Lime (chemical), hydraulic

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:

supporting study

Study period:

June 26, 2007 - July 10, 2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

14 d

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

> 5 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

Ca(OH)2

Basis for effect:

mortality

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

>= 5 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

Ca(OH)2

Basis for effect:

other: biomass

Validity criteria fulfilled:

yes

Remarks:

Adult mortality: ≤ 10% (being 0% after 14 days).

Conclusions:

After 14 days of exposure to Calcium dihydroxide in artificial soil the LC50 for Eisenia fetida was estimated to be higher than 5000 mg Calcium dihydroxide/kg soil d.w. The NOEC was determined to be 5000 mg Calcium dihydroxide/kg soil d.w.

Reference 2

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

experimental study

Adequacy of study:

key study

Study period:

August 16, 2007 - October 11, 2007

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))

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Details on sampling:

Not applicable.

Vehicle:

no

Details on preparation and application of test substrate:

- approx. 24 h before test start, artificial soil was prepared
- deionized water was added to dry components to adjust water content to approx. 40 - 60 % of WHC
- test item incorporated directly into the soil by mixing with a small quantity of finely ground quartz sand (10 g of vessel), such that the required test concentration was achieved once mixed with the artificial soil

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

- Eisenia fetida andrei
- source: original breeding animals were purchased from "W. Neudorff GmbH KG", An der Mühle 3, D-31860 Emmertal
- test animals: reared under ambient laboratory conditions in the test facility
- age of animals used in the test: adult worms (approximately 3 months old with clitellum)
- weight of animals used in the test: 313 - 466 mg/worm
- feeding: weekly during first 4 weeks, by adding ground hourse manure on the soil surface (last feeding at 4 weeks)

ACCLIMATION
- period: approx. 24 h in artificial soil (with food)

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

no

Total exposure duration:

4 wk

Remarks:

4 weeks for adult mortality

Post exposure observation period:

4 weeks for juvenile development

Test temperature:

18-22 °C

pH:

test start: 6.23 - 8.17
test end: 6.05 - 7.71

Moisture:

test start: 34.8-35.0 %
test end: 34.7-35.1 %

Details on test conditions:

- light conditions: light/dark 16/8 h, artificial light (590 lx)
- food: air-dried and ground horse manure (5 g at test start)
- feeding frequency/amount: weekly, amount of manure depended on feeding activity
- test vessels: plastic, 18.3 x 13.6 x 6.4 cm, with lid pervious to air and light
- number of animals/vessel (= replicate): 10
- number of replicates/control group: 8
- number of replicates/treated group: 4
- number of animals/control group: 80
- number of animals/treated group: 40
- amount of soil/test vessel: 556 g dry weight (corresponding to approx. 750 g moist soil)
- total carbon content (%): 4.51 %
- total organic carbon content (%): 4.47 %
- light conditions:
* source: artificial light
* intensity: 590 lx
* duration: 16:8 light:dark

Nominal and measured concentrations:

nominal concentrations: 0, 250, 500, 1000, 2000, 4000 and 5000 mg test item/kg soil d.w.

Reference substance (positive control):

yes

Remarks:

Benlate

Duration:

4 wk

Dose descriptor:

NOEC

Effect conc.:

2 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

Ca(OH)2

Basis for effect:

reproduction

Duration:

4 wk

Dose descriptor:

EC50

Effect conc.:

4 180 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

Ca(OH)2

Basis for effect:

reproduction

Remarks on result:

other: 95% CL: 3346-4665 mg/kg soil d.w.

Duration:

4 wk

Dose descriptor:

NOEC

Effect conc.:

5 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

Ca(OH)2

Basis for effect:

mortality

Details on results:

Validity criteria for control group:
- adult mortality after 4 weeks: <= 10 %
- number of juveniles per replicate: >= 30
- coefficient of variation of reproduction: <= 30%

- Mortality at end of exposure period: 0% in all dose groups
- Total mass of adults at beginning of test: mean between 378.1 and 384.1 mg
- Changes in body weigth of live adults (% of initial weight) at end of exposure period: 31.8% (control), 29.1% (250 mg/kg), 28.8% (500 mg/kg), 32.8% (1000 mg/kg), 29.4% (2000 mg/kg), 26.8% (4000 mg/kg), 22.6% (5000 mg/kg)
- No. of offspring produced: mean numbers:129.5 (control), 123 (250 mg/kg), 126.5 (500 mg/kg), 125.3 (1000 mg/kg), 118 (2000 mg/kg), 77.3 (4000 mg/kg), 32.8 (5000 mg/kg)

Results with reference substance (positive control):

- Results with reference substance valid? Yes
- Relevant effect levels: The number of juveniles was reduced by 67% and 100% compared to control at concentrations of 5 and 10 mg Benlate/kg soil d.w., respectively.

Reported statistics and error estimates:

- probit analysis according to the maximum likelihood methods was used for the calculation of EC50 (software ToxRat Professional 2.09)

Validity criteria fulfilled:

yes

Remarks:

Adult mortality: ≤ 10% (being 0% after 4 weeks). Number of juveniles per replicate: ≥ 30 (being 158, 144, 97, 128, 119, 135, 109 and 146). Coefficient of variation of variation of reproduction: ≤ 30% (being 15.8%).

Conclusions:

Calcium dihydroxide did not show significant adverse effects on survival and growth of the earthworm Eisenia fetida in artificial soil up to 5000 mg Calcium dihydroxide/kg soil dry weight, i.e. the highest rate tested.
The test item showed statistically significantly adverse effects on reproduction at 4000 and 5000 mg Calcium hydroxide/kg soil d.w. Therefore, the NOEC for reproduction was determined to be 2000 mg Calcium dihydroxide/kg soil dry weight. The EC50 for number of juveniles was calculated to be 4180 mg Calcium dihydroxide/kg soil dry weight.

Reference 3

Endpoint:

toxicity to soil macroorganisms except arthropods: short-term

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

6 January 2010 - 11 February 2010

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 207 (Earthworm, Acute Toxicity Tests)

Deviations:

yes

Remarks:

Some of the temperatures were observed to be slightly in excess of the 20 ± 2 °C given in the study plan. However, this deviation was considered not to have affected the outcome or the validity of the test.

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Details on sampling:

Not applicable

Vehicle:

no

Details on preparation and application of test substrate:

- Method of mixing into soil: The test item was prepared by direct addition to the basic soil substrate. An amount of test item (3500 mg) was added to approximately 100 g premix of artificial soil and carefully mixed with a spatula until homogenous. This premix was then incorporated into 3.5 kg (dry weight) of artificial soil with 1000 mL of deionised reverse osmosis water and mixed using a Hobart A200N mixer.
- Controls: The control was prepared in an identical manner using 2.5 kg (dry weight) of artificial soil and 710 mL of deionised reverse osmosis water, without the addition of test item.

Test organisms (species):

Eisenia fetida

Animal group:

annelids

Details on test organisms:

TEST ORGANISM
- Common name: Earthworms
- Source: Original Organics Limited, Langlands Business Park, Devon, UK
- Age at test initiation (mean and range, SD): at least 2 months old with clitellum
- Weight at test initiation (mean and range, SD): 0.44 - 0.52 g (mean 0.49 g)


ACCLIMATION
- Acclimation period: The earthworms were transferred into artificial soil (without test item) approximately 24 hours prior to the start of the test.

Study type:

laboratory study

Substrate type:

artificial soil

Limit test:

yes

Total exposure duration:

14 d

Post exposure observation period:

Earthworms were observed for up to 14 days

Test temperature:

19 to 23 °C

pH:

Test initiation: 5.8 - 7.1
Test termination: 6.3 - 6.6

Moisture:

The actual moisture content of the soil used in the definitive test was 29% of dry weight of basic substrate on Day 0 and 23% and 24% on Day 14.

Details on test conditions:

TEST SYSTEM
- Test container (material, size): 1 L glass beakers were used
- Amount of soil or substrate: The test vessels contained approximately 3.5 kg (dry weight) of artificial soil and the control vessels contained 2.5 kg.
- No. of organisms per container (treatment): 10 earthworms per test and control vessel
- No. of replicates per treatment group: 6
- No. of replicates per control: 4


SOURCE AND PROPERTIES OF SUBSTRATE
- Composition of the artificial soil:
Industrial quartz sand - 70% w/w
Kaolinite clay - 20% w/w
Sphagnum moss peat - 10% w/w
- Maximum water holding capacity (in % dry weight): Nominal: 30% (this level of moisture has been shown to ensure that a good crumb structure is formed within the soil).
- Stability and homogeneity of test material in the medium: No analysis was conducted to determine the homogeneity, concentration or stability of the test item formulation.


OTHER TEST CONDITIONS
- Photoperiod: Continuous illumination
- Light intensity: 684 to 714 lux


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : The weight of each earthworm was recorded on Day 0 before addition to the test or control vessels. Each surviving earthworm was weighed on Day 14.
Any mortalities were recorded 7 and 14 days after the start of the treatment and any sub-lethal effects were recorded daily. The criterion of death was taken to be the absence of any reaction to a physical stimulus at either end of the body.


TEST CONCENTRATIONS
- Range finding study
- Test concentrations: Nominal concentrations: 100 and 1000 mg/kg
- Results used to determine the conditions for the definitive study: The results showed no mortalities at the test concentrations of 100 and 1000 mg/kg.

Nominal and measured concentrations:

Definitive test:
Nominal concentration: 1000 mg/kg of soil

Reference substance (positive control):

yes

Remarks:

Chloroacetamide (nominal concentrations 5.6, 10, 18, 32 and 56 mg/kg)

Duration:

14 d

Dose descriptor:

LC50

Effect conc.:

> 1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Duration:

14 d

Dose descriptor:

NOEC

Effect conc.:

1 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: mortality and sub-lethal effects

Details on results:

- Mortality at end of exposure period: There were no significant mortalities or sub-lethal effects of exposure in 60 earthworms exposed to a test concentration of 1000 mg/kg for a period of 14 days. Two worms are unaccounted for on Day 14 in the 1000 mg/kg test concentration. This was considered to be possibly due to natural causes rather than a toxic effect given that this constituted less than 5% of the total number of earthworms exposed to this concentration and no other effects were observed (see table 1).
- Weight at end of exposure period: There were no significant differences between the control and the 1000 mg/kg test groups in terms of weight on Days 0 and 14 (see table 2).

Results with reference substance (positive control):

- Results with reference substance: The results from the positive control with chloroacetamide are within the normal range for this reference item of 10 to 50 mg/kg.
- Relevant effect levels: 7 day LC50: 44 mg/kg (95% confidence limits 42-45 mg/kg); 14 day LC50: 43 mg/kg (95% confidence limits 41-45 mg/kg); NOEC: 18 mg/kg

Table 1: Cumulative mortality data in the definitive test

Nominal concentration (mg/kg)	Cumulative mortality (initial population: 10)		% mortality
	Day 0		Day 14
Control	-	-	-	-
R1	0	0	0	0
R2	0	0	0	0
R3	0	0	0	0
R4	0	0	0	0
1000	-	-	-	-
R1	0	0	0	0
R2	0	1*	0	10
R3	0	0	0	0
R4	0	0	0	0
R5	0	0	0	0
R6	0	1*	0	10

R1-R6 = Replicates 1 to 6

* Single mortality observed was considered to be possibly due to natural causes and was considered not to affect the validity of the test given that less than 5% mortality occurred at this concentration.

 

 

Table 2: Mean earthworm weights in the definitive test

Nominal concentration (mg/kg)	Mean earthworm weight (g)
	Day 0		Day 14
	Mean	SD	Mean	SD
Control	-	-	-	-
R1	0.51	0.09	0.51	0.09
R2	0.52	0.08	0.49	0.10
R3	0.51	0.08	0.47	0.10
R4	0.49	0.05	0.51	0.10
1000	-	-	-	-
R1	0.47	0.07	0.49	0.06
R2	0.50	0.08	0.51	0.11
R3	0.50	0.09	0.49	0.09
R4	0.51	0.07	0.53	0.09
R5	0.49	0.09	0.47	0.09
R6	0.44	0.08	0.48	0.09

R1-R6 = Replicates 1 to 6

SD = Standard deviation

 

Validity criteria fulfilled:

yes

Remarks:

Mortality of control animals < 10%

Conclusions:

The acute toxicity of the test item, to the earthworm Eisenia foetida, in an artificial soil test gave a 14 day LC50 value of >1000 mg/kg and a NOEC of 1000 mg/kg.

Reference 4

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Justification for type of information:

JUSTIFICATION FOR DATA WAIVING
According to Section 9.4.4 of REACH Regulation 1907/2006, long term toxicity testing on terrestrial invertebrates (except arthropods) is required unless already provided as part of Annex IX requirements. However, such a study is not applicable to calcium carbonate for the following reasons: Calcium carbonate and calcium and carbonate ions are ubiquitous in the environment and are found naturally in soil, water and sediment. Calcium is an important constituent of most soils and the minerals found in soil are mostly compounds of calcium with other substances. Furthermore, calcium carbonate is used as an inorganic fertiliser to improve soil quality, both pH and structure and guarantees an adequate supply of calcium to plants. Furthermore, an acute toxicity study to earthworms was performed according to OECD 207 as a limit test with a calcium carbonate (nano) concentration of 1000 mg/kg soil (Goodband, 2010). There were no significant mortalities or sub-lethal effects of exposure in 60 earthworms exposed to calcium carbonate (nano) for a period of 14 days. As a result, calcium carbonate is considered not acutely toxic to earthworms and hence long term testing is considered to be unnecessary. Given the extensive and continued use of calcium carbonate as a constituent of fertiliser and its natural occurrence in the environment, it is considered that calcium carbonate would not have a detrimental effect on soil macroorganisms. Therefore, the performance of long term toxicity tests on terrestrial organisms is scientifically unjustified.

Reference 5

Endpoint:

toxicity to soil macroorganisms except arthropods: long-term

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

REPORTING FORMAT FOR THE ANALOGUE APPROACH

1. HYPOTHESIS FOR THE ANALOGUE APPROACH
Common functional groups/mechanism of action.

2. SOURCE AND TARGET CHEMICAL(S) (INCLUDING INFORMATION ON PURITY AND IMPURITIES)
Target: Lime (chemical), hydraulic [CAS 85117-09-5; See section 1.2 for information on purity.
Source: calcium dihydroxide [CAS 1305-62-0; EC 215-137-3] 98.2%

3. ANALOGUE APPROACH JUSTIFICATION
In the environment, lime substances rapidly dissociate or react with water. These reactions, together with the equivalent amount of hydroxyl ions set free when considering 100mg of the lime compound (hypothetic example), are illustrated below:
Ca(OH)2 <-> Ca2+ + 2OH-
100 mg Ca(OH)2 or 1.35 mmol sets free 2.70 mmol OH-
Ca(OH)2 + Ca2SiO4 + CaCO3 + 3 H2O <-> 4Ca2+ + SiO2 + CO2 + 8OH-
100 mg lime (chemical) hydraulic or 0.27 mmol sets free 2.16 mmol OH-
It has to be noted that CO32- is not expected to directly release two hydroxyl ions under most environmental conditions (depends on CO2 concentrations and pH) and this is therefore a worst case assumption.
From these reactions it is clear that the effect of lime (chemical) hydraulic will be caused either by calcium or hydroxyl ions. Since calcium is abundantly present in the environment and since the effect concentrations are within the same order of magnitude of its natural concentration, it can be assumed that the adverse effects are mainly caused by the pH increase caused by the hydroxyl ions. Furthermore, the above mentioned calculations show that the base equivalents are within a factor 2 for hydraulic lime (chemical) and calcium hydroxide. As such, it can be reasonably expected that the effect on pH of lime (chemical) hydraulic is comparable to calcium hydroxide for a same application on a weight basis. Consequently, read-across from calcium hydroxide to lime (chemical) hydraulic is justified.

4. DATA MATRIX
Source: No studies available
Target: Klimisch 1 study: NOEC (14d) >= 5000 mg/kg soil dw. for calcium dihydroxide on biomass of the earthworm Eisenia fetida
Klimish 1 study: LC50 (14d) > 5000 mg/kg soil dw. for calcium dihydroxide on mortality of the earthworm Eisenia fetida
Klimisch 1 study: NOEC (4 wk) = 2000 mg/kg soil dw. for calcium dihydroxide on reproduction of the earthworm Eisenia fetida in an artificial soil
Klimish 1 study: EC50 (4 wk) = 4180 mg/kg soil dw. for calcium dihydroxide on reproduction of the earthworm Eisenia fetida in an artifical soil

Reason / purpose for cross-reference:

read-across source

Duration:

4 wk

Dose descriptor:

NOEC

Effect conc.:

2 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

Ca(OH)2

Basis for effect:

reproduction

Duration:

4 wk

Dose descriptor:

EC50

Effect conc.:

4 180 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

Ca(OH)2

Basis for effect:

reproduction

Remarks on result:

other: 95% CL: 3346-4665 mg/kg soil d.w.

Duration:

4 wk

Dose descriptor:

NOEC

Effect conc.:

5 000 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

Ca(OH)2

Basis for effect:

mortality

Description of key information

Klimisch 1 study: NOEC (14d) >= 5000 mg/kg soil dw. for calcium dihydroxide on biomass of the earthworm Eisenia fetida (Friedrich, 2007a)

Klimish 1 study: LC50 (14d) > 5000 mg/kg soil dw. for calcium dihydroxide on mortality of the earthworm Eisenia fetida (Friedrich, 2007a)

Klimisch 1 study: NOEC (4 wk) = 2000 mg/kg soil dw. for calcium dihydroxide on reproduction of the earthworm Eisenia fetida in an artificial soil (Friedrich, 2007b)

Klimish 1 study: EC50 (4 wk) = 4180 mg/kg soil dw. for calcium dihydroxide on reproduction of the earthworm Eisenia fetida in an artifical soil (Friedrich, 2007b)

Klimisch 1 study: LC50 (14d) > 1000 mg/kg soil dw; NOEC (14d) = 1000 mg/kg soil dw for calcium carbonate using the earthworm Eisenia Foetida (Goodband, 2010)

Key value for chemical safety assessment

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

2 000 mg/kg soil dw

Additional information

The short-term toxicity of calcium dihydroxide on mortality and biomass of the earthworm Eisenia foetida (Friedrich, 2007a) was carried out according to OECD test guideline 207. The study is well-documented, all validity criteria are fulfilled. As such a Klimisch 1 score was assigned to the study. After 14 days, no significant effect on both mortality and biomass was observed up to the highest tested dose (5000 mg Ca(OH)2 /kg dw).

The chronic study on the effect of calcium dihydroxide on the reproduction of the earthworm Eisenia foetida (Friedrich, 2007b), was carried out according to OECD test guideline 222. The study is well-documented, all validity criteria are fulfilled. As such a Klimisch 1 score was assigned to the study. The study resulted in a 4w-EC50 of 4180 mg Ca(OH)2 /kg soil dw and a 4w-NOEC of 2000 mg Ca(OH)2 /kg soil dw.

The acute toxicity to earthworms of calcium carbonate (nano) was assessed in a study performed according to OECD TG 207 under GLP (Goodband, 2010). Eisenia foetida were exposed to calcium carbonate in a limit test at 1000 mg/kg in soil for 14 days. There were no significant mortalities or sub-lethal effects observed. Hence, the 14 day LC50 value was >1000 mg/kg soil dw and the NOEC was 1000 mg/kg soil dw. Calcium carbonate is therefore not toxic to earthworms up to a concentration of 1000 mg/kg soil dw.

Long term toxicity tests are not available for calcium carbonate, but given the ubiquitous nature of calcium and carbonate ions in the environment it is considered that calcium carbonate will not have a detrimental effect on soil macroorganisms over an extended time period and it would be expected that any potential toxicity will be limited by its solubility when compared to calcium dihydroxide.

In conclusion, both calcium dihydroxide and calcium carbonate show no effects when tested up to limit doses of 5000 mg/kg dw and 1000 mg/kg dw, respectively in acute tests. In a chronic study, calcium dihydroxide did show significant adverse effects on reproduction at 4000 and 5000 mg/kg soil dw and the NOEC was determined to be 2000 mg/kg soil dw. No explanation for the toxicity observed is given in the study report, however, given the large difference in solubility between calcium dihydroxide and calcium carbonate, it would be expected that such effects will be mitigated when exposure to high concentrations of calcium carbonate occurs. On this basis, it may be concluded that the results for calcium dihydroxide represent a worse-case for grades of lime (chemical) hydraulic containing up to 40% calcium carbonate.