7 = test item treated; (---) concentration not tested

 

Statistical assessments:

Emergence, survival and shoot fresh weight of the seedlings at the end of the test were considered for statistical evaluation. An overview of the effective concentrations EC10 and EC50 as well as the Lowest-Observed-Effect-Concentration (LOEC) and No-Observed-Effect-Concentration (NOEC) is given in the table below (All data refer to nominal test item concentration [mg/kg soil dry weight]):

 

End point	A. cepa	A. sativa	B. rapa	C. sativus	P. sativum	S. lycopersicum
Emergence LOEC	>100	>1,000	>100	>1,000	>316	>316
NOEC	≥100	≥1,000	≥100	≥1,000	≥316	≥316
Survival LOEC	>100	>1,000	>100	>1,000	>316	>316
NOEC	≥100	≥1,000	≥100	≥1,000	≥316	≥316
Shoot fresh weight EC10	37.6	n.d.	(18.8)*	183.6	(12.5)*	n.d.
EC50	95.2	n.d.	n.d.	664.8	(755.5)*	176.8
LOEC	100	>1,000	100	316	100	100
NOEC	31.6	≥1,000	31.6	100	31.6	31.6
()* = not reliable (e.g. extrapolated value exceeding the highest tested test item concentration or confidence limits span more than two test concentrations); n.d.: not determined / calculation not feasible (e.g. no dose-response relationship)

 

Validity criteria fulfilled:

yes

Conclusions:

The effect of the test item N-Cyclohexyl-2-benzothiazolesulfenamide on the emergence and growth of six species of higher terrestrial plants (A. cepa, A. sativa, B. rapa, C. sativus, P. sativum, S. lycopersicum) was tested under laboratory conditions according to OECD Guideline 208 (adopted 19 July 2006).

No effects were found on either seedling emergence or seedling survival.

The lowest observed values for the end points EC10, EC50, LOEC and NOEC were found with shoot fresh weight and were as follows:

EC10 = 37.6 mg/kg dw soil (A. cepa)
EC50 = 95.2 mg/kg dw soil (A.cepa)
LOEC = 100mg/kg dw soil (A.cepa, B. rapa, P. sativum S. lycopersicum)
NOEC = 31.6 mg/kg dw soil (A. cepa, B. rapa, P. sativum, S. lycopersicum)

Executive summary:

To determine possible effects of the test item on the seedling emergence and early growth of higher terrestrial plants, a GLP compliant study according OECD Guideline 208 (adopted 19 July 2006) was performed. The effect of the test item was tested with two monocotyledonous test species, Allium cepa (onion) and Avena sativa (oat), and four dicotyledonous species, Brassica rapa (turnip), Cucumis sativus (cucumber), Pisum sativum (pea) and Solanum lycopersicum (tomato) representing six plant families.

 

The test item was ground with fine quartz sand using a pestle and mortar and subsequently mixed into the test soil at different concentrations rates (dose-response test). The spiked soil was filled into the pots of the respective treatment and the seeds were sown by pressing them into the soil with a pair of tweezers. The seeds had neither been dressed with fungicides nor imbibed with water. Control plants were grown in soil mixed with sand without the test item (control).

 

The tested concentrations were determined for each species following a non-GLP dose range finder and are summariesed below:

 

Tested item concentration [mg/kg soil dw]	A. cepa	A. sativa	B. rapa	C. sativus	P. sativum	S. lycopersicum
	1.0	10.0	1.0	10.0	3.2	3.2
	3.2	31.6	3.2	31.6	10.0	10.0
	10.0	100	10.0	100	31.6	31.6
	31.6	316	31.6	316	100	100
	100	1,000	100	1,000	316	316

 

For each species the control soil received sand which had not been supplemented with the test item.

 

There were four replicates (pots) per treatment with 6 seeds per pot for the monocotyledonous and seven replicates (pots) per treatment with 3 seeds per pot for the dicotyledonous species. Hence, the total number of seeds per species and treatment was at least 24 and 21, respectively. The number of pots and the number of seeds per pot were the same for all treatments of a given test species. Common plant pots made of polypropylene (diameter 11 cm, height 8.5 cm) were used as test pots. Each pot was filled with 470 ± 5 g (A. cepa 475 ± 5 g) soil fresh weight and placed in a separate polystyrene beaker (Kastelplast GmbH, Mainz, Germany) serving as water reservoir.

 

The pots with the reservoirs were distributed randomly in the test room and the water reservoirs were filled with deionised water as necessary to moisten the soil sufficiently. Each pot was covered loosely with a plastic lid to avoid drying up of the soil during germination of the seeds and were removed when seedlings started to emerge.
Pots were re-arranged at minimum weekly intervals.
During the test, deionised water was added to the reservoirs as needed to maintain appropriate soil moisture. Additionally, a nutrient solution (“Substral®”, 7 ml/L H2O) was added as needed judged by the observation of control plants.

 

Results:

The day on which at least 50% of the seedlings had emerged in the untreated control of the respective plant species was defined as day 0 (starting point) of the test. 

 

Seedling emergence and survival:

The number of live seedlings was determined for all species on day 7 and at the end of the test.
Emergence in the control at the end of the test was at least 79.2 %. Survival of seedlings in the controls was 100 %. Hence, the requirements defined by the test guideline regarding emergence (≥70%) and survival of seedlings (≥90%) in the control were fulfilled.

 

No statistically significant effect on either emergence or survival of seedlings was determined in any of the test species at any test item concentrations up to and including either 100 mg/kg soil dry weight (A. cepa, B. rapa) or 316 mg/kg soil dw (P. sativum, S. lycopersicum), or 1000 mg/kg soil dw (A. sativa, C. sativus).

 

Seedling shoot fresh weight:

Shoot fresh weight of each live seedling was determined at the end of the test. Shoot weight of dead seedlings was not considered.
Shoot fresh weight was statistically significantly reduced at test item concentrations of ≥ 100 mg/kg soil dw in A. cepa, B. rapa, P. sativum and S. lycopersicum. Shoot fresh weight of C. sativus was statistically significantly reduced at ≥ 360 mg/kg soil dw. No statistically significant reduction of the shoot fresh weight was observed with A. sativa at test item concentrations up to and including 1000 mg/kg soil dw

 

Statistical assessments:

Emergence, survival and shoot fresh weight of the seedlings at the end of the test were considered for statistical evaluation.

 

The lowest observed values for the end points EC10, EC50, LOEC and NOEC were found with shoot fresh weight and were as follows:

EC10 = 37.6 mg/kg soil dry weight (Allium cepa)

EC50 = 95.2 mg/kg soil dry weight (Allium cepa)

LOEC = 100 mg/kg (Allium cepa, Brassica rapa, Pisum sativum, Solanum lycopersicum)

NOEC = 31.6 mg/kg soil dry weight (Allium cepa, Brassica rapa, Pisum
sativum, Solanum lycopersicum).

Description of key information

The effect of the test item N-Cyclohexyl-2-benzothiazolesulfenamide on the emergence and growth of six species of higher terrestrial plants (A. cepa, A. sativa, B. rapa, C. sativus, P. sativum, S. lycopersicum) was tested under laboratory conditions according to OECD Guideline 208 (adopted 19 July 2006).

No effects were found on either seedling emergence or seedling survival.

The lowest observed values for the end points EC10, EC50, LOEC and NOEC were found with shoot fresh weight and were as follows:

EC10 = 37.6 mg/kg dw soil (A. cepa)
EC50 = 95.2 mg/kg dw soil (A.cepa)
LOEC = 100mg/kg dw soil (A.cepa, B. rapa, P. sativum S. lycopersicum)
NOEC = 31.6 mg/kg dw soil (A. cepa, B. rapa, P. sativum, S. lycopersicum)

Key value for chemical safety assessment

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

37.6 mg/kg soil dw