**2. Material and methods**

**Site description.** Field data were sampled by the investigators on the site of forest remediation in Western Donbass (Ukraine, Dnepropetrovsk region). Soil samples were collected at a depth of 0–10 cm, and fresh excreta (casts) of earthworm *Aporrectodea caliginosa* (Savigny, 1826) were sampled at the surface on the remediation site in a plantation of Norway maple (*Acer plata‐ noides*) (second and third variants of remediation). The first variant of remediation was represented by filling of mining spoil unsuitable for growth of arboreal plants. Top layer of the second variant sampled for assay was represented by humus-free loess-like loam; and top layer of the third variant was represented by a humic filling layer of ordinary chernozem (**Figure 1**).

Earthworm *A. caliginosa* is referred to endogeic soil worms. It is classified as a saprophage, secondary decomposer, nitrogen liberator, and humificator [20, 24].

General description of the forest vegetation and filling remediated soil on the site of mine dump forest recultivation located within the territory of the Western Donbass (Ukraine, Dnepropetrovsk oblast) is shown as follows:

**First variant**. Platform of dump mine spoil was coated with a layer of the same spoil 2 m in thickness. Such variant of remediation was created with the aim to identify environmental suitability or unsuitability of mining spoil for the forest plantation growing. By 2005, trees and bushes had been died off completely within this variant of remediation. Mine spoil of the Western Donbass is a mass heavy loamy in granulometric composition, consisting of aleurolites and argillites; it contains 16–20% of organic carbon. The mine spoil is unsuitable for plant growth in its physicochemical, water, air, and mechanical properties and composition. It is absolutely impermeable, have a higher density, hardness, viscosity, stickiness, and adhesiveness. Such spoil dries to cement condition, and when wet it turns into viscous clay with a high water capacity and lack of air. Agrochemically, mining spoil is represented by nitrogen-free compounds, with trace amount of phosphorus, potassium, calcium, magnesium, sulfur, iron, and minor-nutrient elements. Against this background, pyrite provides especially negative effect (1.5%), contributing to decrease of actual acidity to 3.0 units. Fresh, thrown to the daylight surface, mine spoil has an evaporated residue of not more than 0.4%.

Plants accumulate most of copper into their leaves and seeds, less in roots, and very little in stems [13]. Copper is a component of numerous enzymes insuring normal cells functioning; it takes part in process of chlorophyll formation and other oxidation-reduction processes into plant cells. Copper deficiency in plants causes lowering activity of synthetic processes and leads to accumulation of soluble carbohydrates, amino acids, and other degradation products of complex organic substances; such process leads to withering, turgor loss, chlorosis, delayed shooting stage, and poor seed formation [2]. In animals, copper involved in processes of enzyme activation and it is part of the respiratory proteins such as hemoglobin and hemocyanin [22]. Living organisms-concentrators of copper are well known among both plants and animals (tea plant, mollusks, spiders, etc.). Many animals and plants experience toxicity from copper excess [19]. In most cases, trace elements (particularly copper) come to the animals through trophic chains. Considering representatives of saprophages, it should be noted that

The goal of the article was evaluation earthworm (*Lumbricidae*) tropho-metabolic effect in maintaining capacity of remediated artificial soil to resist from copper contamination. This paper determines quantitatively buffer capacity of artificial soil and earthworm casts from copper contamination, and make a comparison of immobilization capacity between earthworm casts and remediated soil. Soil buffering capacity is maintaining the chemical soil state unchanged under the influence of chemical compounds flow. Assessments of rates of *Lumbri‐ cidae* impact on the environment, particularly the effect of tropho-metabolic activity of earthworms on buffer capacity of the remediated soil are of scientific and practical interest in

**Site description.** Field data were sampled by the investigators on the site of forest remediation in Western Donbass (Ukraine, Dnepropetrovsk region). Soil samples were collected at a depth of 0–10 cm, and fresh excreta (casts) of earthworm *Aporrectodea caliginosa* (Savigny, 1826) were sampled at the surface on the remediation site in a plantation of Norway maple (*Acer plata‐ noides*) (second and third variants of remediation). The first variant of remediation was represented by filling of mining spoil unsuitable for growth of arboreal plants. Top layer of the second variant sampled for assay was represented by humus-free loess-like loam; and top layer of the third variant was represented by a humic filling layer of ordinary chernozem

Earthworm *A. caliginosa* is referred to endogeic soil worms. It is classified as a saprophage,

General description of the forest vegetation and filling remediated soil on the site of mine dump forest recultivation located within the territory of the Western Donbass (Ukraine, Dneprope-

**First variant**. Platform of dump mine spoil was coated with a layer of the same spoil 2 m in thickness. Such variant of remediation was created with the aim to identify environmental

secondary decomposer, nitrogen liberator, and humificator [20, 24].

copper as a trace element is always presented in their body and excreta [17].

relation to soil fertility management.

280 Soil Contamination - Current Consequences and Further Solutions

**2. Material and methods**

trovsk oblast) is shown as follows:

(**Figure 1**).

**Second variant**. Type of forest growing conditions: DL0–1 (dryish loam). Stratigraphic structure of soil profile: loess-like loam: 0–50 cm; tertiary sand: 50–100 cm; mine spoil: 100–700 cm. *Planting with Norway maple*. Type of light structure: half-shade. Type of timber-stand: 10 N. m., height: 8–10 m, average trunk diameter: 100–120 mm, crown closure: 0.6–0.7. Litter from maple leaves is poorly developed; it is mainly accumulated between the tree lines in the relief depressions. The grass cover is missing.

**Third variant**. Type of forest growing conditions: DL0–1 (dryish loam). The remediated bulk soil has the following stratigraphical characteristics: humic topsoil of ordinary chernozem: 0– 50 cm; loess-like loam: 50–100 cm; tertiary sand: 100–150 cm; mine spoil: 150–700 cm. *Planting with Norway maple*. Type of light structure: half-lightened. Type of timber-stand: 10 N. m., height: 8–10 m, average trunk diameter: 100–120 mm, crown closure: 0.5–0.6. The litter layer is well developed; leaves are almost completely decomposed. The topsoil is moist to the touch, well structured. With a depth of 30 cm, it is compacted, occupied densely by maple roots to a depth of 50 cm. The grass cover is missing.

**Sampling and experimental procedures.** Definition of zoogenic participation in the process of stability formation in soil as a saprophages habitat (earthworms, *Lumbricidae*) against contamination by copper was performed by adding different amounts of copper with it absorption from copper solutions. As the methodological basis, recommendations developed by researchers of the National Scientific Center 'Institute for Soil Science and Agrochemistry Research named after O.N. Sokolovsky' were applied [9, 26]. Air-dry sample specimens of soil and earthworm excreta (casts) were placed in cylindrical vessels, filled with a solution of copper sulfate pentahydrate CuSO4 5H2O contained copper in scalar concentrations (from 5 to 40 mg Cu/L), in a ratio of weigh/solution of sulfate of 1/10; suspension has been stirred for 2 h and left to stand for 1 day and filtered. The remaining soil onto the funnel filter was transferred to a glass box and dried to air-dry state. Samples were selected from samples prepared in such manner to determination of mobile forms of copper compounds. As extractant, it used ammonium-acetate buffer (pH = 4.8). Content of extractable copper was determined by the method of atomic spectrophotometry. Quantitative determination of area under curve that characterizes the sustainability of earthworm casts and soil to the flow of toxicant was performed by means of numerical integration using Simpson's formula [18].

**Figure 1.** Variants of artificial soil in experimental-production area of forest reclamation and their stratigraphic structure.

To determine the zoogenic environmental-forming function in formation of soil resistance against copper contamination, we studied immobilization (immobility)-mobilization (mobility) of copper amount in earthworm casts and bulk soils, and participation of earthworm casts in formation of resistance against contamination with copper. To assess the impact of earthworms' tropho-metabolic activity for maintaining resistance of their habitats to copper pollution, we used effect and toxicant immobilization efficiency.

#### **3. Results and their discussion**

Effect of earthworm excreta (casts) on the soil resistance from flow of toxic agents such as high concentrations of copper was investigated on earthworm casts sampled in Norway maple planting. Graphic model of earthworm casts resistance to copper contamination (second and third variants of remediation) are represented in **Figures 2** and **3**. It indicates higher buffering capacity of casts in humus variant.

**Figure 2.** Graphic model of earthworm casts resistance to copper contamination (second variant, humus-free loess-like loam): 1 – Earthworm casts (humus-free loess-like loam, second variant); 2 – Reference.

Environmental Role of Earthworm (*Lumbricidae*) in Formation of Soil Buffering Capacity Against Copper... http://dx.doi.org/10.5772/64722 283

**Figure 1.** Variants of artificial soil in experimental-production area of forest reclamation and their stratigraphic struc-

To determine the zoogenic environmental-forming function in formation of soil resistance against copper contamination, we studied immobilization (immobility)-mobilization (mobility) of copper amount in earthworm casts and bulk soils, and participation of earthworm casts in formation of resistance against contamination with copper. To assess the impact of earthworms' tropho-metabolic activity for maintaining resistance of their habitats to copper

Effect of earthworm excreta (casts) on the soil resistance from flow of toxic agents such as high concentrations of copper was investigated on earthworm casts sampled in Norway maple planting. Graphic model of earthworm casts resistance to copper contamination (second and third variants of remediation) are represented in **Figures 2** and **3**. It indicates higher buffering

**Figure 2.** Graphic model of earthworm casts resistance to copper contamination (second variant, humus-free loess-like

loam): 1 – Earthworm casts (humus-free loess-like loam, second variant); 2 – Reference.

pollution, we used effect and toxicant immobilization efficiency.

**3. Results and their discussion**

282 Soil Contamination - Current Consequences and Further Solutions

capacity of casts in humus variant.

ture.

**Figure 3.** Graphic model of earthworm casts resistance to copper contamination (third variant, humic layer): 1 – Earthworm casts (humic layer, third variant); 2 – Reference.


**Table 1.** Quantitative assessment of earthworm casts and soil resistance against copper contamination.

Results of the study show that in the range of Cu concentration from 50 to 400 mg, effect of casts (Sref – Ssamp) on copper immobilization in the humus-free loess-like loam (second variant) was less than the effect of casts in the humic layer of ordinary chernozem (third variant), with a high level of statistical significance (*p* = 0.0011), and is 197.5 and 373.4 area units, respectively (**Table 1**). The effectiveness of immobilization that reflects resistance degree to contamination by this metal was increased from 23.1% (casts onto the humus-free loess-like loam) to 43.6% (casts on bulk humic layer from ordinary chernozem in the plantings of Norway maple). This, apparently, is due to the fact that the casts formed on loess-like loam is represented by the soilforming rock that contain no organic matter (particularly humus), while the earthworm casts that formed on filling humic layer includes soil organic matter. Furthermore, the presence of humic compositions in earthworm casts is a powerful factor in process of stability formation in remediated soil against the effects of toxic concentrations of copper.

In the context of soil-casts system, effect of casts (Sref – Ssamp) on copper immobilization within concentration range of Cu from 50 to 400 mg is more than the effect of initial bulk soil (respectively 170.6 and 197.5 area units on the second variant with humus-free loess-like loam; 343.7 and 373.4 area units on the third variant with humic layer of ordinary chernozem). In both cases, difference between average effects was statistically significant; values of significance level were 0.03 and 0.045, respectively (**Table 2**).

Efficiency of immobilization in the studied casts (humus-free and humic variants) was more (23 and 43%, respectively) than the efficiency of immobilization in the initial soil: loess-like loam and chernozem (19.9 and 40.1% respectively, see **Table 1**). Efficiency of copper immobilization by casts compared with the corresponding initial soil (variants with loess-like loam and ordinary chernozem coating) was more by 3.1% (the difference between 23 and 19.9%) and 3.5%, respectively (the difference between 43.6 and 40.1%). It evidences for the positive environment-forming role of earthworms (particularly their tropho-metabolic activity) in development of protective and buffer shield of remediated soils and enhances the immobilization ability of the zoogenic soil neoformations—casts—within sites of forest remediation. Thus, earthworm tropho-metabolic activity within different variants of forest remediation sites affects the soil immobilization capacity maintenance (buffering capacity to heavy metals, including copper).


**Table 2.** Statistical evaluation of differences between effects of earthworm cast and bulk soil against copper contamination.
