JP2002210491A - Decomposing method for cyano compound and iron-cyano complex - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a technology by which a cyano compound, especially an iron-cyano complex can be anaerobically decomposed by a microorganism. SOLUTION: In the decomposing method for the cyano compound, the cyano compound, especially the iron-cyano complex is decomposed by applying bacterium belonging to the genus Klebsiella having the ability for decomposing the cyano compound, especially the iron-cyano complex to the object to be treated containing the cyano compound, especially the iron-cyano complex under the anaerobic condition.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

TECHNICAL FIELD The present invention relates to a method for biologically decomposing a cyanide compound, particularly an iron cyano complex.

[0002]

2. Description of the Related Art Cyanide compounds include a group of compounds called cyanide and metal cyano complexes. The cyanide is also called “free cyanide” and is represented by the general formula _An (CN) _x , where A is hydrogen (H), sodium (Na), potassium (K), ammonium (NH) ₄ ),
Calcium (Ca) is the most toxic form of cyanide. Further, the metal cyano complex,
A metal salt of hydrogen cyanide and a metal bonded to an excess of cyanide ion (CN ⁻ ), and represented by the general formula A _n [M (CN)
_x ] _y . Here, M is silver (Ag), gold (A
u), cadmium (Cd), cobalt (Co), copper (C
u), iron (Fe), nickel (Ni), zinc (Zn), and the like, which are dissolved or suspended in a solution.

[0003] The above-mentioned cyanide compound may be contained in industrial wastewater or the like, and is a substance having a property to be removed by a purification treatment. Regarding the form of the cyanide compound in the solution such as the industrial wastewater, much research has been conducted in the field of wastewater treatment, and it has been reported that the form is mostly the free cyanide and the metal cyano complex. I have. As a method of treating wastewater containing these cyanides,
Alkaline chlorine method, ozone oxidation method, electrolytic oxidation method, navy blue method (slightly soluble complex compound precipitation method), acid decomposition combustion method, boiling method (boiled high temperature combustion method), wet thermal decomposition method, adsorption method, etc. are known. ing. However, these physical and chemical cyanide treatment methods may not be applicable to highly stable metal cyano complexes, for example, iron, cobalt, silver, and gold cyano complexes. In addition, reaction conditions such as heating and pressurization are severe, large-scale equipment such as settling tanks and furnaces are required, and the products resulting from these treatments can be used for further secondary treatment. There was a problem in that it sometimes required.

Therefore, as a technique for purifying industrial wastewater and the like while avoiding the various problems described above, so-called bioremediation, which performs a purification treatment using a biological function, has been attracting attention. In removing E. coli, microorganisms having the ability to decompose E. coli were also searched.

At present, regarding microorganisms that decompose the above-mentioned free cyanide such as potassium cyanide and sodium cyanide, Pseudomonas putida, Pseudomonas sp. , Acinetobacter sp. Fusarium sp. And others were isolated and identified.

[0006] With regard to the metal cyano complex,
Nickel - as a decomposition microorganism cyanide complex _{([K 2 Ni (CN)} 4]), Fusarium solani (Fusarium
solani), Trichoderma polysporum (Tri)
chordum polysporum) has been reported. Fusarium oxysporum (Fusarium oxysporum) is a decomposing bacterium of potassium ferrocyanide ([K ₄ Fe (CN) ₆ ]) which is an iron cyano complex.
orum), Citalyum Thermofilum (Scy)
talium thermophilum), Penicillium michizinski (Penicillium m)
iczynski) have been reported (Kno
wles C.W. J. et. al. , Enzyme
and Microbiology Technology
22: 223-231, (1998)).

[0007]

However, all of the above-mentioned cases relating to the decomposition of cyanide compounds including iron cyano complexes are limited to those performed under aerobic conditions, and examples of decomposition under anaerobic conditions have been reported. It has not been.
Here, considering the treatment of the wastewater containing the cyanide, when performing the biological aerobic treatment on the industrial wastewater and the like, as is obvious from the fact that aeration is often performed actively, The industrial wastewater or the like that is simply left standing or flowing in the piping is in an anaerobic state.
Therefore, in order to decompose the cyanide using microorganisms under aerobic conditions, for practical use, facilities for supplying oxygen such as an aeration tank equipped with equipment for aeration are provided for practical use, and the cyanide-decomposition microorganisms are provided. On the other hand, there was a problem that oxygen had to be supplied. For this reason,
There is a problem that the facility for wastewater treatment becomes large and the running cost becomes high.

Further, when the cyan compound is decomposed and removed from the soil containing the cyan compound, it can be said that the soil is in an anaerobic environment except for the surface layer, so that the aerobic decomposition condition of the cyan compound is required. If the treatment is carried out using degrading microorganisms, it is difficult to treat it directly on site. Therefore, in order to remove cyanide using a cyanide-decomposing microorganism that requires aerobic reaction conditions, in order to make the soil environment aerobic, equipment for aeration or stirring the soil is used. There is a problem that it is necessary to provide a large-scale facility for this purpose, and to further require excessive costs for operating the facility. Alternatively, the soil is dug out, and the slurry or the supernatant obtained by mixing the aqueous solution and the dug out soil is stirred and supplied with oxygen while being brought into contact with the cyanide-decomposing microorganisms to perform the treatment. There is a problem that it is necessary to provide a large-scale facility and to operate it with an excessive cost.

Therefore, an object of the present invention is to provide a method for decomposing a cyanide compound by a microbiological technique under anaerobic conditions in view of the above-mentioned drawbacks.

[0010]

Means for Solving the Problems In order to solve the above-mentioned problems, the present inventors screened a microorganism capable of decomposing a cyanide compound even under anaerobic conditions, using the resolution of the cyanide compound as an index. As a result of intensive research, the present inventors have succeeded in separating a novel microorganism having the ability to decompose cyanide compounds even under anaerobic conditions using the atmosphere as a sampling source, and completed the present invention.

That is, the present invention provides a method for decomposing a cyanide compound even under an anaerobic condition.
This is a method of decomposing the cyan compound by causing a novel microorganism belonging to the genus lla) to act on an object to be treated containing the cyan compound. The present invention also provides a cyanide compound capable of decomposing the above-mentioned cyanide compound, which is allowed to act on the object to be treated containing the cyanide compound under aerobic conditions, and thereafter, the cyanide compound is allowed to act on the object to be treated under anaerobic conditions. Is a decomposition method. Specifically, the Klebsiella bacterium refers to Klebsiella pneumoniae (Klebsiella pn
eumoniae) strain AN-1. Further, the cyan compound is specifically a metal cyano complex, and more specifically, an iron cyano complex.

In the present specification, the term "anaerobic conditions" does not mean only strict anaerobic conditions in which the oxygen partial pressure is infinitely close to 0 (see Example 1), but uses microorganisms. It is a condition in which the oxygen partial pressure is reduced to such an extent that it is regarded as "anaerobic" in the field of waste liquid and soil treatment. In these fields, the general range of oxygen partial pressure when wastewater / soil is not subjected to oxygen supply treatment such as aeration / air injection is often referred to as anaerobic condition. In the case of combining the step of supplying oxygen and the step of not supplying oxygen, the step is not limited to the above range of the oxygen partial pressure, and aeration and
Conditions in which air injection or the like is not performed can be referred to as anaerobic conditions.

The present inventors have isolated a novel microorganism according to the present invention as follows. The sterilized ferrocyanide liquid medium placed in the flask was left in the air, and the microorganisms grown in this medium were inoculated on a sterile ferrocyanide agar plate medium, and then cultured at 30 ° C. to separate the formed colonies. . The composition and the preparation method of the ferrocyan agar medium and the ferrocyan liquid medium are as follows.

[0014]

[Table 1]

[0015]

[Table 2]

Further, a sterilized flask prepared by dispensing 100 ml of a sterilized ferrocyanide liquid medium in a flask is prepared, one platinum loop of the above isolated strain is inoculated, and the flask is sealed with a cotton plug so that gas can flow therethrough. Shaking culture (150 to 170 rpm) was performed at 30 ° C. for 1 week. After the cultivation, the ferrocyan (total cyan) content in the culture solution was determined to be "CN of low-quality soil".
Culture (culture solution, bacterial cells) according to the “Pre-operation method for distilling cyan (the method of heating and distilling all the cyanide in soil)” described in “Method for measuring content (Circular water pipe No. 127, paragraph 14.2)” The whole amount was distilled, and the obtained distillate was subjected to a fully automatic cyan measurement apparatus (T-CN501 manufactured by Anatech Yanaco) and the ferrocyanide (total cyan) content was measured and determined. As a result of selecting a strain capable of degrading ferrocyanin in the ferrocyanide liquid medium, a strain having ferrocyanide resolution was obtained, and this strain was named AN-1 strain. The decomposition rate of ferrocyanide was determined by the following equation.

Ferrocyanide decomposition rate = ferrocyan content after culture / ferrocyan content after aseptic shaking × 100
(%)

The AN-1 strain was smeared on an SCD agar plate and cultured at 30 ° C. for 1 day. After confirming that the culture was a pure culture, the taxonomy of the AN-1 strain having the following mycological properties was obtained. The above position is described as "Bergey's Manual of Systematic Bacteriology 1st Edition" (Berg
ery's Manual of Systemati
c Bacteriology 1st Edition
n (1984)) ". The AN
Table 3 shows the bacteriological properties of the -1 strain.

[0019]

[Table 3]

As a result, the AN-1 strain was obtained from Klebsiella pneum
oniae). Here, among known bacteria belonging to the genus Klebsiella, among other species, including Pneumonie species, those having the resolution of cyanide compounds such as iron cyano complexes have not been reported so far. Therefore, the AN-1 strain is a new strain that is distinguished from known strains. Klebsiella pneumoniae A
The strain determined to be the N-1 strain has been deposited with the National Institute of Bioscience and Biotechnology, National Institute of Industrial Science and Technology under the accession number FERM P-17792.

The composition of the SCD agar medium is shown in Table 4 below.
It is as shown in FIG.

[0022]

[Table 4]

The inventors of the present invention have determined the ability of the Klebsiella pneumoniae AN-1 strain, which has decomposed an iron cyano complex, which is a kind of cyanide under aerobic conditions, under anaerobic conditions as described above. As a result of the investigation, it was revealed that the iron cyano complex was decomposed even under anaerobic conditions (see Example 1).

The bacterium of the genus Klebsiella having the ability to decompose a cyanide compound thus obtained is allowed to act on a cyanide-containing object under anaerobic conditions to thereby treat the industrial wastewater and soil. On the other hand, biological treatment can be performed without actively supplying oxygen. Therefore, upon treatment of the cyanide,
Since there is no need to construct and maintain equipment for aeration such as aeration and stirring, the size of the facility can be reduced and running costs can be reduced.

In particular, when treating the cyanide compound in the soil, the soil can be treated in situ without having to dig the soil, which is very useful in terms of shortening the construction period and reducing costs. Further, since the cyan compound in the soil is known to be present as ferrocyan which is an iron cyanide complex, by allowing the Klebsiella bacterium which degrades the ferrocyan to act on the soil to be treated. , A high cyanide removing effect can be obtained.

Further, the inventors of the present invention apply the above-mentioned cyanide-decomposable bacteria of the genus Klebsiella to an object to be treated containing the above-mentioned cyanide compound under aerobic conditions. , The decomposition rate of the cyanide compound was increased as compared with the anaerobic or aerobic single treatment (see Examples 2 and 3). Thereby, by introducing anaerobic decomposition as a post-treatment of the aerobic decomposition of the cyanide by the Klebsiella bacteria, it is possible to increase the decomposition rate of the cyanide compound as compared with the case where the aerobic treatment is performed alone. it can.

The inventors of the present invention have also proposed that the Klebsiella pneumoniae strain AN-1 of the genus Klebsiella according to the present invention grows in a neutral to weakly alkaline pH range similarly to general bacteria. Heading,
It has been clarified that the iron cyano complex can be decomposed and grown under neutral to weakly alkaline growth conditions. Therefore, when using this strain for the treatment of wastewater containing cyanide,
The liquid property is made weakly alkaline, so that the hardly soluble iron cyano complex is dissolved in the wastewater, and can be efficiently decomposed. In addition, the present inventors have found that the Klebsiella pneumoniae AN-1 strain can increase the pH of a culture solution during growth, and the Klebsiella pneumoniae AN-1 strain itself is capable of increasing the iron cyano strain. The complex may be solubilized to create an environment suitable for the decomposition of the iron cyano complex. As described above, the Klebsiella genus bacteria, particularly Klebsiella pneumoniae AN-1 strain, which has a cyanide decomposability for a cyanide-containing treatment target under anaerobic conditions, are neutral to weakly alkaline p-type.
In the H range, a cyanide compound, particularly an iron cyano complex, can be decomposed, so it can be said that the microorganism is suitable for use in wastewater treatment by bioremediation.

Further, since the Klebsiella bacterium is a facultative anaerobic bacterium, it does not require strict control such as inert gas replacement, and can be easily stored and inoculated.

[0029]

EXAMPLES Hereinafter, experimental examples of iron cyano complex decomposition using the microorganism according to the present invention will be described.

Example 1 Klebsiella pneumoniae strain AN-1 (hereinafter referred to as AN-1 strain) was cultured under the following conditions, and the total cyan content in the medium was measured. The decomposition rate was determined from the content. The decomposition rate of the iron cyano complex was determined by the following equation.

Iron cyano complex decomposition rate = total cyan content after culture with addition of AN-1 strain / total cyan content after aseptic shaking × 100 (%)

Into a sterilized serum bottle, 40 ml of sterilized ferrocyanide liquid medium was dispensed, replaced with argon gas, and sealed. Further, in order to completely eliminate oxygen, predetermined amounts of sodium sulfide and cysteine were added, and the mixture was allowed to stand in the presence of oxygen until the color of resazurin, which had a pink color, disappeared.
Thereafter, one loopful of the AN-1 strain was anaerobically inoculated into the serum bottle containing the ferrocyan liquid anaerobic medium, and then cultured statically at 30 ° C. One week after the start of the cultivation, a part of the medium was fractionated, and the total cyan content in the medium was described in the above-mentioned "Method for measuring CN content in low-quality soil (Circular water pipe No. 127, paragraph 14.2)". The measurement was carried out according to the pre-operating method for distillation of cyan (a method for heating and distilling all the cyanide in Shiyan).

As a result, the concentration of the initially added ferrocyanide was 2
In contrast to 5 ppm, the ferrocyanin concentration one week after the start of the culture was reduced to 17.5 ppm. This week of anaerobic digestion resulted in about 30% degradation of ferrocyan.

Example 2 In a 300 ml Erlenmeyer flask, potassium ferrocyanide 10 mg / L, glucose 75 g
/ L, 100 ml of R2 medium containing 1 g / L of urea were added, and one loopful of the AN-1 strain was transplanted at 30 ° C and 170 ° C.
The cells were shake-cultured for 9 days under the condition of rpm (aerobic treatment). Thereafter, the gas in the flask was replaced with nitrogen and sealed, and the flask was left at 30 ° C. for 15 days with gentle stirring with a stirrer.
(Anaerobic treatment). After culturing for a predetermined period, the total amount of cyanide in the flask was measured. The results are shown in FIG. As shown in FIG. 1, after the ferrocyanide is decomposed by about 45% by aerobic treatment, by performing anaerobic treatment, the total decomposition rate becomes about 70%.
%. In the experiment that was carried out at the same time and the aerobic treatment was continued for 24 days, the ferrocyanide decomposition rate was only 45%. From the above results, it is found that the decomposition rate of the cyanide compound can be significantly improved by performing the aerobic treatment and then performing the anaerobic treatment as compared with the case where the aerobic treatment is performed alone. Was.

Example 3 100 ml of the R2 medium was added to a 300 ml Erlenmeyer flask, and one loop of the AN-1 strain was transplanted. After the gas in the flask was replaced with nitrogen, the flask was allowed to stand at 30 ° C. for 7 days with gentle stirring using a stirrer (anaerobic treatment). Then, after replacing the gas in the flask with air, the cells were cultured with shaking at 30 ° C. and 170 rpm for 9 days (aerobic treatment). After culturing for a predetermined period, the total amount of cyanide in the flask was measured. The results are shown in FIG. FIG.
As shown in Fig. 7, after the ferrocyane was decomposed by about 20% by the anaerobic treatment, the total decomposition rate reached about 25% by the aerobic treatment. However, since the ferrocyanide can be decomposed by about 45% when the aerobic treatment is performed during the same period, performing the aerobic treatment after performing the anaerobic treatment is compared with performing the aerobic treatment and then performing the anaerobic treatment. And it is not effective.

From the above examples, it has been clarified that when decomposing a cyanide compound using the AN-1 strain, it is effective to treat under aerobic conditions and then under anaerobic conditions. .

As described above, the Klebsiella pneumoniae AN-1 strain is capable of decomposing an iron cyano complex under anaerobic conditions. It is suitable as a strain to be used. Therefore, the Klebsiella pneumoniae AN
By using the -1 strain, it is possible to efficiently decompose the iron cyano complex in the wastewater.

Further, considering the decomposition of the cyanide compound in the soil, the cyanide compound in the soil is known to exist as ferrocyan which is an iron cyanide complex. To
It is effective if the ferrocyanide can be decomposed by acting on the soil to be treated. From this viewpoint, it is considered that the Klebsiella pneumoniae AN-1 strain having the ferrocyanide-degrading property is suitable for purifying soil containing a cyanide compound. At this time, since the cyanide decomposing treatment by the Klebsiella pneumoniae AN-1 strain is performed in an environment in which microorganisms grow, severe degradation of temperature and temperature, as seen in other physical and chemical decomposition methods.
It does not need to be performed under pressure conditions. Further, since it is possible to decompose the cyanide under both anaerobic and aerobic conditions, the cyanide can be decomposed without excavating the soil or sending air into the soil to supply oxygen. The compound can be decomposed, for example, the soil to be treated is treated with the Klebsiella pneumoniae AN.
It is also possible to add -1 strain and perform the treatment on the spot.

[Brief description of the drawings]

FIG. 1 is a graph showing an embodiment of the present invention in which an anaerobic process is performed after an aerobic process.

FIG. 2 is a graph showing an example of the present invention in which aerobic treatment is performed after anaerobic treatment.

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Claims (6)

[Claims] 1. A method for decomposing an iron cyano complex, wherein a Klebsiella bacterium having the ability to degrade an iron cyano complex is allowed to act on an object to be treated containing an iron cyano complex under anaerobic conditions to decompose the iron cyano complex. 2. Under an anaerobic condition, an object to be treated containing an iron cyano complex is added to a Klebsiella
A method for decomposing an iron cyano complex, wherein the iron cyano complex is decomposed by acting on pneumonie. 3. An object to be treated containing a cyanide under an anaerobic condition is treated with a Klebsiella.
Pneumoniae AN-1 strain (FERM P-1779)
A cyan compound decomposing method of decomposing the cyan compound by applying 2). 4. An aerobic treatment step in which a Klebsiella bacterium having iron cyano complex decomposability is allowed to act on an object to be treated containing an iron cyano complex under aerobic conditions, wherein the aerobic treatment step is performed. A method for decomposing an iron cyano complex, comprising an anaerobic treatment step of allowing the aforementioned Klebsiella bacteria to act on an object to be treated under anaerobic conditions. 5. A method for treating a subject containing an iron cyano complex under aerobic conditions with a Klebsiella having an iron cyano complex decomposability.
An iron cyano complex decomposing method, comprising: an aerobic treatment step in which pneumonia acts; and an anaerobic treatment step in which the Klebsiella pneumoniae acts on the object to be treated that has undergone the aerobic treatment step under anaerobic conditions. 6. An aerobic condition in which an object to be treated containing a cyanide compound is treated with a Klebsiella.
Pneumoniae AN-1 strain (FERM P-1779)
An anaerobic treatment step in which the Klebsiella pneumoniae AN-1 strain (FERM P-17792) is allowed to act on the object to be treated that has been subjected to the aerobic treatment step under anaerobic conditions. A cyan compound decomposing method having a treatment step.