Dipotassium tetraborate

Toxicity to soil macroorganisms except arthropods

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: methods and results well documented

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Version / remarks:

Environment Canada (2002)

Deviations:

not specified

GLP compliance:

not specified

Analytical monitoring:

not specified

Vehicle:

not specified

Details on preparation and application of test substrate:

Test compound added in solution to soil. Soil moisture was 35% (based on wet weight). Tests started after a 1 day equilibration period.

Test organisms (species):

Lumbricus sp.

Animal group:

annelids

Study type:

laboratory study

Substrate type:

other: artificial soil and natural soil

Limit test:

no

Total exposure duration:

14 d

Test temperature:

20°C

pH:

artificial soil: 6.09
clay-loam reference soil: 6.05

Moisture:

35%

Details on test conditions:

TEST SYSTEM
- No. of replicates per treatment group: 4


SOURCE AND PROPERTIES OF ARTIFICIAL SOIL
- Ingredients: 70% silica sand, 20% kaolinite clay, 10% spaghnum peat, 1-3g calcium carbonate per kg soil
- Soil texture
- % sand: 77.3
- % silt: 7.8
- % clay: 14.9
- Total carbon (%): 4.46
- Organic matter (%): 9
- pH water (1:2): 6.09
- eCEC: 18.5 cmol c/kg
- background B concentration: 3.5 mg B/kg dw

SOURCE AND PROPERTIES OF CLAY LOAM REFERENCE SOIL
- collected soil from Alberta (Canada)
- classified as a Delacour Orthic Black Chernozem.
- Soil texture
- % sand: 26.6
- % silt: 43.3
- % clay: 30.1
- Total carbon (%): 6.83
- Organic matter (%): 12.8
- pH water (1:2): 6.05
- eCEC: 34.5 cmol c/kg
- background B concentration: 1.0 mg B/kg dw


OTHER TEST CONDITIONS
- Photoperiod: 16h light, 8h dark

Nominal and measured concentrations:

0-500-1000-2000-4000-6000-8000 mg boric acid/kg dw,
corresponding to 0-87?5-175-350-700-1050-1400 mg B/kg dw.

Reference substance (positive control):

not specified

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

786.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in artificial soil

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

501.4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in artificial soil

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

651.4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in clay loam reference soil

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

447.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in clay loam reference soil

Details on results:

nominal added dosis; background concentration reported

Reported statistics and error estimates:

7 and 14 day LCx values were generated using the Probit, Moving average and the trimmed Spearman-Karber methods.

Validity criteria fulfilled:

not specified

Conclusions:

This study followed the OECD guideline. The standard test and selection criteria were fulfilled.
Test data was not presented.

Reference 2

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: material and methods are well described. test follows OECD guideline and is GLP compliant

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Deviations:

no

Principles of method if other than guideline:

the worms were added randomly to each of the test containers. The deviation is assumed not to have affected the results of this study.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

not specified

Vehicle:

no

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

TEST ORGANISM
- Common name: redworms, brandling worms, tiger worms and red wiggler worms
- Source: commercial hatchery Blades Biological, Crowden-Edenbridge, Kent, England
- Weight at test initiation (mean and range, SD): 355-599 mg


ACCLIMATION
- Acclimation conditions: standard conditions in a horse-manure, garden soil mixture (1:1) at 24 +/- 2°C

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

14 d

Test temperature:

19.4 - 19.7 °C

pH:

6.0-6.2

at start highest test concentration: 6.1
at end highest concentration with survuval: 6.2

Moisture:

54.66%- 54.96%

Details on test conditions:

TEST SYSTEM
- Test container (material, size): one little all glass jars, provided with a glass lid (domestic preserving jars)
- Amount of soil or substrate: 775 g
- No. of organisms per container (treatment): 10
- No. of containers per concentration: 5
- No. of concentrations: 7 + 1 control
- equilibration period: 0 days


SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Sphagnum peat: Pindstrup Sphagnum oriniating from Denmark, DEGA Potgrond, Delft. Arrival date: March 28, 1994.
- Fine industrial sand: M32, Eurogrit BV, Nieuwegein. Arrival date: June 24, 1999.
- Kaolin clay: China Clay Ast containing 85-90% kaolin. Fa Vingerling, Haastrecht. Arrival date: March 18, 1992.
- Composition of artificial soil:
* silica sand: 70%
* kaolinite clay: 20%
* sphagnum peat: 10%
- OM %: 10
- Clay %: 20
- Cb: 3.5 mg B/kg dw


PRETREATMENT OF SOIL
3.4, 9.8, 30.7, 96.4, 300.8, 961.6 and 3001.1 mg Boric acid were accurately weighed out and dissolved in 600ml of ultrapure water. from each of these solutions 100ml was added to 675g of artificial soil respectively yielding nominal concentrations of 1.1, 3.3, 10, 32, 100, 320 and 1000 mg per kg of dry soil. All soil/water mixtures were mechanically mixed.



OTHER TEST CONDITIONS
- Light intensity: 556-582 lux (low intensity illumination)

Nominal and measured concentrations:

0, 1.1, 3.3, 10, 32, 100, 320, 1000 mg boric acid/kg

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

>= 175 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

>= 175 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

> 175 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Details on results:

at the highest test concentration (1000 mg/kg dw) no effects on survival, weight, behaviour and condition were observed after 14 days.
nominal added dosis, background concentration reported.

Reported statistics and error estimates:

To determine the NOEC for weight, a multiple comparison was made between the average weight of the worms per container at each concentration and the average weight in the controls using a two-tailed Dunnett test. A significance level of 99% (a=0.01) and 95% (a=0.05) was employed. In case of 99% significance only that significance is given.
To determine the NOEC for survival, the survival rates at each concentration were compared pair-wise with those in the control using a binomial test. A significance level of 95% was employed.
The NOEC for behaviour and condition was visually determined and was not statistically verified.

At 1000 mg/kg dry soil no adverse effects with respect to survival was observed and the NOEC for survival is therefore equal to or greater than 1000 mg/kg dry soil. At 1000mg per kg of dry soil there is no mortality. Although one of the 40 worms was not recovered at both 3.2 and 320 mg/kg dry soil. This 'mortality' is not taken into account in establishing the test parameters as the OECD guideline allows 10% mortality in the control.

The LC100 is established to be greater than 1000mg/kg dry soil because no total mortality was observed at any of the concentrations tested.

Adverse effects with respect to the behaviour and condition of the test animals were not observed at any concentration tested. The NOEC for these is therefore equal or greater than 1000mg per kg dry soil.

Validity criteria fulfilled:

yes

Conclusions:

Standard test and selection criteria were fullfilled during this study. A standard guideline was followed, the test are well performed and the study is well documented. Unbounded values are not used for PNEC derivation.

Reference 3

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: according to the OECD guideline or national guideline (Environment Canada), standard test and selection criteria fulfilled.

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Deviations:

not specified

Qualifier:

according to guideline

Guideline:

other: Environment Canada (2002)

Deviations:

not specified

GLP compliance:

not specified

Analytical monitoring:

not specified

Vehicle:

not specified

Details on preparation and application of test substrate:

Test compound added in solution to soil. Soil moisture was 35% (based on wet weight). Tests started after a 1 day equilibration period.

Test organisms (species):

Eisenia sp.

Animal group:

annelids

Details on test organisms:

Eisenia andrei
- fully developed clitellum
- aged 2-12 months
- individual wet weight 250-600 mg

Study type:

laboratory study

Substrate type:

other: artificial soil and natural soil

Limit test:

no

Total exposure duration:

14 d

Test temperature:

20°C

pH:

artificial soil: 6.09
clay-loam reference soil: 6.05

Moisture:

35%

Details on test conditions:

TEST SYSTEM
- No. of replicates per treatment group: 4


SOURCE AND PROPERTIES OF ARTIFICIAL SOIL
- Ingredients: 70% silica sand, 20% kaolinite clay, 10% spaghnum peat, 1-3g calcium carbonate per kg soil
- Soil texture
- % sand: 77.3
- % silt: 7.8
- % clay: 14.9
- Total carbon (%): 4.46
- Organic matter (%): 9
- pH water (1:2): 6.09
- eCEC: 18.5 cmol c/kg
- background B concentration: 3.5 mg B/kg dw

SOURCE AND PROPERTIES OF CLAY LOAM REFERENCE SOIL
- collected soil from Alberta (Canada)
- classified as a Delacour Orthic Black Chernozem.
- Soil texture
- % sand: 26.6
- % silt: 43.3
- % clay: 30.1
- Total carbon (%): 6.83
- Organic matter (%): 12.8
- pH water (1:2): 6.05
- eCEC: 34.5 cmol c/kg
- background B concentration: 1.0 mg B/kg dw


OTHER TEST CONDITIONS
- Photoperiod: 16h light, 8h dark

Nominal and measured concentrations:

0-2000-4000-5000-6000-8000-10000 mg boric acid/kg dw,
corresponding to 0-350-700-875-1050-1400-1750 mg B/kg dw.

Reference substance (positive control):

not specified

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

860.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in artificial soil

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

649.9 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in artificial soil

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

696.1 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in artificial soil

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

581.1 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in artificial soil

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

772.7 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in clay loam reference soil

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

673.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in clay loam reference soil

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

689.1 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in clay loam reference soil

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

567.9 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: in clay loam reference soil

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

777.1 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil; asynchronous worms

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

576.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil; synchronous worms

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

693 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil; clitellated worms

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

610.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil; non-clitellated worms

Duration:

7 d

Dose descriptor:

LC50

Effect conc.:

563.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

617 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil; asynchronous worms

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

594.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil; synchronous worms

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

613 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil; clitellated worms

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

533.3 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil; non-clitellated worms

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

795.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: artificial soil

Reported statistics and error estimates:

7 and 14 day LCx values were generated using the Probit, Moving average and the trimmed Spearman-Karber methods.

Validity criteria fulfilled:

yes

Conclusions:

This study followed the OECD guideline or a national guideline (Environment Canada)
The standard test and selection criteria were fulfilled.

Reference 4

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Study was performed according to ISO 10872 guideline and is well documented.

Qualifier:

according to guideline

Guideline:

other: ISO 10872 "Water Quality – Determination of the toxic effect of sediment and soil samples on growth, fertility and reproduction of Caenorhabditis elegans"

Deviations:

no

GLP compliance:

yes

Remarks:

The study has no formal GLP certificate. However, the study was conducted at a GLP-certified laboratory, staff was appropriately trained and standard operating procedures were followed in terms of experimental performance and raw data documentation.

Analytical monitoring:

no

Vehicle:

no

Details on preparation and application of test substrate:

- The test item was dissolved in a sufficient amount of deionised water to prepare a stock solution. This stock solution was diluted with deionised water to obtain one test item solution (= dosage solution) for each test item concentration.
- Method of mixing into soil (if used): The test item was incorporated into the soil by thoroughly mixing in the respective test item solution for approx. 3 to 5 minutes.
- This amount of water/test item-mixture served also to adjust the final soil moisture of 40% of the dry weight.

Test organisms (species):

Caenorhabditis elegans

Animal group:

nematods

Details on test organisms:

The nematodes used in this test, Caenorhabditis elegans were taken from a breeding culture kept at ECT Oekotoxikologie GmbH.
The nematodes were breed on agar plates at 20 ± 2°C. The cultures were fed with Escherichia coli (OP50 uracil-poor strain) and moistened on demand with deionised water.
The nematodes inserted in the test were synchronised juveniles of the first larval stage

Study type:

laboratory study

Substrate type:

natural soil

Limit test:

no

Total exposure duration:

96 h

Test temperature:

20 +- 0.5 °C

pH:

soil pH-value: 5.7 – 5.8

Moisture:

soil moisture: 37.6 – 40.0% of the dry weight

Details on test conditions:

TEST SYSTEM
- Test container (material, size): a well with an area of 3.9 cm² and a volume of 2 to 4 mL in a 12 well multiwell plate
- Amount of soil or substrate: 0.5g fresh weight
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 2
- No. of replicates per control: 6

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographic location: Germany
- Soil name: LUFA 2.2
- Sampling depth (cm): 20
- Soil texture (if natural soil): loamy sand
- Organic carbon (%): 2.29
- Maximum water holding capacity (in % dry weigth): 49.7
- CEC: 10 mval/100g
- Sampling date: December 14, 2007
- Vegetation; uncultivated

OTHER TEST CONDITIONS
- Photoperiod: permanent dark

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : reproduction

Nominal and measured concentrations:

nominal added concentrations: control, 10.0, 31.6, 56.2, 75.0, 100, 237, 562 and 1000 mg H3BO3/kg dry weight soil,
corresponding to control, 1.75, 5.5, 9.8, 13.1, 17.5, 41.4, 98.2 and 174.8 mg B/kg dw

Reference substance (positive control):

no

Duration:

96 h

Dose descriptor:

EC10

Effect conc.:

86.7 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

number of juvenile nematodes

Reported statistics and error estimates:

Data were checked for normality by Shapiro-Wilk´s test on normal distribution and for homogeneity by Cochran’s test. Treatment means were compared by ANOVA and Williams test (p ≤ 0.05, one-sided) and tested for statistically significant differences compared to the control. Probit analysis using Linear Max. Likelihood Regression was used to calculate the EC50,EC20 and EC10 for reproduction.

96 h LOEC for reproduction: 174.8 mg B/kg soil dw (82% inhibition)

96 h NOEC for reproduction: 98.2 mg B/kg soil dw (18.6% inhibition)

96 h EC50 for reproduction: 130.6 mg B/kg soil dw

96 h LOEC for mortality: 98.2 mg B/kg soil dw (55% mortality)

96 h NOEC for mortality: 98.2 mg B/kg soil dw

Concentration [mg test item/kg soil (dw)]	Number of Juveniles[mean ± sd]	Number of Juveniles[% of control]
Control	1250.0 +-334.1	-
10.0	951.0 +- 190.9	76.1
31.6	834.5 +- 402.3	66.8
56.2	1040.5 +- 573.5	83.2
75.0	1141.0 +- 335.2	91.3
100	1163.0 +- 258.8	93.0
237	1203.5 +- 171.8	96.3
562	1017 +- 18.4	81.4
1000	225.0 +- 18.4	18.0

Validity criteria fulfilled:

yes

Conclusions:

This reliable study was performed according to the ISO 10872 guideline and is well documented. A 96-h EC10 value for reproduction of nematods of 86.7 mg B/ kg soil dw is obtained and used for PNEC derivation.

Reference 5

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was performed according to OECD 220 (2004) guideline and is well documented.

Qualifier:

according to guideline

Guideline:

OECD Guideline 220 (Enchytraeid Reproduction Test)

Deviations:

no

GLP compliance:

yes

Remarks:

The study has no formal GLP certificate. However, the study was conducted at a GLP-certified laboratory, staff was appropriately trained and standard operating procedures were followed in terms of experimental performance and raw data documentation.

Analytical monitoring:

no

Vehicle:

no

Details on preparation and application of test substrate:

- The test item was dissolved in a sufficient amount of deionised water to prepare a stock solution. This stock solution was diluted with deionised water to obtain one test item solution (= dosage solution) for each test item concentration.
- Method of mixing into soil (if used): The test item was incorporated into the soil by thoroughly mixing in the respective test item
solution for approx. 5 minutes.
- This amount of water/test item-mixture served also to adjust the final soil moisture of 40-60% of WHCmax.

Test organisms (species):

other: Enchytraeus luxuriosus

Animal group:

other: Enchytraeidae

Details on test organisms:

Enchytraeus luxuriosus (Enchytraeidae, potworms), has been cultured at ECT Oekotoxikologie GmbH since September 2003.
The worms used in the tests were adult (with eggs, i.e. white spots, in the clitellum region).

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

28 d

Test temperature:

18-21 °C

pH:

soil pH-value: 6.5

Moisture:

soil moisture: 40.0-60.0% of the WHCmax

Details on test conditions:

TEST SYSTEM
- Test container (material, size): Test vessels were glass containers (able to be closed tightly) of about 200 mL capacity and with a diameter of about 5 cm.
- Amount of soil or substrate: 20g dry weight
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 2
- No. of replicates per control: 6

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- artificial soil based on OECD guideline 207 (1984). Percentages refer to dry weight:
- 10% Sphagnum peat (no visible plant remains; finely ground and air-dried)
- 20% Kaolin-Clay (kaolinite content > 30%)
- 69.6% Quartz sand (fine sand content with particles between 50 and 200 µm higher than 50%)
- 0.4% Calcium carbonate (CaCO3)
- The moisture content was adjusted to 40 – 60% of WHCmax using deionised water.

OTHER TEST CONDITIONS
- Photoperiod: 16 : 8 h light/dark with a constant light intensity at the substrate surface of 400 – 800 lx
- Feeding: The enchytraeids were fed with about 50 mg of ground oats per test vessel at the beginning of the test (mixed with the test substrate after application of the test item). The organisms were fed an additional 25 mg ground oats once per week until day 21.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Mortality: mean number of dead adult worms as a percentage of the initial number at the start of the test for each concentration and for the control. (Validity criteria for mortality not fulfilled)
- Reproduction: mean number of juveniles per test vessel 28 days after application.

Nominal and measured concentrations:

nominal added concentrations: control, 10, 32, 56, 75, 100, 133, 178, 237, 316, 422, 562 and 1000 mg H3BO3/kg dry weight soil,
corresponding to control, 1.75, 5.6, 9.8, 13.1, 17.5, 23.2, 31.1, 41.4, 55.2, 73.8, 98.2 and 174.8 mg B/kg dw

Reference substance (positive control):

no

Duration:

28 d

Dose descriptor:

EC10

Effect conc.:

17 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

added B

Basis for effect:

reproduction

Remarks:

number of juveniles

Reported statistics and error estimates:

Data were checked for normality by Shapiro-Wilk´s test on normal distribution and for homogeneity by Cochran’s test. Treatment means were compared by ANOVA and Welsh-t test (p ≤ 0.05, one-sided) and tested for statistically significant differences compared to the control.
Probit analysis using Linear Max. Likelihood Regression was used to calculate the EC50, EC20 and EC10 for reproduction.

28 d NOEC for reproduction: 23.2 mg B/kg soil dw (26.2% inhibition)

28 d LOEC for reproduction: 31.1 mg B/kg soil dw (39.7% inhibition)

28 d EC50 for reproduction: 39.9 mg B/kg soil dw

28 d EC20 for reproduction: 22.7 mg B/kg soil dw

28 d NOEC for mortality: >174.8 mg B/kg soil dw

Concentration [mg test item/kg soil (dw)]	Number of Juveniles[mean ± sd]	Number of Juveniles[% of control]
Control	937.5± 191.9	-
10	875.5± 119.5	93.4
32	1024.5± 102.5	109.3
56	1214.5± 211.4	129.5
75	1268.0± 414.4	135.3
100	876.0± 77.8	93.4
133	701.0± 1.4	74.8
178	565.0± 41.0	60.3
237	400.0± 11.3	42.7
316	437.0± 171.1	46.6
422	112.5± 50.2	12.0
562	43.5± 54.5	1.0
1000	38.0± 31.1	4.1

Validity criteria fulfilled:

yes

Conclusions:

This reliable study was performed according to the OECD 220 guideline and is well documented. A 28-d EC10 value for reproduction of Enchytraeus luxuriosus of 17 mg B/ kg soil dw is obtained and is used fpr PNEC derivation.

Reference 6

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was performed according to OECD 220 (2004) guideline and is well documented.

Qualifier:

according to guideline

Guideline:

OECD Guideline 220 (Enchytraeid Reproduction Test)

Deviations:

no

GLP compliance:

yes

Remarks:

The study has no formal GLP certificate. However, the study was conducted at a GLP-certified laboratory, staff was appropriately trained and standard operating procedures were followed in terms of experimental performance and raw data documentation.

Analytical monitoring:

no

Vehicle:

no

Details on preparation and application of test substrate:

- The test item was dissolved in a sufficient amount of deionised water to prepare a stock solution. This stock solution was diluted with deionised water to obtain one test item solution (= dosage solution) for each test item concentration.
- Method of mixing into soil (if used): The test item was incorporated into the soil by thoroughly mixing in the respective test item
solution for approx. 5 minutes.
- This amount of water/test item-mixture served also to adjust the final soil moisture of 40-60% of WHCmax.

Test organisms (species):

other: Enchytraeus crypticus

Animal group:

other: Enchytraeidae

Details on test organisms:

Enchytraeus crypticus (Enchytraeidae, potworms), has been cultured at ECT Oekotoxikologie GmbH since September 2003.
The worms used in the tests were adult (with eggs, i.e. white spots, in the clitellum region).

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

28 d

Test temperature:

19-21 °C

pH:

soil pH-value: 5.9

Moisture:

soil moisture: 40.0-60.0% of the WHCmax

Details on test conditions:

TEST SYSTEM
- Test container (material, size): Test vessels were glass containers (able to be closed tightly) of about 200 mL capacity and with a diameter of about 5 cm.
- Amount of soil or substrate: 20g dry weight
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 2
- No. of replicates per control: 6

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- artificial soil based on OECD guideline 207 (1984). Percentages refer to dry weight:
- 10% Sphagnum peat (no visible plant remains; finely ground and air-dried)
- 20% Kaolin-Clay (kaolinite content > 30%)
- 69.6% Quartz sand (fine sand content with particles between 50 and 200 µm higher than 50%)
- 0.4% Calcium carbonate (CaCO3)
- The moisture content was adjusted to 40 – 60% of WHCmax using deionised water.

OTHER TEST CONDITIONS
- Photoperiod: 16 : 8 h light/dark with a constant light intensity at the substrate surface of 400 – 800 lx
- Feeding: The enchytraeids were fed with about 50 mg of ground oats per test vessel at the beginning of the test (mixed with the test substrate after
application of the test item). The organisms were fed an additional 25 mg ground oats once per week until day 21.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Mortality: mean number of dead adult worms as a percentage of the initial number at the start of the test for each concentration and for the control. (Validity criteria for mortality not fulfilled)
- Reproduction: mean number of juveniles per test vessel 28 days after application.

Nominal and measured concentrations:

nominal added concentrations: control, 1.0, 10, 32, 100, 133, 178, 237, 316, 422, 562, 750 and 1000 mg H3BO3/kg dry weight soil,
corresponding to control, 0.17, 1.75, 5.6, 17.5, 23.2, 31.1, 41.4, 55.2, 73.8, 98.2, 131.1 and 174.8 mg B/kg dw

Reference substance (positive control):

no

Duration:

28 d

Dose descriptor:

EC10

Effect conc.:

22.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

added B

Basis for effect:

reproduction

Remarks:

number of juveniles

Reported statistics and error estimates:

Data were checked for normality by Shapiro-Wilk´s test on normal distribution and for homogeneity by Cochran’s test. Treatment means were compared by ANOVA and Welsh-t test (p ≤ 0.05, one-sided) and tested for statistically significant differences compared to the control.
Probit analysis using Linear Max. Likelihood Regression was used to calculate the EC50, EC20 and EC10 for reproduction.

28 d NOEC for reproduction: 23.2 mg B/kg soil dw (5.2% inhibition)

28 d LOEC for reproduction: 31.1 mg B/kg soil dw (33.1% inhibition)

28 d EC50 for reproduction: 38.5 mg B/kg soil dw

28 d EC20 for reproduction: 27.1 mg B/kg soil dw

28 d NOEC for mortality: >174.8 mg B/kg soil dw

Concentration [mg test item/kg soil (dw)]	Number of Juveniles[mean ± sd]	Number of Juveniles[% of control]
Control	628.7± 92.8	-
1.0	638.0± 148.5	101.5
10	677.5± 87.0	107.8
32	778.0± 48.1	123.8
100	690.0± 200.8	109.8
133	596.0± 15.6	94.8
178	420.5± 43.1	66.9
237	228.0± 29.7	36.3
316	162.0± 8.5	25.8
422	47.5± 4.9	7.6
562	17.5± 9.2	2.8
750	6.0± 1.4	1.0
1000	1.0± 1.4	0.2

Validity criteria fulfilled:

yes

Conclusions:

This reliable study was performed according to the OECD 220 guideline and is well documented. A 28-d EC10 value for reproduction of Enchytraeus crypticus of 22.6 mg B/ kg soil dw is obtained and used for PNEC derivation.

Reference 7

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was performed according to OECD 222 (2004) and ISO 11268-2 Part 2 (1998) guidelines and is well documented.

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

GLP compliance:

yes

Remarks:

The study has no formal GLP certificate. However, the study was conducted at a GLP-certified laboratory, staff was appropriately trained and standard operating procedures were followed in terms of experimental performance and raw data documentation.

Analytical monitoring:

no

Vehicle:

no

Details on preparation and application of test substrate:

- The test item was dissolved in a sufficient amount of deionised water to prepare a stock solution. This stock solution was diluted with deionised water to obtain one test item solution (= dosage solution) for each test item concentration.
- Method of mixing into soil (if used): The test item was incorporated into the soil by thoroughly mixing in the respective test item solution for approx. 5 minutes.

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

The earthworms used in this test, Eisenia fetida (Lumbricidae, Earthworms) are kept at ECT Oekotoxikologie GmbH since February 1994.
Only adult worms (with clitellum) with a fresh weight (FW) between 300 and 600 mg were used. The worms were at least two months, but not more than one year old, however the age of individuals did not differ by more than four weeks. Animals which do fulfil these requirements were taken from a synchronised culture. The worms selected for the test were acclimatised in artificial soil under test conditions at least 24 hours before starting the test.

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

28 d

Remarks:

56 days for reproduction endpoint

Test temperature:

19-21 °C

pH:

soil pH-value: 5.9

Moisture:

soil moisture: 40-60% of WHCmax

Details on test conditions:

TEST SYSTEM
- Test container (material, size): Test vessels were Bellaplast containers consisting of inert (non-toxic) plastic (Polystyrol). The test vessels have a bottom area of 11 x 15.5 cm (=170.5 cm²) and a height of 6 cm. The test vessels were covered by a transparent lid.
- Amount of soil or substrate: 500 g dry weight
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 2
- No. of replicates per control: 6

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- artificial soil based on OECD guideline 207 (1984). Percentages refer to dry weight:
- 5% Sphagnum peat (no visible plant remains; finely ground and air-dried)
- 20% Kaolin-Clay (kaolinite content > 30%)
- 74% Quartz sand (fine sand content with particles between 50 and 200 µm higher than 50%)
- ca. 1% Calcium carbonate (CaCO3)
- The moisture content was adjusted to 40 – 60% of WHCmax using deionised water.

OTHER TEST CONDITIONS
- Photoperiod: 16 : 8 h light/dark with a constant light intensity at the substrate surface of 400 – 800 lx
- Feeding: The adult worms were fed with finely ground cow manure (free of growth promoters, nematicides or similar veterinary pharmaceuticals,
). Before starting the experiment an amount of 5 g food per 500 g soil dry weight was mixed into the artificial soil to be used in the test. After the start of the experiment food was first provided one day after application of the test item and introduction of the adult worms. Thereafter, the adult worms were fed weekly during the first 4 weeks of the test. An amount of 5 g moistened with 10 mL deionised water was spread on the soil surface of each test container. If food remains uneaten the ration was reduced on demand. After removing of the adults on day 28 further 5 g of food was mixed with the artificial soil of each test vessel.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Mortality: mean number of dead adult worms as a percentage of the initial number at the start of the test for each concentration and for the control.
- Biomass: loss/increase in the mean biomass (i.e. the fresh mass of organisms) 28 days after application expressed as a percentage of the initial fresh mass of organisms at the start of the test for each concentration and for the control.
- Reproduction: mean number of juveniles per test vessel 56 days after application.

Nominal and measured concentrations:

nominal added concentrations: control, 75, 100, 133, 178, 237, 316, 422, 562, 750 and 1000 mg H3BO3/kg dry weight soil,
corresponding to control, 13.1, 17.5, 23.2, 41.4, 55.2, 73.8, 98.2, 131.1 and 174.8 mg B/kg dw

Reference substance (positive control):

no

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 174.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

added B

Basis for effect:

mortality

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 174.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

added B

Basis for effect:

growth

Remarks:

biomass production

Duration:

56 d

Dose descriptor:

EC10

Effect conc.:

70.1 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

added B

Basis for effect:

reproduction

Remarks:

number of juveniles

Reported statistics and error estimates:

Mortality and Biomass: No statistical analysis was performed.

Reproduction: Data were checked for normality by Shapiro-Wilk´s test on normal distribution and for homogeneity by Cochran’s test. Treatment means were compared by ANOVA followed by Dunnett’s t-test (p ≤ 0.05, according to the guideline two-sided for biomass and one-sided for reproduction) and tested for statistically significant differences compared to the control. Logit analysis (2-parameter-logistic function) using linear maximum likelihood regression was used to calculate the EC50, EC20 and EC10 for reproduction.

56 d EC50 for reproduction: 84.6 mg B/kg soil dw

56 d EC20 for reproduction: 75.2 mg B/kg soil dw

56 d NOEC for reproduction: 73.8 mg B/kg soil dw (16.6% inhibition)

56 d LOEC for reproduction: 98.2 mg B/kg soil dw (85.3% inhibition)

Concentration [mg test item/kg soil (dw)]	Number of dead adultsDay 28	Mortality (%)Day 28
Control	0	0
75	0	0
100	2	10
133	0	0
178	0	0
237	0	0
316	0	0
422	0	0
562	0	0
750	0	0
1000	0	0

Concentration [mg test item/kg soil (dw)]	Number of Juveniles[mean ± sd]	Number of Juveniles[% of control]
Control	357.3 +- 45.2	100.0
75	320.0 +- 7.1	89.6
100	431.0 +- 80.6	120.6
133	454.0 +- 15.6	127.1
178	406.5 +- 19.1	113.8
237	446.0 +- 50.9	124.8
316	418.0 +- 75.0	117.0
422	298.0 +- 21.2	83.4
562	52.5 +- 9.2	14.7
750	1.0 +- 1.4	0.3
1000	0.0 +- 0.0	0.0

Concentration [mg test item/kg soil (dw)]	Biomass (mg fresh weight)Day 0	Biomass (mg fresh weight)Day 28	% of initial weightDay 28
Control	418.7±65.7	624.3±106.3	149.1
75	427.0±66.0	660.9±137.1	154.8
100	416.2±75.2	591.7±227.2	142.2
133	418.2±58.3	672.4±166.4	160.8
178	421.0±73.9	656.7±98.1	156.0
237	433.2±65.7	680.3±116.3	157.0
316	424.4±45.5	708.6±99.2	167.0
422	416.5±80.0	655.6±107.4	157.4
562	416.3±71.4	639.8±125.9	153.7
750	431.1±61.2	615.6±89.3	142.8
1000	423.6±62.0	484.8±56.4	114.4

Validity criteria fulfilled:

yes

Conclusions:

This reliable study was performed according to the OECD 222 guideline and is well documented. A 56 days EC10 value for reproduction of Eisenia fetida of 70.1 mg B/ kg soil dw is obtained and used for PNEC derivation.

Reference 8

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: according to national guideline (Environment Canada), standard test and selection criteria fulfilled

Qualifier:

according to guideline

Guideline:

other: Environment Canada (2002)

Deviations:

not specified

GLP compliance:

not specified

Analytical monitoring:

not specified

Vehicle:

not specified

Details on preparation and application of test substrate:

Test compound added in solution to soil. Soil moisture was 35% (based on wet weight). Tests started after a 1 day equilibration period.

Test organisms (species):

Eisenia sp.

Animal group:

annelids

Details on test organisms:

Eisenia andrei
- fully developed clitellum
- aged 2-12 months
- individual wet weight 250-600 mg

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

63 d

Post exposure observation period:

not reported

Test temperature:

20°C

pH:

6.09

Moisture:

35% (based on wet weight)

Details on test conditions:

TEST SYSTEM
- Test container (material, size): 500 ml
- Amount of soil or substrate: 350 ml (200 g)
- No. of organisms per container (treatment): 2
- No. of replicates per treatment group: 10
- No. of replicates per control: 10


SOURCE AND PROPERTIES OF ARTIFICIAL SOIL
- Ingredients: 70% silica sand, 20% kaolinite clay, 10% spaghnum peat, 1-3g calcium carbonate per kg soil
- Soil texture
- % sand: 77.3
- % silt: 7.8
- % clay: 14.9
- Total carbon (%): 4.46
- Organic matter (%): 9
- pH water (1:2): 6.09
- eCEC: 18.5 cmol c/kg
- background B concentration: 3.5 mg B/kg dw

OTHER TEST CONDITIONS
- Photoperiod: 16h light, 8h dark

Nominal and measured concentrations:

0-7-14-28-56-113-225-450-900-1800-3600 mg boric acid/kg dw,
corresponding to 0-1.2-2.5-4.9-9.8-19.8-39.4-78.8-315-630 mg B/kg dw

Reference substance (positive control):

not specified

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

315 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: asynchronous cultures

Duration:

63 d

Dose descriptor:

NOEC

Effect conc.:

39.4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(juvenile production)

Remarks on result:

other: asynchronous cultures

Duration:

63 d

Dose descriptor:

NOEC

Effect conc.:

78.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Remarks:

(juvenile dry weight)

Remarks on result:

other: asynchronous cultures

Duration:

63 d

Dose descriptor:

NOEC

Effect conc.:

78.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching success)

Remarks on result:

other: asynchronous cultures

Duration:

63 d

Dose descriptor:

NOEC

Effect conc.:

19.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching failure)

Remarks on result:

other: asynchronous cultures

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

315 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: synchronous culture

Duration:

63 d

Dose descriptor:

NOEC

Effect conc.:

78.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(juvenile production)

Remarks on result:

other: synchronous culture

Duration:

63 d

Dose descriptor:

NOEC

Effect conc.:

78.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Remarks:

(juvenile dry weight)

Remarks on result:

other: synchronous culture

Duration:

63 d

Dose descriptor:

NOEC

Effect conc.:

78.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching success)

Remarks on result:

other: synchronous culture

Duration:

63 d

Dose descriptor:

NOEC

Effect conc.:

>= 78.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching failure)

Remarks on result:

other: synchronous culture

Duration:

28 d

Dose descriptor:

LC50

Effect conc.:

589.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: asynchronous cultures

Duration:

63 d

Dose descriptor:

EC50

Effect conc.:

77.4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(juvenile production)

Remarks on result:

other: asynchronous cultures

Duration:

63 d

Dose descriptor:

EC50

Effect conc.:

95.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Remarks:

(juvenile dry weight)

Remarks on result:

other: asynchronous cultures

Duration:

63 d

Dose descriptor:

EC50

Effect conc.:

120.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching success)

Remarks on result:

other: asynchronous cultures

Duration:

28 d

Dose descriptor:

EC50

Effect conc.:

620 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

mortality

Remarks on result:

other: synchronous culture

Duration:

63 d

Dose descriptor:

EC50

Effect conc.:

99.4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(juvenile production)

Remarks on result:

other: synchronous culture

Duration:

63 d

Dose descriptor:

EC50

Effect conc.:

165.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Remarks:

(juvenile dry weight)

Remarks on result:

other: synchronous culture

Duration:

63 d

Dose descriptor:

EC50

Effect conc.:

86.3 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching success)

Remarks on result:

other: synchronous culture

Duration:

63 d

Dose descriptor:

EC50

Effect conc.:

172.1 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching failure)

Remarks on result:

other: synchronous culture

Details on results:

nominal added dosis; background concentration reported

Reported statistics and error estimates:

ECx values and their associated 95% confidence limits for the chronic tests were analysed by using either a linear or four nonlinear regression models (i.e., logistic, Gompertz, exponential and logistic with hormoesis). If data showed heteroscedascity among treatments, data were weighed with the inverse of the variance of each treatment.
NOEC values were estimated using ANOVA, Dunnett's and Fisher's protected LSD (least significant difference) pairwise comparison test.

Validity criteria fulfilled:

yes

Conclusions:

This study followed the Environment Canada guideline.
The standard test and selection criteria were fulfilled. Reliable NOEC values are used for PNEC derivation.

Reference 9

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: according to THE OECD guideline or national guideline (Environment Canada), standard test and selection criteria fulfilled.

Qualifier:

according to guideline

Guideline:

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

Deviations:

not specified

Qualifier:

according to guideline

Guideline:

other: Environment Canada (2002)

Deviations:

not specified

GLP compliance:

not specified

Analytical monitoring:

not specified

Vehicle:

not specified

Details on preparation and application of test substrate:

Test compound added in solution to soil. Soil moisture was 35% (based on wet weight). Tests started after a 1 day equilibration period.

Test organisms (species):

Eisenia sp.

Animal group:

annelids

Details on test organisms:

Eisenia andrei
- fully developed clitellum
- aged 2-12 months, synchronized
- individual wet weight 250-600 mg

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

56 d

Test temperature:

20°C

pH:

6.09

Moisture:

35% (based on wet weight)

Details on test conditions:

TEST SYSTEM
Environment Canada guideline:
- Test container (material, size): 500 ml
- Amount of soil or substrate: 350 ml (200 g)
- No. of organisms per container (treatment): 2
- No. of replicates per treatment group: 10
- No. of replicates per control: 10

OECD guideline:
- Test container (material, size): 1-2 L
- Amount of soil or substrate: 500-600 g dry weight
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 4
- No. of replicates per control: 8

SOURCE AND PROPERTIES OF ARTIFICIAL SOIL
- Ingredients: 70% silica sand, 20% kaolinite clay, 10% spaghnum peat, 1-3g calcium carbonate per kg soil
- Soil texture
- % sand: 77.3
- % silt: 7.8
- % clay: 14.9
- Total carbon (%): 4.46
- Organic matter (%): 9
- pH water (1:2): 6.09
- eCEC: 18.5 cmol c/kg
- background B concentration: 3.5 mg B/kg dw

OTHER TEST CONDITIONS
- Photoperiod: 16h light, 8h dark

Nominal and measured concentrations:

0-10-16-30-50-100-300-560-1000 mg boric acid/kg dw,
corresponding to 0-1.75-2.8-5.25-8.75-17.5-52.5-98-175 mg B/kg dw.

Reference substance (positive control):

not specified

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

52.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(juvenile production)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

5.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Remarks:

(juvenile dry weight)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

52.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching success)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

52.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching failure)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

98 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(cocoon production)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

77.9 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(juvenile production)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

25.7 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Remarks:

(juvenile dry weight)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

98 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching success)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

136.2 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(cocoon production)

Remarks on result:

other: Environment Canada guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

52.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(juvenile production)

Remarks on result:

other: OECD guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

52.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Remarks:

(juvenile dry weight)

Remarks on result:

other: OECD guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

52.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching success)

Remarks on result:

other: OECD guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

52.5 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching failure)

Remarks on result:

other: OECD guideline

Duration:

56 d

Dose descriptor:

NOEC

Effect conc.:

98 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(cocoon production)

Remarks on result:

other: OECD guideline

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

75.7 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(juvenile production)

Remarks on result:

other: OECD guideline

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

59.4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

growth

Remarks:

(juvenile dry weight)

Remarks on result:

other: OECD guideline

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

90.4 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(hatching success)

Remarks on result:

other: OECD guideline

Duration:

56 d

Dose descriptor:

EC50

Effect conc.:

107.6 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

element

Remarks:

B

Basis for effect:

reproduction

Remarks:

(cocoon production)

Remarks on result:

other: OECD guideline

Details on results:

No significant effect on mortality up to the largest dose tested (175 mg B/kg dw).

Reported statistics and error estimates:

ECx values and their associated 95% confidence limits for the chronic tests were analysed by using either a linear or four nonlinear regression models (i.e., logistic, Gompertz, exponential and logistic with hormoesis). If data showed heteroscedascity among treatments, data were weighed with the inverse of the variance of each treatment.

Validity criteria fulfilled:

yes

Conclusions:

This study followed the OECD guideline or a national guideline (Environment Canada)
The standard test and selection criteria were fulfilled.
Reliable NOEC values are used for PNEC derivation.

Description of key information

In total 36 individual high quality NOEC/EC10 values (for 9 different species) are selected for the PNEC derivation, ranging from 5.2 mg B/kg dw for Eisenia andrei growth to 315 mg B/kg for Eisenia andrei survival (Stantec Consulting & Aquaterra Environmental Consulting, 2003).

Please also refer to read-across statement section 13.

Additional information

All selected NOEC/EC₁₀ values from chronic single-species toxicity tests with invertebrates are summarised in the respective table listing all effects to soil macroorganisms. All these values are considered reliable (Klimisch score 1 and 2) and relevant for the effects assessment. All data in the respective table are based on added nominal values as background concentrations are not always available and the added risk approach will be followed because of the significant difference in bioavailability between boron naturally present in soils and added soluble B. In total 36 individual high quality NOEC/EC₁₀ values (for 9 different species) are selected for the PNEC derivation, ranging from 5.2 mg B/kg dw for Eisenia andrei growth to 315 mg B/kg for Eisenia andrei survival (Stantec Consulting & Aquaterra Environmental Consulting, 2003).

Boric acid and borates have been registered as pesticides effective against a number of terrestrial insects, for example cockroaches, termites and ants. However the studies that establish efficacy as a pesticide are not relevant for use in describing ecotoxicological responses from boron present in soil because these pesticide studies typically involve use of a specially formulated diet or bait or treated article.

 

For example, Durmuş and Büyükgüzel (2008) reported effects on survival and development of larvae of the Greater Wax Moth, Galleria mellonella, when fed artificial diets containing 0.2 % sodium tetraborate. No effects were observed when fed 0.1 % sodium tetraborate. Yang et al. (2000) reported reduction in survival and reproduction of the Caribbean fruit fly Anastrepha suspensa when fed 0.1 to 0.5 % sodium tetraborate. Since the diets were not soil-based (as might occur for an earthworm), such studies cannot be relied upon for derivation of a PNECsoil. The critical boron concentrations in food (approx. 0.1 % or 1000 mg B/kg) are orders of magnitude above the soil effects concentrations of boron for invertebrates (5-300 mg B/kg). Because boron does not magnify through the food-chain and soil to plant bioaccumulation factors for boron derived from tests performed in real soils are generally < 100 and internal boron concentrations in plants are largely controlled by plant-specific requirements for this essential element, it can be concluded that the secondary poisoning of insects through plant material is not critical for boron.