2-{2-chloro-4-mesyl-3-[(tetrahydrofuran-2-ylmethoxy)methyl]benzoyl}cyclohexane-1,3-dione

Toxicity to soil microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to soil microorganisms

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27 Jul - 24 Aug 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 217 (Soil Microorganisms: Carbon Transformation Test)

Version / remarks:

adopted January 21, 2000

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Sächsisches Staatsministerium für Umwelt und Landwirtschaft, Dresden, Germany

Analytical monitoring:

no

Vehicle:

no

Details on preparation and application of test substrate:

AMENDMENT OF SOIL
- Type of organic substrate: glucose concentration was determined as 0.6%

APPLICATION OF TEST SUBSTANCE TO SOIL
- Method: For this purpose soil samples (100 g soil d.w.) were taken, mixed with glucose by means of a hand-stirrer and filled into reaction flasks (500 mL). Afterwards, small vessels containing 18 mL 1 M NaOH solution were placed in the reaction flasks, which were tightly closed and then connected with a respirometer

VEHICLE: no vehicle was used

Test organisms (inoculum):

soil

Total exposure duration:

28 d

Test temperature:

19.7 - 20.6 °C in a climatic room

Moisture:

water holding capacity: 32.46 g/100 g soil d.w.
Water content of soil: 15.42 - 16.14 g/100 g soil d.w. (equivalent to 47.49 - 49.72% of WHC)

Details on test conditions:

TEST SYSTEM
- Testing facility: BioChem agrar; Labor für biologische und chemische Analytik GmbH, Gerichshain, Germany
- Test container: 4 L steel vessel, used lids allowed air exchange
- Amount of soil: 1000 g soil dry weight (= one sub-sample) per vessel
- No. of replicates per concentration: 3
- No. of replicates per control: 3
- No. of replicates per vehicle control: no vehicle was used

SOIL INCUBATION
- Method: incubation of the soil samples was performed as series of individual and equally sized sub-samples (1000 g soil dry weight) of each treatment group, Temperature: 19.7 - 20.6 °C in a climatic room; incubated in the dark; Water content: 12.98 - 16.23 g water/100 g soil d.w. (equivalent to 40 – 50% of WHC); duration: 28 d

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographical reference of sampling site (latitude, longitude): Wassergut Canitz, Germany; GPS-position 12.694435960 degrees East, 51.403774567 degrees North
- History of site: Cultivation at soil removal (2012) and pre-cultivation (2011) fallow ground
- Treatments with fertilizers: none since 2003
- Treatments with plant protection products: last 1990
- Depth of sampling: 20 cm

Soil texture
- % sand: 52.4 (according to USDA), 50.7 (according to DIN 11277)
- % silt: 38.2 (according to USDA), 39.9 (according to DIN 11277)
- % clay: 9.4 (according to USDA and DIN 11277)
- Soil taxonomic classification: sandy loam (according to USDA) and loamy sand (according to DIN 4220)
- Soil classification system: USDA and DIN 4220
- pH (in water): 6.2
- Initial nitrate concentration for nitrogen transformation test (mg nitrate/kg dry weight): 0.44 mg/100 g soil d.w.
- Maximum water holding capacity (in % dry weigth): 32.46 g/100 g soil d.w.
- Cation exchange capacity: 8.8 cmol+/kg soil
- Pretreatment of soil: The soil was removed to a depth of 20 cm as mixed sample, afterwards dried at room temperature and passed through a 2 mm mesh sieve (18.05 – 22.05.2012). Before application, the soil was adapted to test conditions (conditioning phase: 18.07. - 27.07.2012)
- Storage: at approx. 4 °C in containers under aerobic conditions in the dark, 22.05. - 18.07.2012
- Initial microbial biomass as % of total organic C: 1.93% of Corg Carbon content of microbial biomass (corresponds to 27.26 mg C/100 g soil d.w.)

DETAILS OF PREINCUBATION OF SOIL: no additional information

EFFECT PARAMETERS MEASURED: The carbon transformation was determined over a measurement period of 12 hours on sampling days 0 (3 hours after application), 7, 14 and 28 days after application.

VEHICLE CONTROL PERFORMED: no vehicle was used

Nominal and measured concentrations:

nominal concentrations: control (quartz meal only), 0.40 mg test item/kg dry weight soil (0.3 kg test item / 750,000 kg soil dry weight per ha) and 4.00 mg test item/kg dry weight soil (3 kg test item / 750,000 kg soil dry weight per ha)

Reference substance (positive control):

yes

Remarks:

Dinoterb

Duration:

28 d

Dose descriptor:

other: trigger value of 25% inhibition compared with the control

Effect conc.:

> 4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

test item = act. ingr.

Basis for effect:

respiration rate

Details on results:

- Effect concentrations exceeding solubility of substance in test medium: No adverse effects of the test item on carbon transformation in soil were observed at either tested concentrations (0.40 mg/kg dry soil and 4.00 mg/kg dry soil) after 28 days.
- Other: Differences from the control of -1.2% (test concentration 0.40 mg/kg dry soil) and -1.5% (test concentration 4.00 mg/kg dry soil) were measured at the end of the 28-day incubation period.

Results with reference substance (positive control):

- Results with reference substance valid? yes
- Relevant effect levels: reference item Dinoterb caused an inhibition of carbon transformation of -28.1%, -44.5% and -46.3% at 6.80 mg, 16.00 mg and 27.00 mg Dinoterb per kg soil dry weight, respectively, 28 days after application.

Reported statistics and error estimates:

Statistical evaluation of the test results (2-sided Student-t-test for homogeneous variances at 5% significance level and 2-sided Welch-t-test for inhomogeneous variances at 5% significance leve) was performed.

The NOEC was determined to be at 4.00 mg test item/kg soil d.w. (which is equivalent to application rates up to 3 kg test item/ha) since no statistically significant effects were observed at this test item concentration. The EC10 was determined to be >4.00 mg test item/kg soil d.w. since the inhibition at this highest test item concentration was below 10%.

Table 1: Effects on carbon transformation in soil after treatment with Tefuryltrione a.s.

Days after application	Control		0.40 mg test item/kg soil d.w. equivalent to 0.3 kg test item/ha			4.00 mg test item/kg soil d.w. equivalent to 3 kg test item/ha
	O2 consumption [mg/kg soil d.w./h]	CV [%]	O2 consumption [mg/kg soil d.w./h]	CV [%]	Deviation from control [%]1	O2 consumption [mg/kg soil d.w./h]	CV [%]	Deviation from control [%]1
0	10.66	0.2	10.62	0.2	-0.4 n.s.	10.57	0.7	-0.8 n.s.
7	9.96	1.5	9.93	0.8	-0.3 n.s.	9.74	0.9	-2.2 n.s.
14	9.63	1.5	9.68	0.9	+0.5 n.s.	9.68	1.0	+0.5 n.s.
28	9.56	0.1	9.44	1.3	-1.2 n.w.	9.42	0.9	-1.5 n.w.

The calculations were performed with unrounded values.

CV [%] = Coefficient of Variation

1) based on O2 consumption; - = inhibition; + = stimulation

n.s. = No statistically significant difference to the control (Student-t-test for homogeneous variances, 2-sided, p ≤ 0.05)

n.w. = No statistically significant difference to the control (Welch-test for inhomogeneous variances, 2-sided, p ≤ 0.05)

Table 2: Validity criteria for OECD 217 (2000)

Criterion from the guideline	Outcome	Validity criterion fulfilled
The variation between replicate control samples should be less than ± 15%.	The coefficients of variation in the control were maximum 1.5 %.	yes

 

Validity criteria fulfilled:

yes

Remarks:

See Table 2 in "Any other information on results incl. tables".

Reference 2

Endpoint:

toxicity to soil microorganisms

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27 Jul - 25 Sep 2012

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 216 (Soil Microorganisms: Nitrogen Transformation Test)

Version / remarks:

adopted January 21, 2000

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Sächsisches Staatsministerium für Umwelt und Landwirtschaft, Dresden, Germany

Analytical monitoring:

no

Vehicle:

no

Details on preparation and application of test substrate:

AMENDMENT OF SOIL
- Type of organic substrate: soil was enriched with lucerne meal (Wassergut Canitz, 42.83% carbon content, 2.74% nitrogen content, 15.6/1 C/N relation, particle size: <0.25 mm). One additional soil sample (without lucerne meal) was used for determination of the initial NO3-N-content. The initial NO3-N-content was 0.81 mg/100 g soil d.w.

APPLICATION OF TEST SUBSTANCE TO SOIL
- Method: test item was mixed with quartz sand and was mixed with the soil by means of a hand stirrer. For an optimum distribution of the test item in the soil, the test item quartz sand mixture was applied at a ratio of about 10 g/kg soil dry weight. Water was added to the soil to achieve a water content of approximately 45% of WHC. The water content of the soil in each test vessel was determined at test start (after application) and adjusted once a week to the required range of 40 - 50% of WHC.

VEHICLE: no vehicle was used

Test organisms (inoculum):

soil

Total exposure duration:

56 d

Remarks:

The test duration was 56 days but according to the OECD guidleine 216 a duration of 28 days is required.

Test temperature:

19.7 - 21.0 °C in a climatic room

Moisture:

water holding capacity: 32.46 g/100 g soil d.w.
Water content of soil: 14.55 - 15.66 g/100 g soil d.w. (equivalent to 44.84 - 48.23% of WHC)

Details on test conditions:

TEST SYSTEM
- Testing facility: BioChem agrar; Labor für biologische und chemische Analytik GmbH, Gerichshain, Germany
- Test container: 500 mL wide mouth glass flasks (500 mL), screw caps of the flasks used permitted an air exchange
- Amount of soil: 200 g soil dry weight (= one sub-sample) per vessel
- No. of replicates per concentration: 3
- No. of replicates per control: 3
- No. of replicates per vehicle control: no vehicle was used

SOIL INCUBATION
- Method: incubation of the soil samples was performed as series of individual and equally sized sub-samples (1000 g soil dry weight) of each treatment group, Temperature: 19.7 - 21.0 °C in a climatic room; incubated in the dark; Water content: 14.55 - 15.66 g water/100 g soil d.w. (equivalent to 44.84 – 48.23% of WHC); duration: 56 d
SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographical reference of sampling site (latitude, longitude): Wassergut Canitz, Germany; GPS-position 12.694435960 degrees East, 51.403774567 degrees North
- History of site: Cultivation at soil removal (2012) and pre-cultivation (2011) fallow ground
- Treatments with fertilizers: none since 2003
- Treatments with plant protection products: last 1990
- Depth of sampling: 20 cm

Soil texture
- % sand: 52.4 (according to USDA), 50.7 (according to DIN 11277)
- % silt: 38.2 (according to USDA), 39.9 (according to DIN 11277)
- % clay: 9.4 (according to USDA and DIN 11277)
- Soil taxonomic classification: sandy loam (according to USDA) and loamy sand (according to DIN 4220)
- Soil classification system: USDA and DIN 4220
- pH (in water): 6.2
- Initial nitrate concentration for nitrogen transformation test (mg nitrate/kg dry weight): NO3-N content was 0.81 mg /100 g soil d.w.
- Maximum water holding capacity (in % dry weigth): 32.46 g/100 g soil d.w.
- Cation exchange capacity: 8.8 cmol+/kg soil
- Pretreatment of soil: dried at room temperature and passed through a 2 mm mesh sieve (18.05 – 22.05.2012). Before application, the soil was adapted to test conditions (conditioning phase: 18.07. - 27.07.2012)
- Storage: at approx. 4 °C in containers under aerobic conditions in the dark, 22.05. - 18.07.2012
- Initial microbial biomass as % of total organic C: 1.93% of Corg Carbon content of microbial biomass (corresponds to 27.26 mg C/100 g soil d.w.)

DETAILS OF PREINCUBATION OF SOIL: no additional information

EFFECT PARAMETERS MEASURED: The nitrogen transformation was determined on day 0 (after approximately 3 hours) and at intervals of 7, 14, 28, 42 and 56 days after application.

VEHICLE CONTROL PERFORMED: no

Nominal and measured concentrations:

control (quartz meal only), 0.3 kg test item./ha (corresponding to 0.4 mg/ kg soil d.w.) and 3.0 kg a.s./ha (corresponding to 4.00 mg/kg soil d.w.)

Reference substance (positive control):

yes

Remarks:

Dinoterb

Duration:

28 d

Dose descriptor:

other: trigger value of 25% inhibition compared with the control

Effect conc.:

> 0.4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

test item = act. ingr.

Basis for effect:

nitrate formation rate

Duration:

56 d

Dose descriptor:

other: trigger value of 25% inhibition compared with the control

Effect conc.:

> 4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks:

test item = act. ingr.

Basis for effect:

nitrate formation rate

Details on results:

- Other: No adverse effects could be observed at 0.40 mg test item/kg dw at time interval 14-28 days. At a concentration of 4.0 mg/kg dw no adverse effects could be observed at any time interval with the exception of 14-28 days after application.

Results with reference substance (positive control):

In a separate study the reference item Dinoterb caused stimulations of nitrogen transformation of +40.4%, +68.1% and +83.5% at 6.80 mg, 16.00 mg and 27.00 mg Dinoterb per kg soil dry weight, respectively, determined 28 d after application.

Reported statistics and error estimates:

Statistical evaluation of the test results (2-sided Student-t-test for homogeneous variances at 5% significance level and 2-sided Welch-t-test for inhomogeneous variances at 5% significance level) was performed.

Table 1: Effects on nitrogen transformation in soil after treatment with Tefuryltrione a.s.

Time Interval (days)	Control			0.40 mg test item/kg soil dry weight equivalent to 0.3 kg test item/ha				4.00 mg test item /kg dry weight soil equivalent to 3 kg test item/ha
	Nitrate-N1)			Nitrate-N1)			% difference to control	Nitrate-N1)			% difference to control
0-7	1.59	±	0.00	1.48	±	0.13	-6.9 n.d.	1.29	±	0.13	-18.9n.d.
7-14	0.67	±	0.02	0.85	±	0.13	+26.2 n.s.	0.91	±	0.15	+35.5n.w.
14-28	0.82	±	0.02	0.72	±	0.07	-11.9 n.s.	0.58	±	0.07	-29.4*s.
28-42	0.40	±	0.07	-2)	±	-2)	-2)	0.67	±	0.03	+70.5*s.
42-56	0.44	±	0.07	-2)	±	-2)	-2)	0.41	±	0.10	-7.0n.s

The calculations were performed with unrounded values

1) Rate: Nitrate-N in mg/kg soil dry weight/time interval/day, mean of 3 replicates and standard deviation

2) since in this application rate the deviation from the control was below ±25% on day 28, no further evaluations were performed

n.s. = No statistically significant difference to the control (Student-t-test for homogeneous variances, 2-sided, p ≤ 0.05)

n.w. = No statistically significant difference to the control (Welch-t-test for inhomogeneous variances, 2-sided,p ≤ 0.05)

*s. = statistically significantly different to control (Student-t-test for homogeneous variances, 2-sided, p ≤ 0.05)

n.d. = test could not be performed for mathematical reasons

Table 2: Validity criteria for OECD 216 (2000)

Criterion from the guideline	Outcome	Validity criterion fulfilled
The variation of nitrate concentrations between replicate control samples should be less than ± 15%.	The coefficients of variation in the control (NO3-N) were maximum 2.2%.	yes

 

Validity criteria fulfilled:

yes

Remarks:

See Table 2 in "Any other information on results incl. tables".

Description of key information

No adverse effects on the soil carbon transformations observed (OECD 217).

No adverse effects on the soil nitrogen transformations observed (OECD 216).

Additional information

The potential effects of tefuryltrione on nitrogen and carbon transformations by soil microorganisms was investigated using natural soil from an uncultivated site. The studies were performed according to OECD 216 (nitrogen transformation) and 217 (carbon transformation) under GLP conditions. In both studies two application rates were tested (0.3 kg/ha and 3.0 kg/ha corresponding to test concentrations of 0.4 and 4.0 mg test item (=a.s.)/kg soil dw). The test design targets the requirements for plant protection products and they were not initially planned for the derivation of an EC10/NOEC. However, both tests were performed with three replicates. In both studies, a natural soil was mixed with appropriate amounts of the test item in order to achieve the desired concentrations. Test vessels in the carbon transformations test were incubated under controlled conditions for 28 d. The study period in the nitrogen transformations tests was 56 days. At test termination no adverse effects (difference to control < 25 %) of tefuryltrione on the soil nitrogen transformations and carbon transformations were observed. In all studies the validity criteria were met and thus considered valid.