JPH11179337A - Microbiological purification method and apparatus for contaminated soil and groundwater using buoyant granular carriers - Google Patents

Abstract

(57) [Summary] [PROBLEMS] When a microorganism that decomposes pollutant is introduced into an area contaminated with contaminant in soil and decomposed and purified, more efficient contact between the pollutant and the microorganism is achieved. The present invention provides a soil purification method and a device capable of obtaining a higher decomposition and purification effect by providing the same. SOLUTION: Microorganisms that decompose harmful substances are fixed to a floating particulate carrier, and the carrier is floated in a treatment area in soil that has been given an appropriate fluidity by the application of an aqueous medium, and the microorganisms are harmful to microorganisms. Improve contact efficiency with substances.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for purifying an area contaminated with harmful substances in soil using microorganisms, and a purification apparatus used for the method.

[0002]

2. Description of the Related Art As analytical techniques have progressed and chemical substance toxicity evaluation standards have been established, leakage caused by damage to storage devices and the like of chemical substances that have been used in large quantities, and environmental pollution due to human disposal have occurred. Is becoming a problem around the world. Above all, pollution of soil and groundwater by organic compounds such as gasoline and organic chlorine compounds such as PCB and trichlorethylene is remarkable, and there is a strong concern that ecosystems may be impaired through the use of contaminated well water. In addition, the number of contaminated sites will increase dramatically in future investigations, and it is thought that the number of harmful substances will increase with the increase in environmentally controlled substances in the future. It has been demanded.

[0003] Methods for purifying soil and groundwater contaminated with harmful substances can be broadly classified into physicochemical methods and microbiological methods. In physicochemical methods, various methods are selected depending on the harmful substances and the nature of the contamination site, but methods of excavating and heating or incineration of contaminated soil, methods of vacuum extraction from contaminated soil, aeration or adsorption by pumping contaminated groundwater Processing method, and the like. Heating and incineration methods can remove harmful substances almost completely from soil, but it is difficult to purify under buildings because soil excavation is required. Further, since it is often necessary to transport a large amount of excavated soil to the site of the purification treatment, the cost required for the purification treatment becomes enormous.

[0004] The vacuum extraction method is an inexpensive and simple purification method for volatile harmful substances. In addition, since harmful substances can be extracted not only from soil but also from groundwater, it can be said that this is an effective purification method for contaminated soil and groundwater. However, the removal efficiency is low for volatile substances of a few ppm or less, and it takes a year unit time to purify harmful substances such as trichlorethylene to the environmental standard value or less.

[0005] Furthermore, the pumping aeration method of pumping up and treating groundwater is an effective purification method as in the case of the vacuum extraction method. However, a large aeration tower or adsorption tank is installed for high-concentration pollution, which requires a large capital investment. is necessary. In addition, even if the pollution concentration decreases with purification, the initial equipment will operate as it is,
The maintenance cost becomes enormous.

On the other hand, microbiological methods using microorganisms have recently attracted attention as economical and environmentally friendly purification methods as compared with physicochemical methods. In particular, when microbiological purification is performed in situ at the site of contamination, there are two methods: using indigenous degrading microorganisms that naturally inhabit contaminated soil and groundwater, and inoculating microbes with degrading ability from outside. Can be In any case, harmful substances can be decomposed and purified in a short time by using microorganisms having high decomposition activity. Furthermore, purification can be performed by directly injecting degrading microorganisms and nutrients that enhance the degrading activity into soil or groundwater, so that it is also possible to purify under buildings without excavating contaminated soil.

In conventional microbiological purification, a method of adding nutrients, oxygen, inducers, or other chemicals to the soil or groundwater alone or together with the microorganisms in order to allow the microorganisms to survive and enhance the decomposition activity is used. Has been taken. In this method, an effective purification treatment is achieved by maximizing the decomposition activity of microorganisms and efficiently contacting harmful substances with microorganisms. In small-scale experiments in the laboratory, the level of microbial decomposition activity greatly controls the purification efficiency.However, in the case of purification using microorganisms on a large scale like an actual contamination site, the diffusion of harmful substances and microorganisms is usually slow, so Purification speed decreases.

[0008]

In order to purify contaminated soil or groundwater microbiologically in situ, it is preferable to activate decomposed microorganisms and bring the microorganisms into contact with harmful substances. However, it is difficult to almost completely decompose and remove harmful substances even if a large amount of extremely high concentration of degrading microorganisms is introduced into the contamination site (AG Duba et. Al, Environ.
i. Technol., 1996, 30, 1982-1989). In this case, the decomposition activity of the microorganism is extremely high, and it is considered that the cause is that the harmful substance and the microorganism are not in sufficient contact. And
According to the study of the present inventors, in order to carry out microbiological purification at the maximum efficiency, it is necessary to improve the contact efficiency by promoting the transfer of harmful substances or degrading microorganisms, and to devise so that the decomposition rate does not decrease. I got the conclusion. In conventional microbiological in-situ purification, a method has been proposed in which harmful substances are moved by a physical method to improve contact efficiency.
For example, US Pat. No. 5,347,070 describes a method in which a plurality of electrodes are inserted into contaminated soil, an electric current is applied to the soil to increase the temperature, and move harmful substances. Also, US Pat. No. 5,340,570 describes a method in which an electric field is applied to soil and harmful substances are moved by the generated electroosmotic flow. However, the effect on cost and input energy is relatively small, and a microbiological treatment method that increases the contact efficiency at low cost and low energy is desired.

From such a viewpoint, it is conceivable to promote the mobility of microorganisms or microbial particulate carriers injected into soil or groundwater. This is because, even if the transfer of harmful substances existing in soil or groundwater is difficult, the purification efficiency can be improved by increasing the mobility of microorganisms or microbial particulate carriers. In addition, if there is a transfer of harmful substances such as a groundwater flow, the purification treatment becomes more effective.
Enhancing the transfer of microorganisms themselves is a biological research subject, and requires the development of smaller, degraded microorganisms, flagellar microorganisms, and non-adherent microorganisms.

An object of the present invention is to provide a more efficient opportunity for contact between a contaminant and a microorganism when introducing a microorganism that decomposes the contaminant into an area contaminated with the contaminant in the soil and decomposing and purifying the microorganism. The present invention provides a soil purification method and a soil purification device that can provide a higher decomposition and purification effect by providing the same.

[0011]

According to the present invention, there is provided a soil purification treatment method for microbiologically purifying an area contaminated by harmful substances in soil. Fixing to a granular carrier; (b)
Filling a treatment area including the contaminated area with an aqueous medium; and (c) injecting a particulate carrier in which the microorganisms are immobilized into a lower part of the treatment area, and moving the particulate carrier upward from an injection part in the treatment area. And continuously purifying the inside of the treatment area with microorganisms fixed to the granular carrier by floating on the carrier.

The biological soil purification apparatus of the present invention, which can be suitably used in the above-mentioned soil purification method, comprises an aqueous medium for injecting an aqueous medium into a treatment area including an area contaminated by harmful substances in soil. It is characterized by comprising a supply means and a granular carrier injection means for injecting a buoyant granular carrier into the processing region.

In the present invention, a floating carrier is used as a carrier for immobilizing microorganisms that decompose pollutants, and an aqueous medium is filled in a treatment area in soil to ensure appropriate fluidity. Thus, the particulate carrier on which the microorganisms are immobilized is moved by the buoyancy in the treatment area, so that more chances of contact between the pollutants and the microorganisms are secured, and the decomposition efficiency of the pollutants can be improved.

There are several prior examples of particulate carriers on which microorganisms are immobilized. In Japanese Patent Application Laid-Open No. 8-89989, microorganisms are immobilized on a floating particulate carrier, which is introduced from the lower part of a wastewater treatment apparatus, and the upward movement of the apparatus is promoted by utilizing the floating property of the particulate carrier. . In this method, the movement of the particulate carrier causes the microorganisms to first contact the anaerobic zone in the apparatus and then to contact the aerobic zone. That is, in order to bring about an environmental change of microorganisms, mobility is imparted to the granular carrier, and the in-situ purification treatment at the contamination site using the mobility of the microorganism carrier is not intended. Further, Japanese Patent Application Laid-Open No. 8-197081
In Japanese Unexamined Patent Application Publication No. Hei 8-224588, the specific gravity is 0.
A method for biological treatment of sewage by immobilizing microorganisms on a granular carrier of 8 to 1.0 is disclosed. These methods utilize the floating properties of the microbial particulate carrier to prevent the granular carrier from flowing off in the flow direction of the sewage, and to improve the separation and recovery of the treated water and the particulate carrier. Therefore, it is not intended to improve the efficiency of the purification treatment using the floating properties of the granular carrier in the soil.

[0015]

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the present invention will be described in detail.
First, harmful substances gradually penetrate from the surface to the underground due to rainwater, etc., so that harmful substances in non-aquifers slowly diffuse in soil over time. Hazardous substances that reach the aquifer are then directly affected by groundwater flow, and especially soluble toxic substances move with the groundwater. The moving speed of groundwater is as wide as several cm / day to several m / day, but unlike harmful substances in non-aquifers, harmful substances in aquifers diffuse reliably over time. The purification treatment is performed by adding degrading microorganisms, nutrients for increasing the activity, and the like to the soil and groundwater contaminated in this way. Now 10pp
Assume that 5 × 10 ⁸ CFU / ml of degrading microorganisms are present in groundwater (assuming no soil particles) contaminated with m trichlorethylene. It consists of one degrading microorganism with a volume of about 2 fl in 2000 fl water and 9 × 1
This corresponds to the presence of ⁰⁹ trichloroethylene molecules. Trichlorethylene around the microorganism is quickly taken up by the microorganism and decomposed and purified. However, since the continuity of groundwater is interrupted by the soil grains, the diffusion of microorganisms and trichlorethylene in the soil becomes extremely slow. As a result, harmful substances at a distance cannot be brought into contact with microorganisms, so that the decomposition efficiency is reduced. On the other hand, under conditions that can promote the transfer and diffusion of microorganisms, carriers carrying microorganisms, and trichloroethylene, microorganisms can always come into contact with harmful substances, so that the decomposition rate is not reduced and purification efficiency can be improved.

First, microbial materials which can be used in the present invention and which decompose harmful substances include, for example, Saccharomyces, Hansenula, Candida and Micrococcus, which have been confirmed to have various decomposition activities. (Micrococcus), Staphylococcus (Staph)
ylococcus), Streptococcus,
Leuconostoa, Lactobacillus (Lac
tobacillus, Corynebacteriu
m), Arthrobacter, Bacillus (B
acillus), Clostridium, Neisseria, Escherichia, Enterobacter, Serrati
a), Achromobacter, Alcaligenes, Flavobact
erium), Acetobacter, Moraxella, Nitrosomonas,
Nitrobacter, Thiobacillus (Thiob
acillus), Gluconobacter, Pseudomonas, Xanthomonas
onas) and microorganisms belonging to each genus such as Vibrio. The microorganisms can be used alone or in combination of two or more. The degrading microorganism fixed to the granular carrier may be an indigenous microorganism originally inhabiting the contaminated soil or a foreign microorganism having a degrading ability, for example, a microorganism isolated from nature. Also,
Microorganisms that have been artificially mutated or genetically modified can also be used.

Preferred microorganisms include strains J1 and JM1 used in Examples described later. All of these strains have been deposited at the Institute of Biotechnology and Industrial Technology, Institute of Industrial Science and Technology, Ministry of International Trade and Industry, 1-3-3 Higashi, Tsukuba, Ibaraki (postal code 305), and the deposit number (B
P: International deposit under the Budapest Treaty) and the date of deposit are as follows. J1 strain: FERM BP-5102: May 25, 1994
Japan JM1 strain: FERM BP-5352: January 1, 1995
On day 0, the J1 strain and the JM1 strain initially belonged to the genus Corynebacterium, and were labeled "Corynebacterium sp. J."
1 "and" Corynebacterium sp.
JM1 ", but it was confirmed by subsequent examination that it did not belong to the genus Corynebacterium.
The identification display has been changed to “J1 strain” and “JM1 strain”. The JM1 strain is capable of decomposing aromatic compounds and chlorinated ethylene compounds by using an inducer.
A mutant strain obtained by mutating one strain by a mutation operation using a mutagen and capable of decomposing an organic compound without using an inducer.

In the present invention, the aqueous medium introduced into the soil ensures the appropriate fluidity of the soil in the decomposition treatment area including the area contaminated with the harmful substances in the soil, and converts the buoyant particulate carrier into the soil. It is used to move (float) effectively by buoyancy, and is mainly composed of water. An aqueous medium composed of water alone, a growth functional material necessary for the growth of microorganisms, and an activity maintaining functional material for stably expressing the decomposition activity of microorganisms, as required, can be used as long as the predetermined effect can be obtained. An aqueous medium obtained by dissolving or mixing in water can be used.

The growth functional material is a microbial nutrient,
As a result, the microorganisms multiply and survive in soil and groundwater, and decompose harmful substances in soil and groundwater. For example, broth medium, M9 medium, L medium, Malt Extract, MY medium, nitrifying bacteria selective medium, etc. are useful. When the degrading enzyme produced by the microorganism is constitutively expressed, no special activity-maintaining material is required, but when the enzyme activity is expressed by a specific inducer, the inducer is required as the activity-maintaining functional material. It is. For methane-utilizing bacteria, methane,
Inducers include toluene, phenol and cresol for aromatic assimilating bacteria, and ammonium salts for nitrifying bacteria. In addition, an energy source or a mineral for maintaining the activity of the degrading enzyme is also required as the activity maintaining functional material.

The composition and the amount of the aqueous medium to be introduced into the soil are set so that the floating of the floating particulate carrier and the decomposing action of the harmful substances of microorganisms attached or fixed to the particulate carrier can be effectively obtained. However, it is desirable to measure in advance the water content and nutritive value related to the growth and activation of the microorganisms in the soil in the treatment area, and to select in consideration of the measured value. For example, when the treatment area is a groundwater layer, the composition of the aqueous medium is selected with more emphasis on replenishment of nutrients such as microbial nutrients than on filling of soil with moisture.

The amount of the aqueous medium introduced into the soil is determined based on the volume of the soil in the treatment area, the porosity of the soil, and the water content of the soil. The amount of the medium may be calculated, and the soil void may be filled with a larger amount of the aqueous medium.

As the granular carrier to which the microorganisms are fixed, it is preferable to use a granular carrier having a density lower than that of water so as to obtain buoyancy by buoyancy in the treatment area in the soil into which the aqueous medium has been injected. Also, as the shape of the granular carrier,
It can be selected in consideration of the handling property, the applicability to the processing apparatus, and the mobility in the soil. The surface of the carrier may have appropriate irregularities as long as the mobility of the carrier in the soil is not hindered. Such irregularities can rather increase the contact area between the aqueous medium and the carrier and improve the purification effect.

In any case, the granular carrier has a function of providing a comfortable habitat for microorganisms, thereby preventing the predation by other microorganisms and micro-organisms, and preventing the diffusion and disappearance of microorganisms into groundwater. What you have is used.

As the particulate carrier, there can be used many particulate carriers for immobilizing microorganisms which have been used in bioreactors such as the pharmaceutical industry, the food industry, and wastewater treatment systems. For example, porous glass, ceramics, metal oxides, activated carbon, kaolinite, bentonite, zeolite, silica gel, alumina, particulate particulate carriers such as anthracite, starch, agar, chitin, chitosan, polyvinyl alcohol, alginic acid, polyacrylamide, Gelled granular carriers such as carrageenan, agarose and gelatin, cellulose, glutaraldehyde,
Polymer resins such as polyacrylic acid and urethane polymers, and ion exchange resins. Furthermore, natural or synthetic polymer compounds, for example, cotton containing cellulose as a main component,
Hemp, pulp material, or polymer acetate, polyester, polyurethane, or the like obtained by modifying a natural product is also effective.
A material having a lower density than the aqueous medium can be used as a granular carrier as it is, and a material having a higher density can be used after being made porous or hollow to reduce the apparent density. That is, by adjusting the density of the granular carrier, the floating speed can be controlled, and a more effective purification process can be performed according to the condition of the processing region. From a practical viewpoint, as the granular carrier, those having a density (including an apparent density) smaller than the density of water can be suitably used.
As described above, the density of the granular carrier to be used needs to be smaller than the density of water.However, if the density is close to the density of water, a sufficient floating effect may not be obtained. preferable. In addition, it is possible to produce a carrier having a very small apparent density by the hollowing treatment as described above. However, since the pores in the carrier become large, the pores are filled with the aqueous medium, and as a result, they float. The effect may not be obtained. Therefore, a preferable carrier density is 0.01 to 0.8 g / cm ³ , more preferably 0.05 to 0.8 g / cm ³ .
The purification is preferably carried out using a granular carrier of ０．５0.5 g / cm ³ .

The immobilization of the microorganisms on the granular carrier may be performed before the introduction into the soil, or the microorganisms may be immobilized in the soil by introducing only the granular carrier into the soil. In order to more reliably obtain a sufficient amount of microorganisms for purification, it is preferable to fix microorganisms for decomposition on a carrier before putting them into soil.

When the granular carrier is injected into the treatment area in the soil into which the aqueous medium has been injected, an upward force is generated by the buoyancy of the granular carrier. The particle size of the granular carrier is preferably smaller than the average diameter of the soil void, and is preferably about 0.01 mm to 1 mm in consideration of handling of the granular carrier and immobilization rate of microorganisms on the granular carrier. When the particle size of the granular carrier is sufficiently smaller than the soil void, the harmful substance comes into contact with microorganisms and is decomposed while gradually moving upward due to the buoyancy of the granular carrier. On the other hand, in many cases, the particle size of the granular carrier is larger than the soil void,
It becomes difficult for the particulate carrier to float due to the filter effect of the soil. In such a case, it is preferable to employ means for promoting the floating of the granular carrier.

For example, when a gas is injected after the injection of the granular carrier, a void for floating the granular carrier can be created by the buoyancy of the gas and the force of shearing the soil particles. The gas to be injected is air, oxygen, carbon dioxide, nitrogen,
A mixture of hydrogen, helium, neon, argon, carbon monoxide, methane, nitric oxide, nitrogen dioxide, sulfur dioxide, and the like mixed with air can be used.

It is also effective to mechanically vibrate the contaminated soil or groundwater area after injecting the granular carrier. The use of a high-output vibrator or the like makes it possible to vibrate the soil filled with the aqueous medium to secure a gap necessary for the granular carrier to float. Furthermore, when a surfactant such as sodium dodecyl sulfate or Triton-X is dissolved in the aqueous medium, the contact friction between the soil particles and the microbial particulate carrier can be reduced, and the floating property of the particulate carrier can be increased.

The particulate carrier purifies soil and groundwater while rising by buoyancy, and floats to near the ground surface to be collected. The microbial particulate carrier may be used again for the injection treatment as it is, or may be injected after activating the decomposition activity of the microorganism. Further, when the decomposition and removal of the harmful substance from the aqueous medium is completed, the aqueous medium can be taken out as purified water and treated.

In order to decompose harmful substances in soil and groundwater more efficiently, the concentration of harmful substances and oxygen, the concentration of the particulate carrier on which the degrading microorganisms are fixed, or the rising speed of the particulate carrier is measured to determine the amount of gas injected. It is desired to control the injection time, the vibration output, the vibration time, or their time intervals. The locations where these concentrations are measured include a non-aquifer or groundwater layer before filling with an aqueous medium, a purification area after filling with an aqueous medium, or near the ground surface. In addition, it is preferable to control the buoyancy of the particulate carrier so that the concentration of harmful substances and decomposition intermediate products is minimized. If harmful substances and decomposition products are volatile, their concentrations can be measured by gas chromatography or detector tubes. When they are non-volatile or non-volatile, soil can be sampled and pretreated, and measured by liquid chromatography, absorptiometer, or the like. Oxygen can be measured by gas chromatography, a detection tube or an oxygen sensor in the same manner as volatile volatile substances. The number of degraded microorganisms can be counted by a plate count or a flow cytometer after pretreatment of the sampled granular carrier. The ascent rate of the particulate carrier can be determined from the number of particulate carriers in the soil by sampling the soil in the depth direction at appropriate times. Is simple.

FIG. 1 shows an example of a purification apparatus according to the present invention. The soil in FIG. 1 includes a non-aquifer 1, an impermeable layer 2, and a contaminated area 3. There may be a groundwater layer below the non-aquifer 1 and above the impermeable layer 2. First,
The impermeable wall 4 is made of sheet pile or soil solidifying material so as to surround the contaminated area 3. Next, the tank 5 storing the aqueous medium
After that, the area surrounded by the impermeable wall by the pump 6 is filled with the aqueous medium. Further, a purifying material containing a particulate carrier having immobilized decomposed microorganisms is injected into the lower part of the contaminated area through an injection pipe 9 by a pump 8 from a tank 7. A single tube having an opening for press-fitting at the tip or side can be used as the injection tube. When it is desired to perform the injection operation repeatedly while changing the injection depth, a method of combining a manchette pipe 11 having a rubber sleeve 10 and a sleeve pipe 13 having a packer 12 is useful. That is, the sleeve pipe 13
To move the packer 1 up and down at a predetermined position.
Inflate 2 The purifying material is pumped through the sleeve pipe 13 to the portion between the upper and lower packers, and is pressed into the soil through the rubber sleeve 10.

When the soil void is sufficiently large, the injected microbial particulate carrier decomposes and purifies the contaminated area while rising by buoyancy. When the soil void is small, air is injected from a position below the granular carrier injection region by a pump 15 from the tank 14 storing the air. Thereby, upward movement of the granular carrier can be promoted. The microbial particulate carrier that has floated to near the surface of the ground is recovered by the pump 16 and returned to the tank 7 as it is or after activating the activity.

Each part constituting the purification treatment apparatus according to the present invention can be composed of commonly used members and devices.

[0034]

The present invention will be described below by way of examples, which do not limit the scope of the present invention in any way. “%” Is “% by weight”. Example 1 (Fixation of Degraded Microorganism J1 Strain on Floating Granular Carrier and Phenol Degradation) (1) First, strain J1 was applied to an agar plate medium containing 0.1% yeast extract added to M9 medium to form a colony. did. 8 liters of M9 medium containing 0.1% yeast extract was placed in a 10-liter culture tank, and the particle size was 0.1 m.
m and 8 g of a porous cellulose granular carrier (Visco Pearl, manufactured by Rengo) having a density of 0.1 g / cm ³ were mixed. The J1 colony that had grown therein was inoculated and stirred at 15 ° C. while aerating air to allow the microorganism to grow for about 2 days. A part of the cellulose granular carrier was taken out, placed in a new M9 medium, vigorously stirred and shaken to detach the adhered microorganisms, and the number of fixed microorganisms was determined by a plate count method. As a result, it was found that about 5 × 10 ⁹ J1 strains were fixed per 1 cm ^{3 of} the granular carrier. M9 medium composition (in 1 liter of medium): Na ₂ HPO ₄ : 6.2 g KH ₂ PO ₄ : 3.0 g NaCl: 0.5 g NH ₄ Cl: 1.0 g (pH 7.0) (2) Inner diameter 32 cm, high A 30 cm cylindrical stainless steel pot was filled with gravel having a water content of 5%. The weight of the gravel filled at this time was 43 kg, and the average pore diameter determined by eye was 0.1 kg.
7 mm. Further, an injection tube (diameter: 1 cm) for injecting a granular carrier immobilized with microorganisms was attached from the lower center of the stainless steel pot. The gravel was contaminated with phenol such that the phenol concentration in the gravel was 10 ppm with respect to the water in the gravel layer, and the test was performed. 6.8 ion-exchanged water in which 1.2% disodium hydrogen phosphate, 0.6% potassium dihydrogen phosphate, 0.4% ammonium chloride, and 0.1% sodium chloride are dissolved in the stainless steel pot
One liter (hereinafter, a buffer solution) was added. Next, 8 g of the granular carrier to which the strain J1 was fixed was mixed with 0.7 liter of the above buffer solution, and the mixture was injected from the lower part of the stainless steel pot through an injection tube. The water at the center of the pot immediately after the injection was sampled, and the phenol concentration was determined by the JIS method (JIS K012-
1993, 28.1). As a result, the phenol concentration was 1.3 ppm, and the calculated value was 1.4 ppm.
Almost matched.

(3) The granular carrier decomposes the phenol in the stainless steel pot while passing and rising through the gap of the gravel.
Two hours after the injection of the granular carrier, when a part of the granular carrier floated on the surface of the stainless steel pot, the water content in the center of the pot was sampled, and the phenol concentration was measured according to JIS.
Measured according to the method. As a result, the phenol concentration was 0.6
ppm, indicating that phenol can be efficiently decomposed even in the upper part of the injection region by using the floating particulate carrier.

Comparative Example 1 (Fixation of Degraded Microorganism J1 onto Non-Floating Granular Carrier and Degradation of Phenol) (1) In the same manner as in Example 1 (1), J1 was immobilized on the granular carrier. The granular carrier used had a particle size of 0.18 mm and a density of 1.03 g / cm ³ (microcarrier, manufactured by Asahi Kasei) 8
0 g. Example 1 (1) and was determined the number of microorganisms that have been fixed in the same manner, approximately 4 to particulate support 1 cm ³ per
It was found that × 10 ⁹ J1 strains were fixed.

(2) A gravel layer contaminated with phenol was prepared in the same manner as in Example 1 (2), and 6.8 liter of a buffer solution was added thereto. Granular carrier 8 with J1 strain immobilized
0 g was mixed with 0.7 liter of a buffer solution, and the mixture was injected from the lower part of the stainless steel pot through an injection tube. The water content at the center of the pot immediately after the injection was sampled, and the phenol concentration was measured according to the JIS method. As a result, the phenol concentration was 1.2 ppm with respect to the water in the gravel layer, which almost coincided with the calculated value of 1.4 ppm.

(3) Two hours after the injection of the granular carrier, water in the center of the pot was sampled, and the phenol concentration was adjusted to J.
It was measured according to the IS method. As a result, the phenol concentration was 1.2 ppm, and it was found that phenol could not be decomposed in the upper part of the injection region when the non-floating particulate carrier was used.

Example 2 (Fixation of Degraded Microorganism JM1 onto Floating Granular Carrier
(Chloroethylene degradation) (1) First, add 0.1% yeast extract to M9 medium
The JM1 strain was spread on the agar plate medium, and colonies were created.
Was. 0.1% yeast extract in a 10 liter culture tank
8 liters of M9 medium containing
m, density 0.1 g / cm^ThreePorous cellulose granular carrier
(Visco Pearl, made by Rengo) 8 g was mixed. In this
Inoculated with JM1 colonies grown in
The microorganisms were allowed to grow for about 2 days while stirring with stirring. cell
Take out part of the loin granular carrier and place it in a new M9 medium.
And vigorously shake to shake off the adhered microorganisms.
The number of fixed microorganisms was determined by the rate counting method.
As a result, the granular carrier 1 cm ^ThreeAbout 5 × 10 per⁹Individual
JM1 strain was found to be fixed.

(2) A cylindrical stainless steel pot having an inner diameter of 32 cm and a height of 30 cm was filled with gravel having a water content of 5%. At this time, the weight of the filled gravel was 43 kg, and the average pore diameter measured by eye was 0.7 mm. Further, an injection tube (diameter: 1 cm) for injecting a granular carrier immobilized with microorganisms was attached from the lower center of the stainless steel pot. The trichlorethylene concentration in the gravel is 50
The sample was contaminated with trichlorethylene so as to be ppm and used for the test. 6.8 liters of the buffer solution was placed in the stainless steel pot, and then 8 g of the granular carrier on which the JM1 strain was immobilized.
Was mixed with 0.7 liter of a buffer solution and injected from the lower part of the stainless steel pot through an injection tube. The water in the center of the pot immediately after the injection was sampled, and the concentration of trichlorethylene was measured by a solvent extraction method using n-hexane. As a result, the trichlorethylene concentration was 3.5 ppm
Which was approximately half the calculated value of 7 ppm assuming that trichlorethylene did not volatilize from the pot.
It was considered that about half the amount of trichlorethylene was dissipated from the pot by the pouring operation.

(3) The granular carrier decomposes trichlorethylene in the stainless steel pot while passing through and rising through the gaps in the gravel. Two hours after the injection of the granular carrier, when a part of the granular carrier came to the surface of the stainless steel pot, water in the center of the pot was sampled, and the concentration of trichlorethylene was measured by a solvent extraction method using n-hexane. As a result, the trichlorethylene concentration was 1.8 pp
m, and it was found that trichlorethylene can be efficiently decomposed even in the upper part of the injection region by using the floating particulate carrier.

Comparative Example 2 (Fixation of Degrading Microorganism JM1 on Non-Floating Granular Carrier and Degradation of Trichlorethylene) (1) In the same manner as in Example 2 (1), JM1 was added to the granular carrier.
The strain was fixed. The granular carrier used had a particle size of 0.18 mm and a density of 1.03 g / cm ³ (microcarrier, manufactured by Asahi Kasei).
80 g. The number of fixed microorganisms was determined in the same manner as in Example 2 (1).
10 ⁹ strain JM1 was found to be fixed.

(2) In the same manner as in Example 2 (2), a gravel layer contaminated with trichlorethylene was prepared, and 6.8 liter of a buffer solution was added thereto. 80 g of the granular carrier on which the JM1 strain was immobilized was mixed with 0.7 liter of a buffer solution,
This was injected from the lower part of the stainless steel pot through the injection tube. Sampling the water in the center of the pot immediately after the injection,
When the trichlorethylene concentration was measured by a solvent extraction method using n-hexane, the trichlorethylene concentration was 3.3 ppm.

(3) Two hours after the injection of the granular carrier, the water content in the center of the pot was sampled, and the trichloroethylene concentration was measured according to a solvent extraction method using n-hexane. As a result, the trichlorethylene concentration was 3.1 ppm
It was found that trichloroethylene could not be decomposed in the upper part of the injection region when a non-floating particulate carrier was used.

Example 3 (Phenol decomposition using a floating particulate carrier and air injection) (1) In the same manner as in Example 1 (1), the particle size was 0.1 mm.
The J1 strain was immobilized on 8 g of a porous cellulose granular carrier (Visco Pearl, manufactured by Rengo) having a density of 0.1 g / cm ³ .

(2) A cylindrical stainless steel pot having an inner diameter of 32 cm and a height of 30 cm was filled with sand having a water content of 10%. The weight of the sand at this time was 35 kg, and the average pore diameter measured by eye was 0.1 mm. Further, an injection tube (diameter: 1 cm) for injecting a granular carrier immobilized with microorganisms was attached from the lower center of the stainless steel pot. The sand was contaminated with phenol such that the phenol concentration in the sand became 10 ppm with respect to the water in the sand layer, and the sand was subjected to a test. 8.9 liters of the buffer solution was placed in the stainless steel pot, and then a mixture of 8 g of the granular carrier immobilized with the J1 strain and 0.7 liter of the buffer solution was injected from the lower part of the stainless steel pot through an injection tube. Water at the center of the pot immediately after the injection was sampled, and the phenol concentration was measured according to the JIS method. As a result, the phenol concentration was 1.9 ppm, and the calculated value 2.
It almost coincided with 0 ppm.

(3) Since the pore size of the sand is almost the same as that of the granular carrier, the granular carrier cannot float due to the filter effect of the sand. Therefore, air was injected from the injection pipe after the injection of the granular carrier to promote the floating of the granular carrier. Air injection is performed once an hour for 5 minutes at a flow rate of 100 ml / min. After 10 hours from the injection of the granular carrier, water at the center of the pot is sampled when a part of the granular carrier floats on the surface of the stainless steel pot. Then, the phenol concentration was measured according to the JIS method. As a result, the phenol concentration was 0.2 ppm, and it was found that phenol can be efficiently decomposed even in the upper part of the injection region by using both the floating particulate carrier and air injection.

Example 4 (Phenol decomposition using a floating particulate carrier and soil vibration) (1) In the same manner as in Example 1 (1), the particle size was 0.1 mm.
The J1 strain was immobilized on 8 g of a porous cellulose granular carrier (Visco Pearl, manufactured by Rengo) having a density of 0.1 g / cm ³ .

(2) In the same manner as in Example 3 (2), a sand layer contaminated with phenol was prepared, and a buffer solution and a granular carrier were injected. The water in the center of the pot immediately after the injection was sampled, and the phenol concentration was measured according to the JIS method. The phenol concentration was 1.8 ppm with respect to the water in the sand layer, which was almost the same as the calculated value of 2.0 ppm.

(3) Since the pore size of the sand is almost the same as that of the granular carrier, the granular carrier cannot float due to the filter effect of the sand. Therefore, the soil was vibrated after the injection of the granular carrier to promote the floating property of the granular carrier. Vibro (10
0 W) for 10 minutes once an hour. Eight hours after the injection of the granular carrier, when a part of the granular carrier floated on the surface of the stainless steel pot, water at the center of the pot was sampled, and the phenol concentration was measured according to the JIS method. As a result, the phenol concentration was 0.4 ppm, and it was found that phenol can be efficiently decomposed even in the upper part of the injection region by using the floating granular carrier and soil vibration together.

Comparative Example 3 (Phenol Decomposition Using Floating Granular Carrier) (1) In the same manner as in Example 3 (1),
The J1 strain was immobilized on 8 g of a porous cellulose granular carrier (Visco Pearl, manufactured by Rengo) having a density of 0.1 g / cm ³ .

(2) In the same manner as in Example 3 (2), a sand layer contaminated with phenol was prepared, and a buffer solution and a granular carrier were injected into the sand layer. The water in the center of the pot immediately after the injection was sampled, and the phenol concentration was measured according to the JIS method. The phenol concentration was 1.8 relative to the water in the sand layer.
ppm, which almost coincided with the calculated value of 2.0 ppm.

(3) Ten hours after the injection of the granular carrier, water at the center of the pot was sampled, and the phenol concentration was measured according to the JIS method. As a result, the phenol concentration was 1.7 ppm, and it was found that phenol in the upper part of the injection area could not be decomposed unless air injection or soil vibration was used together.

[0054]

According to the present invention, efficient microbiological purification of contaminated soil and groundwater has become possible.

[Brief description of the drawings]

FIG. 1 is a diagram showing an outline of a soil purification process in the present invention.

[Description of Signs] 1 Non-aquifer 2 Impermeable layer 3 Contaminated area 4 Impermeable wall 5, 7, 14 Tank 6, 8, 15, 16 Pump 9 Injection pipe 10 Rubber sleeve 11 Manchette pipe 12 Packer 13 Sleeve pipe

──────────────────────────────────────────────────の Continued on front page (51) Int.Cl. ⁶ Identification symbol FI C12N 1/00 C12N 1/00 B 1/20 1/20 DF 11/02 11/02 // (C12N 1/20 C12R 1 : 01)

Claims (24)

[Claims] 1. A method for microbiologically purifying an area contaminated by a harmful substance in soil, comprising: (a) fixing a microorganism capable of decomposing the harmful substance to a floating particulate carrier; (C) injecting a particulate carrier in which the microorganisms are immobilized into a lower portion of the treatment region, and injecting the particulate carrier upward from an injection portion in the treatment region. Continuously purifying the inside of the treatment area with microorganisms fixed to the granular carrier by flotation. 2. The treatment area includes a groundwater layer.
The soil purification treatment method according to item 1. 3. The soil purification treatment method according to claim 1, wherein the harmful substance is an organic compound. 4. The soil purification treatment method according to claim 3, wherein the organic compound is phenol. 5. The soil purification treatment method according to claim 3, wherein the harmful substance is an organic chlorine compound. 6. The soil purification method according to claim 5, wherein the organochlorine compound is trichloroethylene. 7. The microorganism according to claim 1, wherein the microorganism is a wild strain.
The soil purification method according to any one of the above. 8. The wild-type strain is a J1 strain (FERM BP-
5102). The soil purification treatment method according to claim 7, wherein 9. The soil treatment method according to claim 1, wherein the microorganism is a mutant obtained by mutating a wild strain. 10. The mutant strain is a JM1 strain (FERM B).
P-5352). 11. The soil purification treatment method according to claim 10, wherein the density of the granular carrier in the treatment area is lower than the density of water. 12. The particle size of the granular carrier is 0.01 mm.
The soil purification treatment method according to claim 10 or 11, wherein the distance is from 1 to 1 mm. 13. The soil purification treatment method according to claim 10, wherein the granular carrier comprises a polymer compound. 14. The soil purification treatment method according to claim 1, wherein the aqueous medium contains a material that exhibits or maintains the activity of decomposing the harmful substance by the microorganism. 15. The soil purification treatment method according to claim 1, wherein a gas is injected after injecting the particulate carrier into the treatment area to promote the movement of the particulate carrier by buoyancy. 16. The soil purification method according to claim 15, wherein the gas is air. 17. The method according to claim 1, wherein the processing target area is vibrated to promote movement of the granular carrier by buoyancy.
The soil purification method according to any one of the above. 18. The soil purification treatment method according to claim 17, wherein a vibro is used to vibrate the treatment area. 19. An aqueous medium supply means for injecting an aqueous medium into a treatment area including a toxic substance contaminated area in soil, and a particulate carrier injecting means for injecting buoyant particulate carriers into the treatment area. And a processing device for microbiological purification. 20. The processing apparatus according to claim 19, further comprising a movement promoting means for promoting movement of the granular carrier supplied into the processing area by buoyancy. 21. The processing apparatus according to claim 19, further comprising a granular carrier extracting means for collecting the granular carrier above the processing area. 22. The processing apparatus according to claim 19, further comprising a granular carrier moving unit that supplies the collected granular carrier to the granular carrier injection unit again. 23. The processing apparatus according to claim 19, further comprising a treated water extracting means for extracting an aqueous medium treated by the microorganism from within the treatment target area. 24. The processing apparatus according to claim 19, wherein microorganisms having an action of decomposing the harmful substance are fixed to the granular carrier.