Registration Dossier

Administrative data

Endpoint:
toxicity to soil microorganisms
Type of information:
experimental study
Adequacy of study:
key study
Study period:
22 November - 20 December 1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995
Report Date:
1995

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
other: Council Directive 91/414/EEC (Annex II, 8.5) 'Effects on soil non-target microorganisms'
Deviations:
not specified
Qualifier:
according to
Guideline:
other: EPPO guideline 'Recommended Laboratory tests for assessing hazard to soil microflora'
Deviations:
not specified
Qualifier:
according to
Guideline:
other: FAO guideline 'Annex to revised guidelines on environmental criteria for the registration of pesticides'
Deviations:
not specified
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Test material: Simazine technical ai
- Appearance: White powder
- Batch No.of test material: 74
- Expiration date of the batch: September 1996
- Purity: 97.7%

STABILITY AND STORAGE CONDITIONS OF TEST MATERIAL
- Storage condition of test material: In the dark at 4°C

Sampling and analysis

Analytical monitoring:
no
Details on sampling:
Nitrogen transformation: Quadruplicate samples (replicates A, B, C and D) of each of the control and treated soil groups were withdrawn approximately 3 hours after preparation (Day 0) and at 14 and 28 days after preparation and the samples transferred to a deep-freeze at approximately -20°C to await analysis. Analysis for ammonium, nitrite and nitrate ion concentrations was performed using a continuous flow colourimetric autoanalyser.

Carbon mineralisation: The NaOH in the respirometer traps was removed and replaced with 40 mL fresh 1 M NaOH approximately 3 hours after preparation (Day 0) and at 2, 7, 14, 21 and 28 days after preparation. Analysis was performed by autotitration.

Test substrate

Vehicle:
no
Details on preparation and application of test substrate:
EXPERIMENTAL DESIGN:
The following mixtures of soil, lucerne meal, distilled water and test substance : sand mixture were prepared:
Group 1: Soil + water
Group 2: Soil + lucerne meal + sand + water
Group 3: Soil + lucerne meal + Simazine technical ai : sand + water

AMENDMENT OF SOIL
- Type of organic substrate: Lucerne meal having a nitrogen content of approximately 3.0% (C:N ratio 13.8:1) was used and 22,4 g was added to give a maximum of 0.5% of the soil dry weight.

APPLICATION OF TEST SUBSTANCE TO SOIL
- Method: Simazine technical ai was first mixed into dry fine sand so that 20.47 mg Simazine technical ai was contained in 44.85 g sand per 5 kg soil (4485 g dry weight). For soil treatment Groups 2 and 3, the lucerne meal sand and test substance : sand mixture were distributed evenly in several aliquots over the surface of the test soil (spread as a thin layer of approximately 2.5 cm) and then thoroughly mixed after each aliquot addition.

Test organisms

Test organisms (inoculum):
soil

Study design

Total exposure duration:
28 d

Test conditions

Test temperature:
20°C ± 2°C
Moisture:
45% of its maximum water holding capacity (35.04%)
Details on test conditions:
TEST SYSTEM
- Test container (type, material, size): Glass jars (nitrogen transformation); respirometer flasks (carbon mineralisation)
- Amount of soil: 100 g
- No. of replicates per concentration: 4 replicates (nitrogen transformation); 3 replicates (carbon mineralisation)
- No. of replicates per control: 4 replicates (nitrogen transformation); 3 replicates (carbon mineralisation)

SOIL INCUBATION
- Method: in the dark at 20°C ± 2°C

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographical reference of sampling site (latitude, longitude): Clifton, Nottinghamshire, England, OS map reference SK 542 338
- History of site: Uncropped for last 2 years. Last crop oilseed rape harvested in 1992
- Vegetation cover: Bare soil/ very light weed infestation
- Treatments with pesticides or fertilizers: Not received treatment during the preceding 2 years
- Depth of sampling: 20 cm
- % sand: 61.03%
- Soil taxonomic classification: Sandy loam
- pH (in water): 6.7
- Organic carbon: 1.3%
- Water holding capacity: 35.04%
- Cation exchange capacity: 15.0 mEq/100 g
- Initial microbial biomass as % of total organic C: 1.57%
Nominal and measured concentrations:
3000 g active ingredient (ai) per hectare (twice the maximum expected concentration, 2 x MEC) equivalent to 20.47 mg ai/ 5 kg soil
Reference substance (positive control):
yes
Remarks:
Dinoseb acetate: applied to the test soil at a concentration of 40 mg/kg soil to determine effects on nitrogen transformation and at a concentration of 100 mg/kg soil to determine effects on carbon mineralisation.

Results and discussion

Effect concentrationsopen allclose all
Key result
Duration:
28 d
Dose descriptor:
other: Single dose of 4.094 mg/kg
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
other: Loss of ammonium
Remarks on result:
other: Conversion of ammonium to nitrate is not affected at the single concentration tested of 4.094 mg/kg.
Key result
Duration:
28 d
Dose descriptor:
other: Single dose at 4.094 mg/kg
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
nitrate formation rate
Remarks on result:
other: Nitrate formation was actually slightly higher than the control without the simazine
Key result
Duration:
28 d
Dose descriptor:
other: Single dose of 4.094 mg/kg
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
respiration rate
Remarks on result:
other: respiration of C was comparable with control
Details on results:
Nitrogen transformation in the soils amended with lucerne meal (Group 2 control soil and Group 3 Simazine technical ai treated soil) was evident from the decline in ammonium concentrations and increase in nitrate concentrations over the first 14 days. At the end of the 28 day study period the deviation in measured activity in Group 3 soil compared with Group 2 soil was less than 25% for both ammonium (+8.33%) and nitrate (+16.13%) concentrations.

Carbon mineralisation in the Group 2 and Group 3 soils amended with lucerne meal was evident from the cumulation of CO2 over the first 14 days. At the end of the 28 day study period the deviation in cumulative mean ppm C as CO2/g soil for Group 3 compared with Group 2 was less than 25% at +1.00%.

The EPPO Decision-making scheme for the environmental risk assessment of plant protection products states that 'if deviation from the control treatment at the end of the test is <25%' then the test substance is 'categorised as low risk to soil microflora'.
Results with reference substance (positive control):
- Results with reference substance valid: Yes
- Relevant effect levels: After 28 days of incubation it was demonstrated that the treated soil showed deviations in measured activity greater than 25% compared to the untreated control soil in all parameters examined.
Reported statistics and error estimates:
Since the variability in Group 1 frequently differed from that in the other two groups, Group 1 was omitted from the analyses, which therefore provided a comparison between Groups 2 and 3 only.
No significant differences were found in ammonium, nitrate or carbon levels between Groups 2 and 3.

Any other information on results incl. tables

Table 1: Ammonium as mg N/g soil (mean of 4 replicates)

Day

Group

Group 3 : Group 2 deviation (%)

1

2

3

0

0.71

16.20

16.74

+3.33

14

0.27

1.90

2.23

+17.37

28

0.14

0.24

0.26

+8.33

 

Table 2:Nitrate as mg N/g soil (mean of 4 replicates)

Day

Group

Group 3 : Group 2 deviation (%)

1

2

3

0

12.02

15.12

14.77

-2.31

14

17.65

34.80

35.92

+3.22

28

21.42

48.16

55.93

+16.13

 

Table 3: Carbon mineralisation – cumulative mean (of 3 replicates) ppm C as CO2/g soil

Day

Group

Group 3 : Group 2 deviation (%)

1

2

3

0

0.00

31.54

19.18

-39.19

2

8.26

348.52

352.71

+1.20

7

29.80

668.11

673.84

+0.86

14

54.26

808.54

814.26

+0.71

21

73.60

887.79

897.72

+1.12

28

89.38

943.16

952.57

+1.00

Applicant's summary and conclusion

Validity criteria fulfilled:
not specified
Conclusions:
The results from the study demonstrated that nitrogen transformation and carbon mineralisation in soil treated with Simazine technical ai were comparable to the results obtained from the control soil. The deviations in measured activity at the end of the study period were less than 25% for all parameters examined.

Under anticipated conditions of field use, Simazine technical ai can be considered to present a low risk to soil microflora.