Didecyldimethylammonium chloride

Toxicity to soil macroorganisms except arthropods

Link to relevant study record(s)

Reference

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2008-01-16 to 2008-07-15

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 222 (Earthworm Reproduction Test (Eisenia fetida/Eisenia andrei))

Version / remarks:

DIN ISO 11268-2:1998 (E), BBA Guideline

Deviations:

yes

Remarks:

Fresh Lufa 2.2 was used instead of artificial soil

Principles of method if other than guideline:

LUFA 2.2 soil, classified as a loamy sand, was used rather than the soil recommended by the OECD guideline as it is considered a reasonable worst case and this laboratory has experience with this soil. Moreover, the OECD soil contains 20 % kaolinite, a clay type which may influence the test results if used in conjunction with a cationic surfactant. Fresh rather than dried soil was used as this is closer to the soil that would be found in the field and is therefore expected to be more environmentally realistic.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

yes

Details on preparation and application of test substrate:

- Method of mixing into soil (if used): The necessary amounts of the test item were dispersed in the amount water necessary to adjust a humidity of 50-70% of the maximum water holding capacity. Subsequently, the test medium was thoroughly mixed into the soil with a long handled blender for at least four minutes and filled into the test vessels.
- Control: Fresh LUFA 2.2 standard soil moistened with demineralised water without test or reference item
- Vehicle control: Fresh LUFA 2.2 standard soil mixed with quartz sand and moistened with demineralised water without test or reference item

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

TEST ORGANISM
- Common name: Eisenia fetida SAVIGNY 1826, subspecies: Eisenia fetida fetida
- Source: Own breeding (Source: Bundesanstalt fuer Verbraucherschutz und Landwirtschaft (BVL), Messeweg 11/12, D-38104 Braunschweig)
- Age at test initiation (mean and range, SD): 9-10-month, not variating more than 1 month
- Weight at test initiation (mean and range, SD): 0.53 +/- 0.04 g


ACCLIMATION
- Acclimation period: 2 days prior to test start
- Acclimation conditions (same as test or not):same as test
- Health during acclimation (any mortality observed): healthy

Study type:

laboratory study

Substrate type:

natural soil

Limit test:

no

Total exposure duration:

56 d

Remarks:

removement of adults after 28 days

Test temperature:

20 +/- 2 °C

pH:

Table 1: pH-Values of the Test Media
Measurement of pH and of test medium was based on mixed samples of all replicates per treatment
and control.
Application rate pH-value
[mg/kg soil DW] day 0 day 56
Control 5.618 6.52
Vehicle Control 5.710 6.72
125 5.773 6.66
250 5.749 6.67
500 5.779 6.72
1000 5.914 6.75

Moisture:

Table 1: Moisture of the Test Media
Measurement of moisture content of test medium was based on mixed samples of all replicates per
treatment and control.
Application rate Moisture [%] of dry weight

[mg/kg soil DW] day 0 day 55
Control 24.7 35.6
62.5 24.3 34.3
125 25.3 32.6
250 25.0 31.5
500 23.6 33.8
1000 25.0 32.7

Details on test conditions:

TEST SYSTEM
- Test container (material, size): round plastic boxes, ID: 15.0 cm, bottom surface area: 177 cm3, height: 14 cm; transparent and perforated lids enabled sufficient gas exchange and light penetration and prevented the test substrate from drying
- Amount of soil or substrate: 600 g DW
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 4
- No. of replicates per control: 8
- No. of replicates per vehicle control: 4


SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographic location: Germany / Rheinland Pfalz / Hanhofen, "Grosser Striet", No. 585, owner: Appel, Hanhofen
- Pesticide use history at the collection site: None (former 4 years)
- Collection procedures: Sampling date 2007-10-30, batch F2.24407
- Sampling depth (cm): approximately 20 cm
- Soil texture (if natural soil)
- % sand: 0.63 - 2.0 mm: 0.7 %, 0.2 - 0.63 mm: 44.4 %, 0.063 - 0.2 mm: 35.7 %
- % silt: 0.02 - 0.063 mm: 5.2 %, 0.006 - 0.02 mm: 3.9 %, 0.002 - 0.006 mm: 3.4 %
- % clay: 6.7
- Soil taxonomic classification: loamy sand
- Organic carbon (%): 2.29
- Maximum water holding capacity: 49.7 g/100 g DW
- CEC: 10 mval/100 g
- Pretreatment of soil: Drying (room temperature, until sievable), presieving 10 mm, last sieving 2 mm, moistening at approximately 19 % of water holding capacity
- Storage (condition, duration): 4 - 7 °C for 43 days prior to use



OTHER TEST CONDITIONS
- Photoperiod: 16 h light - 8 h dark
- Light intensity: 400 - 800 lx


EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
Adult mortality, biomass, behaviour and pathological symptoms after 28 days
Reproductive output after 55 days

VEHICLE CONTROL PERFORMED: yes


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2
- Justification for using less concentrations than requested by guideline: Due to the results of the preliminary test no effects were expected.
- Range finding study (NON-GLP)
- Test concentrations: 100 - 500 - 1000 - 2000 mg/kg soil DW
- Results used to determine the conditions for the definitive study: No mortality or significant change of live weight was determined.

Nominal and measured concentrations:

Nominal application rates: 62.5- 125 - 250 - 500 - 1000 mg/kg soil dry weight (dw)

Reference substance (positive control):

yes

Remarks:

1.0 - 2.5 - 5.0 mg Carbendazim/kg soil DW

Key result

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

ca. 125 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

other: reproduction, mortality, body weight

Key result

Duration:

28 d

Dose descriptor:

EC50

Effect conc.:

ca. 509 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Basis for effect:

reproduction

Details on results:

- Mortality at end of exposure period: No significant mortality (> 10 %) was observed in any of the tested test item and control groups.
- Total mass of adults at beginning of test: 0.49 g (mean)
- Changes in body weigth of live adults (% of initial weight) at end of exposure period:
Control: 11.1 +/- 6.7, 62.5 mg/kg soil: 11.9 +/- 4.9, 125 mg/kg soil DW: 15.4 +/- 4.0, 250 mg/kg soil DW: 8.7 +/- 5.8, 500 mg/kg soil DW: 9.4 +/- 6.7, 1000 mg/kg soil DW: 17.9 +/- 8.5
- No. of offspring produced:
Control: 416 +/- 48.1, 62.6 mg/kg soil DW: 435 +/- 57.9, 125 mg/kg soil DW: 453 +/- 67.6, 250 mg/kg soil DW: 298 +/- 59.7, 500 mg/kg soil DW: 239 +/- 31.5, 1000 mg/kg soil DW: 58 +/- 35.3
- No. of unhatched cocoons: Not determined
- Morphological abnormalities: none
- Behavioural abnormalities: None
- Other biological observations: No significant mortality was observed

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: EC50 (Reproduction) = 1.19 mg/kg

Reported statistics and error estimates:

One Way Analysis of Variance (ANOVA and Dunnett's Method) was carried out for determination of statistically significant differences between control and vehicle control and further between pooled control and test item concentrations. Prior to Analysis of Variance, a Normality Test and an Equal Variance Test were conducted. P-values for both normality and equal variance test are 0.05. The a-value for ANOVA test (acceptable probability of incorrectly concluding that there is a difference) is a = 0.05.

Adult mortality:

No significant mortality was observed in any of the tested doses of the test item and the control groups.

Behaviour and Pathological Symptoms:

No pathological and behavioural symptoms were found at the tested application rates and control groups.

Live weight:

At test start earthworms had a mean weight of 0.49 g. The body weights of earthworms in tested doses and control groups increased during test period. Effects on body live weights after 28 days were tested for statistical significance by One Way Analysis of Variance (ANOVA). There was no statistically significant difference when compared to the control.

Reproduction rate:

The average number of juveniles of treated groups ranged between 453 and 58 individuals. Mean number of juveniles found in control groups were between 316 and 460. The coefficient of variation calculated for the reproduction of control was lower than 30 %. Statistically significant difference to the control groups was observed for the 250, 500 and 1000 mg/kg dw application rate (ANOVA).

Validity criteria fulfilled:

yes

Conclusions:

Under the study conditions, the EC50 for reproduction was 509 (95% CI: 412-629) mg/kg soil dw. The NOEC was 125 mg/kg soil dw.

Executive summary:

The effects of the test substance (40% purity) on mortality, biomass and reproductive potential of the earthworm Eisenia fetida (Savigny) were determined according to the DIN ISO Method 11268-2 and OECD Guideline 222, in compliance with GLP. The study was conducted under static conditions over 8 weeks with concentrations of 62.5, 125, 250, 500 and 1000 mg a.i./kg soil dry weight (dw) applied to natural soil (LUFA 2.2). Forty test organisms, with an individual weight of 0.30 to 0.60 g, were divided into 4 replicate groups of 10 test organisms. After 28 days, no significant mortality and no pathological symptoms or changes in behaviour of the adult earthworms were observed at any of the tested application rates and in the control. The individual body live weights increased in the control as well as in test substance groups, with no significant differences between the groups. After further four weeks, the reproduction rate (average number of juveniles) was 416 in the control and ranged between 58 and 435 in the test substance groups. Compared to the control, reproduction was significantly reduced 250, 500 and 1000 mg a.i./kg soil dw. Under the study conditions, the EC50 for reproduction was 509 (95% CI: 412-629) mg/kg soil dw. The NOEC was 125 mg/kg soil dw (Winkelmann, 2008).  

Description of key information

Based on the results of the study, the EC50 and NOEC for reproduction was 509 and 125 mg/kg soil dw respectively.

Key value for chemical safety assessment

Short-term EC50 or LC50 for soil macroorganisms:

509 mg/kg soil dw

Long-term EC10, LC10 or NOEC for soil macroorganisms:

125 mg/kg soil dw

Additional information

The effects of the test substance (40% purity) on mortality, biomass and reproductive potential of the earthworm Eisenia fetida (Savigny) were determined according to the DIN ISO Method 11268-2 and OECD Guideline 222, in compliance with GLP. The study was conducted under static conditions over 8 weeks with concentrations of 62.5, 125, 250, 500 and 1000 mg a.i./kg soil dry weight (dw) applied to natural soil (LUFA 2.2). Forty test organisms, with an individual weight of 0.30 to 0.60 g, were divided into 4 replicate groups of 10 test organisms. After 28 days, no significant mortality and no pathological symptoms or changes in behaviour of the adult earthworms were observed at any of the tested application rates and in the control. The individual body live weights increased in the control as well as in test substance groups, with no significant differences between the groups. After further four weeks, the reproduction rate (average number of juveniles) was 416 in the control and ranged between 58 and 435 in the test substance groups. Compared to the control, reproduction was significantly reduced 250, 500 and 1000 mg a.i./kg soil dw. Under the study conditions, the EC50 for reproduction was 509 (95% CI: 412-629) mg/kg soil dw. The NOEC was 125 mg/kg soil dw (Winkelmann, 2008).