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Toxicity to soil microorganisms

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Reference
Endpoint:
toxicity to soil microorganisms
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2016
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 216 (Soil Microorganisms: Nitrogen Transformation Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Specific details on test material used for the study:
None
Analytical monitoring:
yes
Details on sampling:
On day 0 and on day 28, the water content and the nitrate content of each replicate was determined. From each replicate, a sample containing 12.5 g wet soil (corresponding to 11 g dry soil) was taken for nitrate analysis.
Vehicle:
no
Details on preparation and application of test substrate:
The test system was delivered by LUFA Speyer, Obere Langgasse 40, 67346 Speyer. Major components were sand (around 57.5%) and silt (around 35.6%). Water holding capacity (WHC) was 35.6%. The soil was not treated with plant protection products within the last year and no organic fertilizer was applied for at least 6 months. No inorganic fertilizer was applied for at least 3 month as required in the guideline.
The soil used was freshly collected and stored in the test facility in the dark for 8 days under aerobic conditions at 18.0 - 21.2 °C. A mixture of 3 parts of whole wheat flour and 1 part of soya wheat flour with a C/N ratio of 13.56/1 (determined by elemental analysis performed at Mikroanalytisches Labor Pascher, Rema-gen, Germany) was used as a nitrogen source for the test.
The soil was divided into six portions of 900 g. The required amount of 4.3 g organic substrate was given to each portion and the soil samples were homogenised. To each portion the required amount of test item was added using 8.5 g of quartz sand (in which the relevant amount of test item was previously mixed) and 77 mL deionised water which was also used for adjustment of the soil moisture and then thoroughly mixed. Each portion was divided into 3 replicates of 300 g each and incubated in glass jars. The amount of organic substrate corresponded to the recommended value in the guideline (5 g substrate per 1000 g dry weight of used test soil)
Test organisms (inoculum):
soil
Total exposure duration:
28 d
Remarks:
None.
Test temperature:
19.1°C - 22.1°C (min - max)
Moisture:
14% - 14.7% (min - max), (per kg soil dw)
Details on test conditions:
TEST SYSTEM
- Testing facility: LAUS
- Test container (type, material, size): glass jar of 1 L
- Amount of soil: 300 g
- No. of replicates per concentration: 3
- No. of replicates per control: 3

SOIL INCUBATION
- Method: The replicates were left to stand for 28 days. During this period, the replicates were manually homogenised twice a week. Each week, the water content of each test vessel was adjusted to maintain the water content of the start of the test.

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Source: LUFA Speyer
- Batch F 2.3 1216
- Geographical reference of sampling site (latitude, longitude): Offenbach, Germany
- Sampling date: 23 March 2016
- History of site: Site not used in the past two years
- Vegetation cover: Uncultivated
- Treatments with pesticides: None for at least one year.
- Treatments with organic fertilizer: None for at least 6 months.
- Treatments with inorganic fertilizer: None for at least 3 months.
- Accidental contamination: None known
- Depth of sampling: 20 cm
- Soil texture:
- % sand: 54.8-60.2%
- % silt: 32.8 - 38.4%
- % clay: 5.3 - 8.5%
- Soil taxonomic classification: Silty sand
- pH (in water): 5.9
- Initial nitrate concentration for nitrogen transformation test (mg nitrate/kg dry weight): 77.6 mg NO3 per kg soil dry weight
- Maximum water holding capacity (in % dry weigth): 35.6%
- Cation exchange capacity (meq/100g): 7.6 ± 0.8
- Storage (condition, duration): The soil used was freshly collected and stored in the test facility for 1 week under aerobic conditions at 18.8 - 21.1 °C
- Initial microbial biomass as % of total organic C: 3.26 - 3.57 % of organic carbon
Nominal and measured concentrations:
Nominal concentrations: 19 / 58 / 175 / 524 / 1572 mg/kg soil dry weight.
Concentrations not analytically measured.
Reference substance (positive control):
no
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 572 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
nitrate formation rate
Details on results:
The quantities of nitrate in mg nitrate/kg dry weight soil formed in the treated samples after 28 days were compared to that formed in the control. From these data, the % inhibition values for each test concentration were calculated. “Nitrate formed” means “nitrate measured at D28” minus “nitrate measured at D0”. The % inhibition values were expressed in % of the nitrate formed in the control group (table 1). There were no significant inhibition of nitrate production at any concentration tested. The NOEC is considered superior or equal to 1572 mg/kg soil dw.
Results with reference substance (positive control):
None.
Reported statistics and error estimates:
Calculation of results was performed with the help of validated software. The estimation of the biological results was accomplished using the software ToxRat® Professional, version 3.2.1., default template for OECD 216 study.

Table 1: Nitrate production in % compared to blank control. Negative inhibition implies a boost of nitrate production compared to the blank control.

Concentration in mg/kg
dry matter

Inhibition of Nitrate production in %
compared to blank control

19 mg/kg

-7.6

58 mg/kg

-3.6

175 mg/kg

-7.7

524 mg/kg

-6.7

1572 mg/kg

-15.1

Table 2: Measured nitrate concentrations in replicate vessels at Day 0 and D28

Concentration in

mg/kg
dry matter

Vessel No.

NO3Concentration

in mg/kg dry soil

Day 0

Day 28

Blank control

1

81.54

270.30

2

74.30

266.77

3

75.75

273.45

19 mg/kg

4

75.01

283.75

5

76.23

279.93

6

73.65

284.14

58 mg/kg

7

86.75

282.53

8

78.93

281.87

9

76.64

277.46

175 mg/kg

10

77.88

282.49

11

75.12

284.49

12

80.15

289.64

524 mg/kg

13

77.96

282.88

14

76.06

280.84

15

78.68

286.64

1572 mg/kg

16

78.58

313.57

17

77.03

291.35

18

76.14

293.30

QC Samples

QC 1

50.72

48.50

QC 2

49.99

49.14

% Recovery

100.31

97.64

 

Table 3: Water content in replicate vessels at Day 0 and Day 28.

Concentration in

mg/kg
dry matter

Vessel No.

Water content

in % (dry matter)

Day 0

Day 28

Blank control

1

14.46

14.55

2

14.22

14.67

3

14.19

14.68

19 mg/kg

4

14.16

14.33

5

14.09

14.56

6

14.03

14.50

58 mg/kg

7

14.30

14.61

8

14.16

14.57

9

14.08

14.45

175 mg/kg

10

14.11

14.44

11

14.08

14.32

12

14.13

14.36

524 mg/kg

13

14.08

14.54

14

14.00

14.13

15

13.99

14.45

1572 mg/kg

16

13.84

14.37

17

13.87

14.18

18

13.96

14.50

 

Validity criteria fulfilled:
yes
Conclusions:
The study was performed using 5 treatments of 19 / 58 / 175 / 524 / 1572 mg/kg nominal concentration. Sieved soil was exposed to the test item for 28 days. After 0 and 28 days, the nitrate content of the soil of each replicate was determined. A blank control was tested. There were no significant inhibition of nitrate production compared to the control at any tested concentration. NOEC is therefore considered superior or equal to 1572 mg/kg soil dry weight.
Executive summary:

This study investigated the effect of bis(dibutyldithiocarbamato-S,S')copper on soil microorganisms involved in nitrification of organic matter. The study was performed according to the OECD 216 guideline. The experimental design was in triplicate, using 5 treatments of 19 / 58 / 175 / 524 / 1572 mg/kg soil dry weight nominal concentration and a control. A sieved natural soil was used. Each replicate was exposed to a relevant amount of test item for 28 days. After 0 and 28 days, the nitrate content of the soil of each replicate was determined. There were no significant inhibition of nitrate production compared to the control at any tested concentration. NOEC is therefore considered superior or equal to 1572 mg/kg soil dry weight.

Description of key information

This study investigated the effect of bis(dibutyldithiocarbamato-S,S')copper on soil microorganisms involved in nitrification of organic matter. The study was performed according to the OECD 216 guideline. The experimental design was in triplicate, using 5 treatments of19 / 58 / 175 / 524 / 1572 mg/kg soil dry weight nominal concentration and a control. A sieved natural soil was used. Each replicate was exposed to a relevant amount of test item for 28 days. After 0 and 28 days, the nitrate content of the soil of each replicate was determined. There were no significant inhibition of nitrate production compared to the control at any tested concentration. NOEC is therefore considered superior or equal to 1572 mg/kg soil dry weight.

Key value for chemical safety assessment

Additional information