WO2007111479A1

WO2007111479A1 - Soil improver with white-rot fungi and methods of using thereof for plant restoration and recover of heavy metals contaminated soil around abandoned mine area

Info

Publication number: WO2007111479A1
Application number: PCT/KR2007/001519
Authority: WO
Inventors: Kyu Jin Yum; Jae Cheon Lee; Sim Hee Han
Original assignee: Korea Forest Research Institute
Priority date: 2006-03-28
Filing date: 2007-03-28
Publication date: 2007-10-04
Also published as: KR20070097622A; KR100903666B1

Abstract

The present invention relates a soil improver comprising sludge, sawdust (a waste from mountain village) and white rot fungus, wherein the amount of white rot fungus is more than 107 cells per unit gram of the sludge and sawdust. The soil improver manufactured by adding white rot fungus into the base material comprising sludge and sawdust according to the present invention is a matured soil improver so that it is difficult to distinguish from upland soil; a economical soil because it can be quickly manufactured with the help of added white rot fungus; and a high effective soil improver because it contains a effective composition of sludge and degraded material of sawdust. By adding the soil improver according to present invention, the plantation of the soil of abandoned mine having low pH and nutrient shortage can be restored and heavy metals can be removed.

Description

[ Description]

[ Invention Title]

SOIL IMPROVER WITH WHITE-ROT FUNGI AND METHODS OF USING THEREOF FOR PLANT RESTORATION AND RECOVER OF HEAVY METALS CONTAMINATED SOIL AROUND ABANDONED MINE AREA

[Technical Field]

The present invention relates to a soil improver and methods of using thereof. The soil improver, according to the present invention, facilitates plant growth and removal of heavy metals from abandoned mine areas. More specifically, the present invention relates to a soil improver manufactured by adding white rot fungi into the starting material of waste sludge and sawdust. This has outstanding effects on the promotion of plant growth in the forest area of abandoned mines and on removal of heavy metals from the soil of abandoned mines .

[Background Art] In our country, a lot of sludge has been produced due to the increase of sewage and waste water which are increased because water usage has been increased as result of industrial development and improved living standards .

Sludge is one of the sources of contamination. It causes high costs and soil and ocean environmental contamination because, for preventing contamination of outdoor environment, sludge is converted into a solid form by concentration, dehydration, and drying etc. and then buried into soil or abandoned into the ocean with sludge delivery ships.

Even though sludge contains a sufficient amount of organic material and low concentration of various kinds of contamination material, so that it has sufficient value as an organic material resource, only 2.9% of sludge is reused by recycling.

In domestic research, there were some researches on recycling of agricultural or livestock byproducts, which are a research on composting of agricultural or livestock byproducts, a research of preparation of an agricultural fertilizer by mixing them with sewage water, and a research of synergistic effects produced by the combination of damaged soil of mines with sewage sludge. But all of these researches were only conducted with a laboratory scale.

The techniques of manufacturing soil improver from active sludge and sawdust were disclosed in Korean patent applications numbers 1999-0023234, 2000-0028866, 2000- 0008634, and 1997-0075288, and the technique of composting active sludge and sawdust by adding yeast was disclosed in Korean patent applications numbers 1995-0007037.

As mentioned above, the composting technique adding sawdust into sludge facilitates composting speed by increasing aeration property during composting. However the sawdust is comprised of cellulose, hemicellulose and lignin which are non biodegradable and are degraded by only the specific microorganism producing cellulase and ligninase. Therefore there are problems that sometimes the sawdust is not degraded in certain environmental conditions of composting, such as the temperature, and the moisture etc.

To solve the above problems, the inventors of present invention make the present invention based on the finding that the composting is facilitated by the addition of white rot fungi in the early stage of composting.

[Disclosure] [Technical Problem]

Accordingly, to solve the above problems occurring in the prior art, an object of the present invention is to provide a soil improver which is made from the starting materials of sludge and sawdust by a sufficient and rapid composting.

Another object of the present invention is to provide a method of restoration of plant. Still another object of the present invention is to provide a method of removal of heavy metal in abandoned mine area with the soil improver.

[Technical Solution]

The present invention provides a soil improver manufactured by adding white rot fungi which is capable of degrading saw dust, to the starting materials comprising sludge and sawdust. The soil improver has prominent effects on removal of heavy metal and promotion of plant growth in abandoned mine area.

[Description of Drawings]

FIG. 1 shows a variation of composting temperature according to the addition of white rot fungi which facilitate composting.

FIG. 2 shows a difference in chlorophyll content in selected trees according to the treatment of the soil improver .

FIG. 3 shows a difference in growth response in selected trees according to the treatment of the soil improver. (A) 0 cc treatment of soil improver; (B) 500 cc treatment of soil improver

FIG. 4 shows development of selected tree in upper- earth part and rhizosphere in under-earth part according to the treatment of the soil improver. (A) 0 cc treatment of soil improver (B) 500 cc treatment of soil improver (c) 1000 cc treatment of soil improver

FIG. 5 shows difference of microorganism activity in coal soil according to the treatment of the soil improver with an cellulose fabric assay method.

FIG. 6 shows variation of harmful heavy metal concentration in effluent water according to time course after treatment of the soil improver [Best Mode]

For said purpose, the present invention provide a soil improver comprising sludge, degraded sawdust and white rot fungi, wherein the amount of white rot fungi is more than 10⁷ cells per unit gram of sludge and sawdust. If the amount of white rot fungi is less than 10⁷ cells, the composting process of sawdust is slowed or insufficiently- composted. As used herein, the term "white rot fungi" refer to a

fungus capable of degrading cellulose, hemicellulose and

lignine in trees. White rot fungi can be one or more species

selected from the Genus group consisting of Genus Trametes,

Genus Pleurotus, Genus Phanerochaete, Genus Fusarium, and

Genus Phanerochaete. More specifically, white rot fungi is a

species selected from the group consisting of Trametes

versicolor, Pleurotus ostreatus, Phanerochaete sordida,

Trametes hirsutus, Fusarium culmorum, and Phanerochaete

chrysosporium. The soil improver according to the present

invention is manufactured by composting of the mixture of

sludge and added materials at suitable temperature, more

specifically at an initial water content rate being in a

range of between 65 and 67%.

After mixing sludge and sawdust, normal composting in

terms of composting temperature was done at initial water content rate in the range of between 65 and 67% and the

sawdust mixing rate was 30, 40 or 50%. When the sawdust

mixing rate was 30%, 40% or 50%, the maximum composting

temperature was about 52 ^°C, about 54 ^°C and about 69 °C respectively. As the amount of organic material content

before composting was 60, 63 and 67% respectively, the

amount of organic material content has a positive

relationship with the maximum composting temperature.

However, if the initial water content was more than 70% in

sludge, suitable composting was not occurred even in the

case that sawdust mixing ratio was more than 30% because the

composting temperature was not increased to be more than

50 ^°C due to the initial water content rate being more than 65-67%. Thus the initial mixing ratio of composting

materials should be decided in the aspect of initial water

content rate (65-67%) .

The second aspect of the present invention provides

methods of preparation of soil improver by adding white rot

fungi into the materials comprising sludge and sawdust. More

specifically, the white rot fungi can be more or more

species selected form the group consisting of Trametes

versicolor, Pleurotus ostreatus, Phanerochaete sordida,

Trametes hirsutus_r Fusarium culmorum, Phanerochaete chrysosporium and the deposited microorganism of deposition number KCCM 10725P.

The third aspect of present invention provides a

method of plant restoration by introducing the soil improver

to abandoned mine area. More specifically, the present

invention provides a method that the soil improver is

covered on coal soil with more than 25% ratio by weight of

the soil improver to the surface soil of coal soil in the

abandoned mine area.

As used herein, the term "surface soil" refers to dark

grey topsoil layer which has mineral materials and organic

decays. Soil is made up of horizontal layers formed parental

material because soil surface influence greatly by climate

and actively occurred differentiation of layer. From the

bottom layer, soil is made up with (J) unweathered rock

(bedrock) layer (R horizon) , (2) regolith layer affected

little by soil formation action (C horizon) CD subsoil

deposited and precipitated with various materials in soil (B

horizon) ® topsoil made up of humus (decomposed organic

matter) mixed with mineral particle (A horizon) and (B

organic layer in a top (0 horizon) . The topsoil layer is

suitable for plant growth due to its good nutrient

absorption and maintenance capacity. The depth of surface soil is average 30 cm, becoming to 1 m in thick place.

The forth aspect of the present invention provides

methods comprising steps of (a) introducing the soil

improver according to the present invention into the soil of

abandoned mine area; and (b) planting a plant able to remove

heavy metals. More specifically the soil improver covers

coal surface soil of abandoned mine area in the ratio of the

soil improver to coal surface soil being more than 25%. The

plant able to remove heavy metal is one or more selected

from the group consisting of Betula schmidtii, Betula

costata, Pinus Densiflora, Fraxinus rhynchophylla, Fraxinus

sieboldiana, Rhododendron mucromulatum, Weigela subsessilis,

Acer pseudosieboldianum, Rhododendron schlippenbachii, Salix

hulteni, Betula platyphylla, Salix koreensis, Alnus

japonica, Stephanandra incisa, Mignolia sieboldii,

Callicarpa japonica, Acer mono, Quercus mongolica, Corylus

sieboldiana and Tilia amurensis.

Hereinafter, the present invention will be

described in further detail with reference to examples.

It is to be understood, however, that these examples

are for illustrative purposes only and are not to be construed to limit the scope of the present invention.

[Mode for Invention] [Example 1] PREPARATION OF SOIL IMPROVER

In case that a large amount (more than 70%) of sawdust is needed to be inputted into the original sludge having high water content (more than 90%) , the sawdust mixed by 5% with dominant bacterial species and a culture solution of white rot fungi and then introduced into the sludge in the initial stage of decomposition. The dominant bacterial species is the mesophilic and thermophilic bacteria founded in a decomposition period. Fig. 1 and Table 1 show the change of decomposition temperature, the characteristics of decomposition progress, and physical and chemical properties (organic matter, water content, pH and C/N) .

< Table 1 > physical and chemical properties with or without white-rot fungi in the presence of mesophilic or thermophilic microorganisms

As a result of physical and chemical properties analysis of the mixture with or without white-rot fungi in the presence of mesophilic or thermophilic microorganisms, water content without white-rot fungi decreased from 62.36% to 45.53% and content of organic matter with white-rot fungi decreased from 51.86% to 25.92%. Before decomposition, C content (%) with or without white-rot fungi was analyzed 36.64% and 33.88%, respectively. After decomposition, C content (%) with or without white-rot fungi decreased 29.98% and 31.26%, respectively. Consequently even if sawdust was added in great amount more than 70%, all of two groups with or without white-rot fungi satisfied the standard of composted soil. When white- rot fungi was added in the presence of thermophilic microorganisms, the water content in a final mature soil decreased due to high exothermic reaction (more than 50^°(J in a second decomposition step, and high decomposition temperature (about 40^°(J was maintained during post decomposition step after 30 days. As shown in Fig. 1, the total decomposition period was about 60 days and was lengthened about two times than that of the decomposition progress having relatively lower sawdust input (30% sawdust) . But high-quality soil improver could be produced by addition of white-rot fungi degrading sawdust, even if the sludge is mixed with more than 70% of sawdust having non-degrading lignin as a main component .

[Example 2] METHOD OF MANUFACTURING SOIL IMPROVER

An optimal decomposition process to highly produce soil improver for plant restoration around abandoned mine area was developed by using sewage sludge and sawdust added decomposition stimulating microorganism.

Sawdust was mixed by 5% with mesophilic or thermophilic microorganisms and the culture solution of white rot fungi degrading cellulose in sawdust. The appropriate mixing ratio of sludge to the sawdust was determined to be 65-67% of the water content of mixture. Aeration was done by forced air injection at the bottom of decomposition reactor. Aeration was carried out at a rate of 0.2i air/min per Ii compost soil during first decomposition step and then carried out at a rate of O.li air/min per It mature soil in the second decomposition step to maintain appropriate temperature of decomposition because as decomposition was carried out, the volume of compost soil decreased. After finishing stable decomposition by added white-rot fungi and mesophilic or thermophilic microorganism which degrade organic matter, generate heat more than 65 ^°C in decomposition reactor and thereby remove moisture, the compost mixture was transferred to post decomposition reactor and then packed to be easily handled as a soil improver.

[Example 3] METHODS OF PLANT RESTORATION USING SOIL IMPROVER

(1) Selection of indigenous tree species for the plant restoration of the abandoned coal mine lands in Taeback area, Korea

(a) The content of hazardous components in the body of nature imported tree species on the abandoned coal mine lands

We investigated the content of hazardous components in the body of picked trees on the abandoned coal mine lands . Mn was found 111.3 mg/kg with the highest concentration and As was found 0.21mg/kg with the lowest concentration. But Cd of 10 atoms wasn't detected in the tree body.

(b) Physiological activity of indigenous tree species on the abandoned mine area

(i) Differences in the content of malondialdehyde (MDA) and H₂O₂ between leaves Malondialdehyde (MDA) was a material produced by peroxydation of lipid in a living body and was used as an indicator of damaged levels from various stress (Kim and Lee, 1994; Davis and Swanson, 2001) . The content of malondialdehyde (MDA) measured in leaves of Betula costata and Betula schmidtii Regel was different not only between tree species but also between regions. Specially, the malondialdehyde (MDA) content of Betula costata and Betula schmidtii Regel growing on the abandoned coal mine lands was higher than that growing in forests around the abandoned coal mine lands . These results suggest that the environment on the abandoned coal mine lands have various stress factors and trees on the abandoned coal mine lands are damaged by various stress factors. Also, the content of H₂O₂ measured in leaves differs between tree species and regions. But differences between regions were not larger than the differences between tree species. That is, the H₂O₂ content in leaves of Betula costata showed differances between regions, while the H₂O₂ content in leaves of Betula schmidtii Regel showed no difference between regions. H₂O₂ is a strong oxidant, which is produced through degradation of O₂ by superoxide dismutase

(SOD) in thylakoid membrane. H₂O₂ concentration is regulated by catalase and peroxydase and is maintained at a proper level in the biological world. In case of plants or photosynthetic algae, additional ascorbate peroxidase (APX) exists, which plays an important role in the removal of H₂O₂ within chloroplasts, a photosynthetic machinery (Foyer, 1993) . As H₂O₂ has specific membrane penetration ability and relatively its long span, it can influence to over all cells as well as photosynthetic machinery including photosystem I (PS I) and photosystem II (PS II) . Particularly, photosystem II which is not protected by ascorbate peroxidase could be damaged seriously by H₂O₂ accumulation (Prasad et al., 1994). As the H₂O₂ increase in leaves was resulted from the accumulation of H₂O₂ in the body, the Betula costata in the investigated regions was damaged more than Betula schmidtii Regel by H₂O₂. It is estimated that Betula costata is more sensitive to stress than Betula schmidtii Regel.

(ii) Differences in the activity between nitrate reductase and antioxygenase in leaves

The activity of nitrate reductase in leaves is used as a criterion to measure indirectly the availability of nitrogen concentration in plant growing soil and to determine plant responses by environmental stress (Srivastava, 1980; Hogberg et al., 1986). The nitrate reductase activity of Betula costata and Betula schmidtii Regel on the abandoned coal mine lands showed large differences between regions as well as between tree species. The nitrate reductase activity of Betula schmidtii Regel was 129.5μmol/g higher than Betula costata. The nitrate reductase activity of Betula costata and Betula schmidtii Regel growing in forests around the abandoned coal mine lands showed higher than the activity of them growing on the abandoned coal mine lands . The reason why there was large difference in the nitrate reductase activity between tree species is that soil conditions growing each tree species and nitrogen use efficiency differ between tree species (Adams and Attkwill, 1982; Crick and Grime, 1987) . Also, the low nitrate reductase activity of trees on the abandoned coal mine lands is related to low nitrogen content in the abandoned coal mine soil, which is because unsuitable environmental conditions in the abandoned coal mine soil influence the enzyme activity by acting as a stress for trees growing on the abandoned coal mine lands (Srivastava, 1980) .

Meanwhile antioxygenase, a superoxide dismutase (SOD) , activity was difference between trees, however, there was no difference between regions (Table 2) .

< Table 2 > statistical analysis of the nitrate reductase (NR) activity, superoxide dismutase (SOD) activity, H₂O₂ content and malondialdehyde content in leaves of Betula costata and Betula schmidtii Regel growing naturally in areas and forests around on the abandoned coal mine lands in Taeback area, Korea

*, ** and *** means differences in significance levels of 0.01, 0.001 and 0.0001 respectively.

Results showed that trees on the abandoned coal mine lands were influenced by various stress. Nevertheless, the superoxide dismutase (SOD) activity was not increased. This is because available energy resources required for defense actions were not supplied sufficiently. As shown in decrease of nitrate reductase activity in leaves of trees on the abandoned coal mine lands, it is thought that low content of nitrogen in the abandoned coal mine soil may influence on the production of enzymes related to remove stress.

(iii) Differences in the carbohydrate concentration in leaves

There was no difference in the carbohydrate concentration measured in leaves of Betula costata and Betula schmidtii Regel between trees on the abandoned coal mine lands and forests around there, but there was a difference between regions. But unlike Betula schmidtii Regel, there was no difference in carbohydrate concentration in leaves of Betula costata between regions. In the case of Betula schmidtii Regel, glucose concentration in leaves of populations growing in forests around the abandoned coal mine lands was higher than populations growing on the abandoned coal mine lands. On the contrary, starch concentration in leaves of populations growing on the abandoned coal mine lands was higher than populations growing in forests around there.

It is thought that the decrease in glucose concentration in leaves of trees on the abandoned coal mine lands relates to tolerance responses to stress. In other words, it is thought that the decrease in glucose concentration is because glucose is used as a source of

NADPH for maintaining optimal anti-oxidant materials

(Robinson and Rowland, 1996) , and because consumed glucose is not supplied since stress factors inhibit sucrose hydrolysis. Under the normal condition, starch is accumulated in leaves during daytime and is transported to sink after being hydrolyzed and converted to sucrose at night. However, sometimes the increase in concentration of starch and total carbohydrate in the pollution area is observed. This is thought that hydrolysis of starch and transportation of sucrose are inhibited by pollutants (Samarakoon and Rauser, 1979) . Specially, it is known that plants supplied with very low nitrogen accumulate much more starch in leaves than plants supplied with sufficient nitrogen (Ericsson, 1979; Balsberg-Pahlsson, 1989) .

When we put together various phenomenon appeared in trees on the abandoned coal mine lands in the present investigation, trees on the abandoned coal mine lands were greatly damaged by inhibition of various metabolisms and tolerance expressions from stresses. It is thought that the damage of trees would increase because low content of nitrogen in abandoned coal mine lands does not help to have tolerance expressions against stress. Accordingly, in order to prevent damages and increase stress tolerance of trees in abandoned coal mine lands, it is urgently demanded the supply of nutrients to be used as various energy sources of metabolism. (c) Selection of tolerance tree in a damaged area by mine and raising seedlings

The selection for tolerance tree in a damaged area by mine was based on results of the vegetation investigation and plant body analysis. It was founded that Betula costata and Betula schmidtii Regel were best tree species among naturally imported tree species on the abandoned coal mine lands. A tree species was selected among naturally imported tree species on the abandoned coal mine lands, marked at the spot and then collected seeds. The germination test by using collected seeds showed a very low germination rate. That is, the germination rate of Betula costata was less than 12% and that of Betula schmidtii Regel was less than 8%. After finishing the germination tests, the trees were raised by- tree species and individually to obtain tolerance families.

(2) Growth promotion effects by the addition of soil improver

Because it was thought to be difficult to uniformly mix with coal soil when compost soil was supplied in the fields, ports having coal soil in down layer (50% volume ratio) , and compost soil and peat in upper layer

(50% volume ratio) of 75% : 25% (v: v) , 50%:50%, 25%:75% and 0%:100% were prepared and then planted mustard plumules having strong tolerance and good development of roots. After growing for 45 days, the growth of mustard was analyzed and the heavy metals concentration absorbed into leaves and roots was measured. After finishing tests, we investigated the microbial activity in the down coal soil, investigated residual heavy metals contents in the soils of upper layer and down layer, examined the effects of plant nutrients, and examined optimal mixture ratios, examined whether damage had happened by hazardous toxic heavy metals, and found suitable mixing ratio according to organic fertilizing by upper part.

After growing for 45 days, the roots of mustard grew into the coal soil in down layer without significant damage symptoms. After final harvesting, the mustard growth was investigated and the results were showed in Table 3.

Generally, the more the content of organic matters in compost soil was increased, the more the growth of mustard was stimulated. The optimal mixture ratio was the sample added 50% of compost soil. Table 4 showed the contents of chlorophyll and water-soluble proteins.

To analyze the data and statistics, ANOVA analysis was done on the average value of each data and the difference betweens average values were analyzed by using Duncan's multiple range test. A statistical analysis carried out using statistical analysis program SPPS (version 10.2) .

< Table 3 > the growth of mustard growing in soil including mature soil and in mixture soil including coal soil

< Table 4 > the contents of chlorophyll and water- soluble proteins of mustard growing in soil including mature soil and in mixture soil including coal soil

Finally, the optimal mixing ratio is from 25% to 50% in the aspect of plant nutrient. For investigating the cause of plant growth promotion in soil microbiological aspect, the enzyme activity of soil microorganism was measured. This showed that the microorganism' s activity was greatly increased in both upper mixed soil and down coal soil (Table 5).

< Table 5 > the microbial enzyme activity in soil with compost sludge, and coal soil cultivated mustard

Overall, the microbial activity of the mixed soil of compost soil and peat in upper layer increased following to the amount of organic matter. This showed that microbial activity was dependent on the added amount of organic matter. The more meaningful and interesting things is that microbial activity is significantly increased even if only upper layer was provided organic matter. When compared with the microbial activity of coal soil before providing organic matter, microbial activity of down layer was increased in a time dependent manner when the mixed soil of compost soil and peat was laid in the upper part . The enzyme activity of xylanase degrading high molecular polysaccharide was increased about 4 times, the enzyme activity of invertase involving degradation of carbon source of polysaccharide and glycoside from xylanase' s degradation activity was increased about 3-4 times, and the enzyme activity of β-glycosidase which was not detected before furnishing compost soil was increased to 2/3 levels of common surface soil. The protease activity and urease involving degradation of organic nitrogen were increased about 20 times, and about 100 150 times respectively. The enzyme activity of phosphatase involving degradation of organic phosphate showed little difference. The enzyme activity of arylsulfatase which was not detected before furnishing organic matter was increased about 1/2 levels. Finally, the enzyme activity of dehydrogenase for using carbon source as energy source was slightly increased. As a result, after furnishing compost soil to upper part, significant amount of eluted organic matter moved to coal soil in a down part and microorganism involved in the material circulation of coal soil. Particularly, the enzyme activities relating to organic nitrogen degrading was obviously increased. The degradation of organic carbon was definitely increased. Particularly, while the influx of organic phosphate was happen little, the degradation enzyme activity was increased according to the increase of influx of organic sulfur. Therefore by addition of compost soil, coal soil was converted from low microbial activity to high microbial activity to be a biologically active soil. This results shows that the present invention provide a platform to increase soil productivity by plant restoration through material circulation of coal soil and stabilization of ecosystem.

(Example 4) The method of removing heavy metals with soil improver

For investigating heavy metal absorption of coal soil by using plant, the concentration of heavy metals in the mixed soil in upper layer, the coal soil in down layer, and harvested leaves and roots of mustard were analyzed (Table 6 and 7) . This results show that the concentrations of heavy metals in compost mixed soil and coal soil are definitely decreased by plant's absorption, and that the removal of hazardous heavy metals by plants is excellent. Therefore the present invention could be used for soil recovery.

[Table 6] The amount of hazardous heavy metals in compost mixed soil and coal soil in which mustard grows

[Table 7] The amount of hazardous heavy metals in the part of mustard cultivated in compost mixed soil and coal soil

The results of nutrient salts and heavy metals in eluted water during 180 days of growth periods after planting tolerant trees showed in tables 8 to 10. In a briefly, the nutrient salts in eluted water from coal soil having plants had low concentration of nutrient salts (K, Ca, Mg, Na) than that of coal soil not having plants, which was prominent in initial growth stage. Also, in case that coal soil had plants, the more compost soil added, the more the absorption ratio of nutrient salts increased. It is thought that the nutrient salts supplied in the form of compost soil facilitates the growth of trees so that the absorption of nutrient salts by trees increases, thus, there are low concentration of nutrient salts in eluted water.

This shows definite effects of nutrient supply by compost soil. On the contrary, high toxic heavy metals such as Cr, As and Mn were not found regardless of the presence of plant, the amount of compost soil, treatment methods etc. In case of Fe and Cu, it had a tendency of reduction of concentration in the eluted water by planting plant.

<Table 8> Nutrient salts and hazardous heavy metals and effects of restoration of plant restoration in eluted water from compost soil added coal soil

<Table 9> Nutrient salts and hazardous heavy metals and effects of restoration of plant restoration in eluted water from compost soil added coal soil (110 days after plantation)

<Table 10> Nutrient salts and hazardous heavy metals and effects of restoration of plant restoration in eluted water from compost soil added coal soil (180 days after plantation)

In the result of nutrient salt and hazardous heavy metals accumulated in tolerant trees, Betula costata and

Betula schmidtii Regel after harvest, there was a close relationships between the element concentration accumulated in unit dry weight and added compost soil. Total amount of elements accumulated in individual trees increased as the amount of compost soil increased. It is thought that the total amounts of elements accumulated increases because the more the amounts of compost soil increases, the more the total dry weight of tolerant trees increases. Also, it is thought that since most of the investigated heavy metals except Fe and Pb were contained much more in compost soils, the more compost soil increases, the more the concentration of heavy metals would increases. However, in the aspect of Fe which was more than 40 times higher in coal soil than that in compost soil, the accumulated Fe amounts was from about 3 times to 9 times regardless of tree species or treatment methods of compost soil, suggesting that hazardous heavy metals in coal soil was efficiently removed by planting tolerant tree and by their growth.

The total accumulation amount of hazardous heavy metals was no big difference between tree species, however, it had a tendency that separate treatments methods was higher amount than that of the mixed treatments methods among compost soil treatment methods. The 25% of compost soil addition rate determined in growth aspect had good efficiency in removing heavy metals. Therefore the compost soil addition to coal soil promotes the activity of soil microorganism converting organic matter to inorganic material, increases nutrients supply and simultaneously remove heavy metals, suggesting that tolerant tree grown in a mixture of compost soil and coal soil could remove heavy metals.

(Example 5) Field evaluation of application in destructed coal soil

(1) Research methods Field evaluation was carried out in a mine-destructed coal soil in Taebaek, Gangwon-Do. One year old seedling of Betula schmidtii Regel selected by National Forest Research Institute was planted in a abandoned mine restoration region operated by National Forest Research Institute in the middle of August, 2003. After digging a hole having 20 cm depth, 30 cm width in the mine-destructed coal soil for planting, three experiment groups, such as a control group not having the developed compost soil, one group having 500 cc of compost soil treatment (xl) , the other group having 1000 cc of compost soil treatment (x2) were randomly distributed in a manner of 5 replicates per each treatment. In each replicates being planted tree in a soil, 8 pieces of cellulose fabrics was set in 5 cm depth and periodically taken away to evaluate microbial activity according to time course after addition of soil improver. After planting, tree growths, amount of chlorophyll and variation of growth was periodically monitored.

(2) Growth response of selected tree according to treatment of soil improver

For investigating the growth response of tree according to treatments of soil improver, the quantity of chlorophyll of selected tree species measured on October 1 and November 1. Measuring results showed that the quantity of chlorophyll was increased 2 times or more after treatment of soil improver, suggesting that nutrient supplement effects by soil improver was definite (Fig 2) . In a research of new root development measured on April 8, 2004, new root development of trees was very quickly progressed by the treatment of soil improver (Fig. 3) . Also in a development of rhizosphere, soil improver treatment group was superior than control group (Fig 4) . After 280 days from planting, the growth parameters of invested tree were shown in Table 11.

<Table 11> Tree growth analysis according to a treatment of soil improver in a mine-destructed coal soil

The soil improver treatment group (xl, x2) was 2 and more times in total tree biomass and total dried biomass. The more soil improver treatment increase, the more the biomass of tree increase. This result shows that by treatment of soil improver in a mine-destructed coal soil, the growth of selected tree species is facilitated and the tree's biological activity was maintained rightly, thus, inducing successful settlement.

(3) Evaluation of microbial activity of coal soil according to soil improver

By using cellulose fabric methods, evaluation of microbial activity after treatment of soil improver was carried out according to time course. This result showed in Fig. 5.

In the control group that the soil improver did not treated, cellulose was degraded little after 8 months from treatment, however, in the treatment group, 500cc treatment group (xl) and 1000 cc treatment group (x2) , cellulose was quickly degraded during more or less 40 days after treatment to a extent that it was difficult to identify the original shape of cellulose. The degradation extent of cellulose was proportional to the amount of treatment of soil improver.

(4) Confirmation of plant restoration effect in a mine- destructed coal soil

To investigate plant restoration effect by plant growth promotion and removal of hazardous heavy metal by treatment of soil improver, the variation of heavy metal quantity in eluted water sampled specific manner showed in Fig. 6. The concentration of hazardous heavy metal in eluted waters after 40 days and 190 days from planting were decreased as time progressed in all kinds of investigated heavy metals except Fe and Al. In the case of Fe and Al mainly derived from coal soil, the concentration of Fe and Al was slowly increased in the control group which was not treated soil improver as time progressed, while the concentration of Fe in soil improver treatment group significantly decreased. This shows phytoremediation effects by removal of heavy metals, planting selected trees and treatment of soil improver. The removal of heavy metal is thought to have close relation with tree' s growth promotion effects and their absorption promotion.

(Example 6) Method of treatment of soil improver on a surface soil in mine-destructed coal soil

With the soil improver, it was investigated the growth of Betula costata and Betula schmidtii Regel, promotion effects on soil microorganism activity and plant restoration by removal of heavy metals. One year of seedlings of tolerant tree species, Betula costata and Betula schmidtii Regel, provided by National Forest Research Institute were culture on the soil from coal soil, compost soil and common yellow earth prepared by composition ratio of Table 12. Group I is a experiment that it has separately upper layer having mixtures of compost soil and yellow earth and down layer having coal soil (separation) ; Group II is a experiment with mixed soil without upper layer and down layer (mixing) . After 6 months cultivation, tree growth and soil microorganism activity was investigated and nutrient element and heavy metals in eluted water was investigated.

<Table 12> Mixing ratio for manufacturing compost soil, yellow earth and coal soil to investigate tolerant tree growth

Compost Yellow Coal soil Sum (%) soil (%) earth (%) (%)

Sludge 0 0 50 50 100

Sludge 12. 5 12.5 37.5 50 100

Sludge 25 25 25 50 100

Sludge 37. 5 37.5 02.5 50 100

The results of compost soil effect obtained by using the seedlings of tolerant trees, Betula costata and Betula schmidtii Regel growing naturally in abandoned mine area were shown in Table 13. From the result of both planting method (a) separation method, which had separately upper layer comprising the mixture of compost soil and yellow earth, and down layer comprising coal soil; (b) mixing method which completely mixed compost soil and coal soil, Betula schmidtii Regel was more tolerant and adaptive than Betula costata overall. <Table 13> Growth analysis of tolerant tree species naturally growing in abandoned mine area grown on a coal soil added compost soil

In the aspects of growth rates, dried body weights and quantity of chlorophyll, compost soil treatment was better effect than without compost soil treatment regardless of methods of treatment and tree species. It is expected to facilitate plant growth by treating compost soil into coal soil. The 25% of addition rate of compost soil is highly effective regardless of treatment methods.

[industrial Applicability] The soil improver manufactured by adding white rot fungus into the base material comprising sludge and sawdust according to the present invention is a matured soil improver so that it is difficult to distinguish from upland soil in shape; a economical soil because it can be quickly manufactured with the help of added white rot fungus; and a high effective soil improver because it contains a effective composition of sludge and degraded material of sawdust .

By adding the soil improver according to present invention into the soil of abandoned mines, the plantation of the destructed soil of abandoned mine having low pH and nutrient shortage can be restored; the exposure of slope can be reduced with a sticky property of sludge; the leakage of heavy metals to the around soil of abandoned mines and rivers can be reduced due to its relatively high pH (6-8) compared with the pH of the soil of abandoned mine

(pH 2-4); and the heavy metal removal from the soil of abandoned mine can be achieved by promoting the absorption of heavy metals to plant. Also, the method of manufacturing soil improver can be applied to the sludge having 90% and more water content .

Claims

[CLAIMS]

[Claim l]

A soil improver comprising sludge, degraded sawdust and white rot fungus, wherein the amount of white rot fungus is more than 10⁷ cells per unit gram of the sludge and sawdust.

[Claim 2]

The soil improver according to claim 1, wherein the white rot fungus is one or more species selected from the group consisting of Trametes versicolor, Pleurotus ostreatus, Phanerochaete sordida, Trametes hirsutus, Fusarium culmorum, Phanerochaete chrysosporium and the deposited microorganism of deposition number KCCM 10725P.

[Claim 3] The microorganism of deposition number KCCM 10725P for facilitating composting process of a soil improver.

[Claim 4]

A method of manufacturing a soil improver by adding white rot fungus into a base material comprising sludge and sawdust.

[Claim 5]

The method of manufacturing a soil improver according to claim 4, wherein the white rot fungus is one or more species selected from the group consisting of Trametes versicolor, Pleurotus ostreatus, Phanerochaete sordida, Trametes hirsutus, Fusarium culmorum, Phanerochaete chrysosporium and the deposited microorganism of deposition number KCCM 10725P. [Claim β] The method of manufacturing a soil improver according to any one of claim 4 or 5, wherein a sludge fermenting bacteria is further added.

[Claim 7] A method of plant restoration by introducing the soil improver according to claim 1 into the soils of abandoned mines [Claim 8]

The method of plant restoration according to claim 7, wherein a surface coal soil in abandoned mines is cover with the soil improver by more than 25 % ratio by weight of the soil improver to the surface coal soil. [Claim 9]

A method of removing heavy metals comprising the steps of:

(a) introducing the soil improver according to claim 1 to the soil of abandoned mines; and

(b) planting a plant with heavy metal removing activity to the soil which is introduced with the soil improver. [Claim 10]

The method of removing heavy metals according to claim 9, wherein a surface coal soil in abandoned mines is cover with the soil improver by more than 25 % ratio by weight of the soil improver to the surface coal soil. [Claim 11]

The method of removing heavy metals according to any one of claim 9 or 10, wherein the plant with heavy metal removing activity is one selected from the group consisting of Betula schmidtii, Betula costata, Pinus Densiflora, Fraxinus rhynchophylla _r Fraxinus sieboldiana, Rhododendron mucromulatum, Weigela subsessilis, Acer pseudosieboldianum, Rhododendron schlippenbachii, Salix hulteni, Betula platyphylla, Salix koreensis, Alnus japonica, Stephanandra incisa, Mignolia sieboldii, Callicarpa japonica, Acer mono, Quercus mongolica, Corylus sieboldiana and Tilia amurensis.